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Sample records for polarized madin-darby canine

  1. Sorting of membrane and fluid at the apical pole of polarized Madin-Darby canine kidney cells.

    PubMed

    Leung, S M; Ruiz, W G; Apodaca, G

    2000-06-01

    When fluid-phase markers are internalized from opposite poles of polarized Madin-Darby canine kidney cells, they accumulate in distinct apical and basolateral early endosomes before meeting in late endosomes. Recent evidence suggests that significant mixing of apically and basolaterally internalized membrane proteins occurs in specialized apical endosomal compartments, including the common recycling endosome and the apical recycling endosome (ARE). The relationship between these latter compartments and the fluid-labeled apical early endosome is unknown at present. We report that when the apical recycling marker, membrane-bound immunoglobulin A (a ligand for the polymeric immunoglobulin receptor), and fluid-phase dextran are cointernalized from the apical poles of Madin-Darby canine kidney cells, they enter a shared apical early endosome (

  2. Sorting of Membrane and Fluid at the Apical Pole of Polarized Madin-Darby Canine Kidney Cells

    PubMed Central

    Leung, Som-Ming; Ruiz, Wily G.; Apodaca, Gerard

    2000-01-01

    When fluid-phase markers are internalized from opposite poles of polarized Madin-Darby canine kidney cells, they accumulate in distinct apical and basolateral early endosomes before meeting in late endosomes. Recent evidence suggests that significant mixing of apically and basolaterally internalized membrane proteins occurs in specialized apical endosomal compartments, including the common recycling endosome and the apical recycling endosome (ARE). The relationship between these latter compartments and the fluid-labeled apical early endosome is unknown at present. We report that when the apical recycling marker, membrane-bound immunoglobulin A (a ligand for the polymeric immunoglobulin receptor), and fluid-phase dextran are cointernalized from the apical poles of Madin-Darby canine kidney cells, they enter a shared apical early endosome (≤2.5 min at 37°C) and are then rapidly segregated from one another. The dextran remains in the large supranuclear EEA1-positive early endosomes while recycling polymeric immunoglobulin receptor–bound immunoglobulin A is delivered to a Rab11-positive subapical recycling compartment. This latter step requires an intact microtubule cytoskeleton. Receptor-bound transferrin, a marker of the basolateral recycling pathway, has limited access to the fluid-rich apical early endosome but is excluded from the subapical elements of the Rab11-positive recycling compartment. We propose that the term ARE be used to describe the subapical Rab11-positive compartment and that the ARE is distinct from both the transferrin-rich common recycling endosome and the fluid-rich apical early endosome. PMID:10848634

  3. Toxin sensitivity of the calcium-dependent rubidium efflux in Madin-Darby canine kidney cells.

    PubMed

    Tauc, M; Gastineau, M; Poujeol, P

    1993-01-29

    86Rb+ efflux was measured on polarized Madin-Darby canine kidney cells under A23187 or ATP stimulation. This efflux, inhibited by barium, Leiurus quinquestriatus hebraeus venom and charybdotoxin was attributed to the stimulation of Ca(++)-activated maxi K+ channels. Snake venom from Dendroaspis polylepis did not alter the stimulation as well as did apamine. ATP was effective on both the apical and basolateral membranes and the Ca(++)-activated maxi K+ channels were predominantly found on the basolateral membrane. This study presents the physiological evidence that dendrotoxin is ineffective on the epithelial Ca(++)-activated maxi K+ channel present in MDCK cells. PMID:7678959

  4. Microtubule-acting drugs lead to the nonpolarized delivery of the influenza hemagglutinin to the cell surface of polarized Madin-Darby canine kidney cells.

    PubMed

    Rindler, M J; Ivanov, I E; Sabatini, D D

    1987-02-01

    The synchronized directed transfer of the envelope glycoproteins of the influenza and vesicular stomatitis viruses from the Golgi apparatus to the apical and basolateral surfaces, respectively, of polarized Madin-Darby canine kidney (MDCK) cells can be achieved using temperature-sensitive mutant viruses and appropriate temperature shift protocols (Rindler, M. J., I. E. Ivanov, H. Plesken, and D. D. Sabatini, 1985, J. Cell Biol., 100:136-151). The microtubule-depolymerizing agents colchicine and nocodazole, as well as the microtubule assembly-promoting drug taxol, were found to interfere with the normal polarized delivery and exclusive segregation of hemagglutinin (HA) to the apical surface but not with the delivery and initial accumulation of G on the basolateral surface. Immunofluorescence analysis of permeabilized monolayers of influenza-infected MDCK cells treated with the microtubule-acting drugs demonstrated the presence of substantial amounts of HA protein on both the apical and basolateral surfaces. Moreover, in cells infected with the wild-type influenza virus, particles budded from both surfaces. Viral counts in electron micrographs showed that approximately 40% of the released viral particles accumulated in the intercellular spaces or were trapped between the cell and monolayer and the collagen support as compared to less than 1% on the basolateral surface of untreated infected cells. The effect of the microtubule inhibitors was not a result of a rapid redistribution of glycoprotein molecules initially delivered to the apical surface since a redistribution was not observed when the inhibitors were added to the cells after the HA was permitted to reach the apical surface at the permissive temperature and the synthesis of new HA was inhibited with cycloheximide. The altered segregation of the HA protein that occurs may result from the dispersal of the Golgi apparatus induced by the inhibitors or from the disruption of putative microtubules containing tracks

  5. Characterization of hormone-sensitive Madin-Darby canine kidney cells

    SciTech Connect

    Lin, M.C.; Beckner, S.K.; Darfler, F.J.

    1985-01-01

    The paper describes the optimal conditions for maintaining hormone responsiveness, measurement of intracellular AMP, and the characteristics of several types of hormone sensitivity in Madin-Darby canine kidney cells. Cyclic AMP is measured by radioimmunoassay with (/sup 175/I) as tracer. The responsiveness of the kidney cells to glucagon, vasopressin, isoproterenol, and prostaglandin in presented.

  6. EphrinA1-EphA2 Signal Induces Compaction and Polarization of Madin-Darby Canine Kidney Cells by Inactivating Ezrin through Negative Regulation of RhoA*

    PubMed Central

    Wakayama, Yuki; Miura, Koichi; Sabe, Hisataka; Mochizuki, Naoki

    2011-01-01

    The epithelial cells exhibit either a columnar or a flat shape dependent on extracellular stimuli or the cell-cell adhesion. Membrane-anchored ephrinA stimulates EphA receptor tyrosine kinases as a ligand in a cell-cell contact-dependent manner. The mechanism through which ephrinA1/EphA2 signal regulates the cell morphology remains elusive. We demonstrate here that ephrinA1/EphA2 signal induces compaction and enhanced polarization (columnar change) of Madin-Darby canine kidney epithelial cells by regulating Ezrin, a linker that connects plasma membrane and actin cytoskeleton. Activation of EphA2 resulted in RhoA inactivation through p190RhoGAP-A and subsequent dephosphorylation of Ezrin on Thr-567 phosphorylated by Rho kinase. Consistently, the cells expressing an active mutant of Ezrin in which Thr-567 was replaced with Asp did not change their shape in response to ephrinA1. Furthermore, depletion of Ezrin led to compaction and enhanced polarization without ephrinA1 stimulation, suggesting the role for active Ezrin in keeping the flat cell shape. Ezrin localized to apical domain irrespective of ephrinA1 stimulation, whereas phosphorylated Ezrin on the apical domain was reduced by ephrinA1 stimulation. Collectively, ephrinA1/EphA2 signal negatively regulates Ezrin and promotes the alteration of cell shape, from flat to columnar shape. PMID:21979959

  7. The Recycling Endosome of Madin-Darby Canine Kidney Cells Is a Mildly Acidic Compartment Rich in Raft Components

    PubMed Central

    Gagescu, Raluca; Demaurex, Nicolas; Parton, Robert G.; Hunziker, Walter; Huber, Lukas A.; Gruenberg, Jean

    2000-01-01

    We present a biochemical and morphological characterization of recycling endosomes containing the transferrin receptor in the epithelial Madin-Darby canine kidney cell line. We find that recycling endosomes are enriched in molecules known to regulate transferrin recycling but lack proteins involved in early endosome membrane dynamics, indicating that recycling endosomes are distinct from conventional early endosomes. We also find that recycling endosomes are less acidic than early endosomes because they lack a functional vacuolar ATPase. Furthermore, we show that recycling endosomes can be reached by apically internalized tracers, confirming that the apical endocytic pathway intersects the transferrin pathway. Strikingly, recycling endosomes are enriched in the raft lipids sphingomyelin and cholesterol as well as in the raft-associated proteins caveolin-1 and flotillin-1. These observations may suggest that a lipid-based sorting mechanism operates along the Madin-Darby canine kidney recycling pathway, contributing to the maintenance of cell polarity. Altogether, our data indicate that recycling endosomes and early endosomes differ functionally and biochemically and thus that different molecular mechanisms regulate protein sorting and membrane traffic at each step of the receptor recycling pathway. PMID:10930469

  8. Toxin pharmacology of the ATP-induced hyperpolarization in Madin-Darby canine kidney cells.

    PubMed

    Tauc, M; Gastineau, M; Poujeol, P

    1992-03-23

    The effects of Leiurus quinquestriatus hebraeus (LQH) venom, mamba venom, Buthus tamulus (BT) venom, purified apamin and synthetic charybdotoxin on the membrane hyperpolarization induced by extracellular ATP were examined in Madin-Darby canine kidney cells. For this we used a membrane potential probe (bisoxonol) to determine the potential variations. The relation between bisoxonal fluorescence and membrane potential was established by treating Madin-Darby canine kidney cells suspended in solutions containing various external sodium concentrations with gramicidin. Extracellular ATP induced a rapid hyperpolarization that was blocked by LQH venom and synthetic charybdotoxin. BT venom also blocked the response but at a much higher concentration than that of LQH. Mamba venom (Dendroaspis polylepis) and apamin did not modify the ATP-induced hyperpolarization. We concluded that the ATP induced hyperpolarization was due to the augmentation of the potassium conductance probably through Ca(2+)-activated K+ channels sensitive to charybdotoxin but not to mamba venom. The interaction previously described between charybdotoxin and dendrotoxin (the main toxin of mamba venom) was not observed in our case. PMID:1373656

  9. Protein 4.1N is required for translocation of inositol 1,4,5-trisphosphate receptor type 1 to the basolateral membrane domain in polarized Madin-Darby canine kidney cells.

    PubMed

    Zhang, Songbai; Mizutani, Akihiro; Hisatsune, Chihiro; Higo, Takayasu; Bannai, Hiroko; Nakayama, Tomohiro; Hattori, Mitsuharu; Mikoshiba, Katsuhiko

    2003-02-01

    Protein 4.1N was identified as a binding molecule for the C-terminal cytoplasmic tail of inositol 1,4,5-trisphosphate receptor type 1 (IP(3)R1) using a yeast two-hybrid system. 4.1N and IP(3)R1 associate in both subconfluent and confluent Madin-Darby canine kidney (MDCK) cells, a well studied tight polarized epithelial cell line. In subconfluent MDCK cells, 4.1N is distributed in the cytoplasm and the nucleus; IP(3)R1 is localized in the cytoplasm. In confluent MDCK cells, both 4.1N and IP(3)R1 are predominantly translocated to the basolateral membrane domain, whereas 4.1R, the prototypical homologue of 4.1N, is localized at the tight junctions (Mattagajasingh, S. N., Huang, S. C., Hartenstein, J. S., and Benz, E. J., Jr. (2000) J. Biol. Chem. 275, 30573-30585), and other endoplasmic reticulum marker proteins are still present in the cytoplasm. Moreover, the 4.1N-binding region of IP(3)R1 is necessary and sufficient for the localization of IP(3)R1 at the basolateral membrane domain. A fragment of the IP(3)R1-binding region of 4.1N blocks the localization of co-expressed IP(3)R1 at the basolateral membrane domain. These data indicate that 4.1N is required for IP(3)R1 translocation to the basolateral membrane domain in polarized MDCK cells. PMID:12444087

  10. Transepithelial transport of aliphatic carboxylic acids studied in Madin Darby canine kidney (MDCK) cell monolayers

    SciTech Connect

    Cho, M.J.; Adson, A.; Kezdy, F.J. )

    1990-04-01

    Transport of 14C-labeled acetic, propionic (PA), butyric, valeric, heptanoic (HA), and octanoic (OA) acids across the Madin Darby canine kidney (MDCK) epithelial cell monolayer grown on a porous polycarbonate membrane was studied in Hanks' balanced salt solution (HBSS) at 37{degrees}C in both apical-to-basolateral and basolateral-to-apical directions. At micromolar concentrations of solutes, metabolic decomposition was significant as evidenced by (14C)CO2 production during the OA transport. The apparent permeability (Pe) indicates that as lipophilicity increases, diffusion across the unstirred boundary layer becomes rate limiting. In support of this notion, transport of OA and HA was enhanced by agitation, showed an activation energy of 3.7 kcal/mol for OA, and resulted in identical Pe values for both transport directions. Analysis of Pe changes with varying alkyl chain length resulted in a delta G of -0.68 +/- 0.09 kcal/mol for -CH2-group transfer from an aqueous phase to the MDCK cells. When the intercellular tight junctions were opened by the divalent chelator EGTA in Ca2+/Mg2(+)-free HBSS, transport of the fluid-phase marker Lucifer yellow greatly increased because of paracellular leakage. PA transport also showed a significant increase, but OA transport was independent of EGTA. Although albumin also undergoes paracellular transport in the presence of EGTA and OA binds strongly to albumin, OA transport in EGTA solution was unchanged by albumin. These observations indicate that transmembrane transport is the major mechanism for lipophilic substances. The present study, together with earlier work on the transport of polar substances, shows that the MDCK cell monolayer is an excellent model of the transepithelial transport barrier.

  11. Three-dimensional imaging of cholesterol and sphingolipids within a Madin-Darby canine kidney cell

    DOE PAGESBeta

    Yeager, Ashley N.; Weber, Peter K.; Kraft, Mary L.

    2016-01-08

    Metabolic stable isotope incorporation and secondary ion mass spectrometry(SIMS) depth profiling performed on a Cameca NanoSIMS 50 were used to image the 18O-cholesterol and 15N-sphingolipid distributions within a portion of a Madin-Darby canine kidney (MDCK) cell. Three-dimensional representations of the component-specific isotope distributions show clearly defined regions of 18O-cholesterol and 15N-sphingolipid enrichment that seem to be separate subcellular compartments. Furthermore, the low levels of nitrogen-containing secondary ions detected at the 18O-enriched regions suggest that these 18O-cholesterol-rich structures may be lipiddroplets, which have a core consisting of cholesterol esters and triacylglycerides.

  12. Three-dimensional imaging of cholesterol and sphingolipids within a Madin-Darby canine kidney cell.

    PubMed

    Yeager, Ashley N; Weber, Peter K; Kraft, Mary L

    2016-06-01

    Metabolic stable isotope incorporation and secondary ion mass spectrometry (SIMS) depth profiling performed on a Cameca NanoSIMS 50 were used to image the (18)O-cholesterol and (15)N-sphingolipid distributions within a portion of a Madin-Darby canine kidney (MDCK) cell. Three-dimensional representations of the component-specific isotope distributions show clearly defined regions of (18)O-cholesterol and (15)N-sphingolipid enrichment that seem to be separate subcellular compartments. The low levels of nitrogen-containing secondary ions detected at the (18)O-enriched regions suggest that these (18)O-cholesterol-rich structures may be lipid droplets, which have a core consisting of cholesterol esters and triacylglycerides. PMID:26746168

  13. Iterative sorting of apical and basolateral cargo in Madin-Darby canine kidney cells.

    PubMed

    Treyer, Aleksandr; Pujato, Mario; Pechuan, Ximo; Müsch, Anne

    2016-07-15

    For several decades, the trans-Golgi network (TGN) was considered the most distal stop and hence the ultimate protein-sorting station for distinct apical and basolateral transport carriers that reach their respective surface domains in the direct trafficking pathway. However, recent reports of apical and basolateral cargoes traversing post-Golgi compartments accessible to endocytic ligands before their arrival at the cell surface and the post-TGN breakup of large pleomorphic membrane fragments that exit the Golgi region toward the surface raised the possibility that compartments distal to the TGN mediate or contribute to biosynthetic sorting. Here we describe the development of a novel assay that quantitatively distinguishes different cargo pairs by their degree of colocalization at the TGN and by the evolution of colocalization during their TGN-to-surface transport. Keys to the high resolution of our approach are 1) conversion of perinuclear organelle clustering into a two-dimensional microsomal spread and 2) identification of TGN and post-TGN cargo without the need for a TGN marker that universally cosegregates with all cargo. Using our assay, we provide the first evidence that apical NTRp75 and basolateral VSVG in Madin-Darby canine kidney cells still undergo progressive sorting after they exit the TGN toward the cell surface. PMID:27226480

  14. Neutrophil-mediated transport of liposomes across the Madin Darby canine kidney epithelial cell monolayer.

    PubMed

    Cho, M J; Scieszka, J F; Cramer, C T; Thompson, D P; Vidmar, T J

    1989-01-01

    Targeted drug delivery to peripheral blood neutrophils (PMNs) should be of therapeutic potential in various disease states. In addition, substances taken up by PMNs in the circulation may be delivered to an extravascular site via the naturally occurring cell infiltration. The present study employs an in vitro chemotaxis model to test whether particulate drug carriers such as liposomes can be transported across a cellular barrier by migrating PMNs. The system contained 10(7) human PMNs/ml, 0.3-micron liposomes at a total lipid concentration of 2.5 mM, and 10% autologous human serum in the apical side of a confluent Madin Darby canine kidney (MDCK) epithelial cell monolayer of 4.71 cm2. The MDCK cells were grown on a polycarbonate membrane with 3-micron pores without any extracellular matrix, and 10(-7) M f-Met-Leu-Phe was added to the basolateral side as a trigger of chemotaxis. The aqueous phase of the reverse-phase evaporation vesicles (REVs) contained lucifer yellow CH (LY) and [14C]sucrose. The lipid bilayer of the REVs was spiked with [3H]dipalmitoylphosphatidylcholine (DPPC). Transmission electron micrographs showed that, in response to the formyl peptide, PMNs adhered to the apical surface of MDCK cells, emigrated across the MDCK cell layer, passed through the 3-micron pores in the polycarbonate membrane, and finally, appeared in the bottom well. Epifluorescence micrographs showed that most, if not all, of the migrated PMNs contained punctate fluorescence derived from LY. Transport data over a 3.5-hr period indicated that those markers that appeared in the basal side were indeed transported by phagocytosis of REVs by PMNs and that intact serum was an essential component in the process.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:2717523

  15. Characterization of a Madin-Darby canine kidney cell line stably expressing TRPV5.

    PubMed

    den Dekker, Els; Schoeber, Joost; Topala, Catalin N; van de Graaf, Stan F J; Hoenderop, Joost G J; Bindels, René J M

    2005-07-01

    To provide a cell model for studying specifically the regulation of Ca2+ entry by the epithelial calcium channel transient receptor potential-vanilloid-5 (TRPV5), green fluorescent protein (GFP)-tagged TRPV5 was expressed stably in Madin-Darby canine kidney type I (MDCK) cells. The localization of GFP-TRPV5 in this cell line showed an intracellular granular distribution. Ca2+ uptake in GFP-TRPV5-MDCK cells cultured on plastic supports was threefold higher than in non-transfected cells. Moreover, apical Ca2+ uptake in GFP-TRPV5-MDCK cells cultured on permeable supports was eightfold higher than basolateral Ca2+ uptake, indicating that GFP-TRPV5 is expressed predominantly in the apical membrane. Patch-clamp analysis showed the presence of typical electrophysiological features of GFP-TRPV5, such as inwardly rectifying currents, inhibition by divalent cations and Ca2+-dependent inactivation. Moreover, the TRPV5 inhibitor ruthenium red completely inhibited Ca2+ uptake in GFP-TRPV5-MDCK cells, whereas Ca2+ uptake in non-transfected cells was not inhibited. The characterized GFP-TRPV5-MDCK cell line was used to assess the regulation of TRPV5. The protein kinase C activator phorbol 12-myristate 13-acetate and the cAMP-elevating compounds forskolin/3-isobutyl-1-methylxanthine, 8-Br-cAMP and PGE2 stimulated TRPV5 activity in GFP-TRPV5-MDCK cells by 121+/-7, 79+/-5, 55+/-4 and 61+/-7%, respectively. These compounds did not affect Ca2+ uptake in non-transfected cells. In conclusion, the GFP-TRPV5-MDCK cell line provides a model to specifically study the regulation of TRPV5 activity. PMID:15924239

  16. Apical sorting of a voltage- and Ca2+-activated K+ channel alpha -subunit in Madin-Darby canine kidney cells is independent of N-glycosylation.

    PubMed

    Bravo-Zehnder, M; Orio, P; Norambuena, A; Wallner, M; Meera, P; Toro, L; Latorre, R; González, A

    2000-11-21

    The voltage- and Ca(2+)-activated K(+) (K(V,Ca)) channel is expressed in a variety of polarized epithelial cells seemingly displaying a tissue-dependent apical-to-basolateral regionalization, as revealed by electrophysiology. Using domain-specific biotinylation and immunofluorescence we show that the human channel K(V,Ca) alpha-subunit (human Slowpoke channel, hSlo) is predominantly found in the apical plasma membrane domain of permanently transfected Madin-Darby canine kidney cells. Both the wild-type and a mutant hSlo protein lacking its only potential N-glycosylation site were efficiently transported to the cell surface and concentrated in the apical domain even when they were overexpressed to levels 200- to 300-fold higher than the density of intrinsic Slo channels. Furthermore, tunicamycin treatment did not prevent apical segregation of hSlo, indicating that endogenous glycosylated proteins (e.g., K(V,Ca) beta-subunits) were not required. hSlo seems to display properties for lipid-raft targeting, as judged by its buoyant distribution in sucrose gradients after extraction with either detergent or sodium carbonate. The evidence indicates that the hSlo protein possesses intrinsic information for transport to the apical cell surface through a mechanism that may involve association with lipid rafts and that is independent of glycosylation of the channel itself or an associated protein. Thus, this particular polytopic model protein shows that glycosylation-independent apical pathways exist for endogenous membrane proteins in Madin-Darby canine kidney cells. PMID:11069304

  17. Apical sorting of a voltage- and Ca2+-activated K+ channel α-subunit in Madin-Darby canine kidney cells is independent of N-glycosylation

    PubMed Central

    Bravo-Zehnder, Marcela; Orio, Patricio; Norambuena, Andrés; Wallner, Martin; Meera, Pratap; Toro, Ligia; Latorre, Ramón; González, Alfonso

    2000-01-01

    The voltage- and Ca2+-activated K+ (KV,Ca) channel is expressed in a variety of polarized epithelial cells seemingly displaying a tissue-dependent apical-to-basolateral regionalization, as revealed by electrophysiology. Using domain-specific biotinylation and immunofluorescence we show that the human channel KV,Ca α-subunit (human Slowpoke channel, hSlo) is predominantly found in the apical plasma membrane domain of permanently transfected Madin-Darby canine kidney cells. Both the wild-type and a mutant hSlo protein lacking its only potential N-glycosylation site were efficiently transported to the cell surface and concentrated in the apical domain even when they were overexpressed to levels 200- to 300-fold higher than the density of intrinsic Slo channels. Furthermore, tunicamycin treatment did not prevent apical segregation of hSlo, indicating that endogenous glycosylated proteins (e.g., KV,Ca β-subunits) were not required. hSlo seems to display properties for lipid-raft targeting, as judged by its buoyant distribution in sucrose gradients after extraction with either detergent or sodium carbonate. The evidence indicates that the hSlo protein possesses intrinsic information for transport to the apical cell surface through a mechanism that may involve association with lipid rafts and that is independent of glycosylation of the channel itself or an associated protein. Thus, this particular polytopic model protein shows that glycosylation-independent apical pathways exist for endogenous membrane proteins in Madin-Darby canine kidney cells. PMID:11069304

  18. Sphingomyelin regulates the transbilayer movement of diacylglycerol in the plasma membrane of Madin-Darby canine kidney cells.

    PubMed

    Ueda, Yoshibumi; Makino, Asami; Murase-Tamada, Kotono; Sakai, Shota; Inaba, Takehiko; Hullin-Matsuda, Françoise; Kobayashi, Toshihide

    2013-08-01

    Diacylglycerol (DAG) is a key component in lipid metabolism and signaling. Previous model membrane studies using DAG analogs suggest their rapid membrane transbilayer movement. However, little is known about the DAG distribution and dynamics in cell membranes. Using live-cell fluorescence microscopy, we monitored the transbilayer movement of DAG with the yellow fluorescent protein-tagged C1AB domain from protein kinase C-γ (EYFP-C1AB), which selectively binds DAG. When HeLa cells were treated with Bacillus cereus phospholipase C (Bc-PLC) to produce DAG on the outer leaflet of the plasma membrane, intracellularly expressed EYFP-C1AB probe accumulated at the plasma membrane, indicating the transbilayer movement of the outer leaflet DAG to the inner leaflet. This Bc-PLC-induced translocation of EYFP-C1AB probe to the plasma membrane was not observed in the sphingolipid-enriched plasma membrane of Madin-Darby canine kidney cells, but was recovered after cell treatment with sphingomyelinase or preincubation with an inhibitor of sphingolipid biosynthesis. The inhibitory effect of sphingomyelin (SM) on the transbilayer movement of DAG was reproduced in model membranes using a fluorescent short-chain DAG analog. These results demonstrate that the SM content on the outer leaflet regulates the transbilayer movement of DAG in the plasma membrane, thus providing new insights into the dynamics of DAG in cell pathophysiology. PMID:23682124

  19. Analysis of transcriptomic and proteomic profiles demonstrates improved Madin-Darby canine kidney cell function in a renal microfluidic biochip.

    PubMed

    Snouber, Leila Choucha; Letourneur, Franck; Chafey, Philippe; Broussard, Cedric; Monge, Matthieu; Legallais, Cécile; Leclerc, Eric

    2012-01-01

    We have evaluated the influence of the microfluidic environment on renal cell functionality. For that purpose, we performed a time lapse transcriptomic and proteomic analysis in which we compared gene and protein expressions of Madin-Darby canine kidney cells after 24 h and 96 h of culture in both microfluidic biochips and plates. The transcriptomic and proteomic integration revealed that the ion transporters involved in calcium, phosphate, and sodium homoeostasis and several genes involved in H(+) transporters and pH regulation were up-regulated in microfluidic biochips. Concerning drug metabolism, we found Phase I (CYP P450), Phase II enzymes (GST), various multidrug resistance genes (MRP), and Phase III transporters (SLC) were also up-regulated in the biochips. Furthermore, the study shows that those inductions were correlated with the induction of the Ahr and Nrf-2 dependent pathways, which results in a global cytoprotective response induced by the microenvironment. However, there was no apoptosis situation or cell death in the biochips. Microfluidic biochips may thus provide an important insight into exploring xenobiotic injury and transport modifications in this type of bioartificial microfluidic kidney. Finally, the investigation demonstrated that combining the transcriptomic and proteomic analyses obtained from a cell "on chip" culture would provide a pertinent new tool in the mechanistic interpretation of cellular mechanisms for predicting kidney cell toxicity and renal clearance in vitro. PMID:22095740

  20. Synergistic antioxidant activity of resveratrol with genistein in high-glucose treated Madin-Darby canine kidney epithelial cells

    PubMed Central

    CHU, CHISHIH; LU, FUNG-JOU; YEH, RANG-HUI; LI, ZIH-LING; CHEN, CHING-HSEIN

    2016-01-01

    Resveratrol (Re), a stilbenoid, is associated with a potential benefit in controlling certain biomarkers in type II diabetes. Genistein (Ge), a phytoestrogen, may act as an antioxidant and thus may diminish damaging effects of free radicals in tissues. In the present study, a potential synergistic antioxidant effect of an Re/Ge combination on high-glucose (HG) incubation in Madin-Darby canine kidney (MDCK) epithelial cells was evaluated. Compared with the treatment of Re or Ge alone, the Re/Ge combination synergistically decreased intracellular reactive oxygen species (ROS) and hydroxyl radicals in MDCK cells. This synergistic antioxidant effect correlated with the inhibition of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase expression and an increase in γ-glutamylcysteine synthetase expression. In addition, mitochondrial complex I, NADPH oxidase, xanthine oxidase and lipoxygenase contributed towards ROS overproduction when the MDCK cells were incubated with HG. In conclusion, the Re/Ge combination synergistically enhanced the antioxidant effect in HG-incubated kidney cells, possibly through an enhanced antioxidant regulation mechanism. The Re/Ge combination may be a potential benefit against oxidative stress in diabetes mellitus. PMID:26998274

  1. Failure-to-thrive syndrome associated with tumor formation by Madin-Darby canine kidney cells in newborn nude mice.

    PubMed

    Brinster, Lauren R; Omeir, Romelda L; Foseh, Gideon S; Macauley, Juliete N; Snoy, Philip J; Beren, Joel J; Teferedegne, Belete; Peden, Keith; Lewis, Andrew M

    2013-08-01

    Tumors that formed in newborn nude mice that were inoculated with 10(7) Madin-Darby canine kidney (MDCK) cells were associated with a failure-to-thrive (FTT) syndrome consisting of growth retardation, lethargy, weakness, and dehydration. Scoliosis developed in 41% of affected pups. Pups were symptomatic by week 2; severely affected pups became moribund and required euthanasia within 3 to 4 wk. Mice with FTT were classified into categories of mild, moderate, and severe disease by comparing their weight with that of age-matched normal nude mice. The MDCK-induced tumors were adenocarcinomas that invaded adjacent muscle, connective tissue, and bone; 6 of the 26 pups examined had lung metastases. The induction of FTT did not correlate with cell-line aggressiveness as estimated by histopathology or the efficiency of tumor formation (tumor-forming dose 50% endpoint range = 10(2.8) to 10(7.5)); however, tumor invasion of the paravertebral muscles likely contributed to the scoliosis noted. In contrast to the effect of MDCK cells, tumor formation observed in newborn mice inoculated with highly tumorigenic, human-tumor-derived cell lines was not associated with FTT development. We suggest that tumor formation and FTT are characteristics of these MDCK cell inocula and that FTT represents a new syndrome that may be similar to the cachexia that develops in humans with cancer or other diseases. PMID:24209967

  2. Depletion of E-Cadherin Disrupts Establishment but Not Maintenance of Cell Junctions in Madin-Darby Canine Kidney Epithelial Cells

    PubMed Central

    Capaldo, Christopher T.

    2007-01-01

    E-cadherin forms calcium-dependent homophilic intercellular adhesions between epithelial cells. These contacts regulate multiple aspects of cell behavior, including the organization of intercellular tight junctions (TJs). To distinguish between the roles of E-cadherin in formation versus maintenance of junctions, Madin-Darby canine kidney (MDCK) cells were depleted of E-cadherin by RNA interference. Surprisingly, reducing E-cadherin expression had little effect on the protein levels or localization of adherens junction (AJ) or TJ markers. The cells underwent morphological changes, as the normally flat apical surface swelled into a dome. However, apical–basal polarity was not compromised, transmembrane resistance was normal, and zonula occludin protein 1 dynamics at the TJs were unchanged. Additionally, an E-cadherin/Cadherin-6 double knockdown also failed to disrupt established TJs, although β-catenin was lost from the cell cortex. Nevertheless, cells depleted of E-cadherin failed to properly reestablish cell polarity after junction disassembly. Recovery of cell–cell adhesion, transepithelial resistance, and the localization of TJ and AJ markers were all delayed. In contrast, depletion of α-catenin caused long-term disruption of junctions. These results indicate that E-cadherin and Cadherin-6 function as a scaffold for the construction of polarized structures, and they become largely dispensable in mature junctions, whereas α-catenin is essential for the maintenance of functional junctions. PMID:17093058

  3. Effects of cranberry extract on prevention of urinary tract infection in dogs and on adhesion of Escherichia coli to Madin-Darby canine kidney cells.

    PubMed

    Chou, Hsin-I; Chen, Kuan-Sheng; Wang, Hsien-Chi; Lee, Wei-Ming

    2016-04-01

    OBJECTIVE To determine effects of cranberry extract on development of urinary tract infection (UTI) in dogs and on adherence of Escherichia coli to Madin-Darby canine kidney (MDCK) cells. ANIMALS 12 client-owned dogs (in vivo experiment) and 6 client-owned dogs (in vitro experiment). PROCEDURES 12 dogs with a history of recurrent UTI received an antimicrobial (n = 6) or cranberry extract (6) orally for 6 months. Dogs were monitored for a UTI. For the in vitro experiment, cranberry extract was orally administered to 6 dogs for 60 days. Voided urine samples were collected from each dog before and 30 and 60 days after onset of extract administration. Urine was evaluated by use of a bacteriostasis assay. An antiadhesion assay and microscopic examination were used to determine inhibition of bacterial adherence to MDCK cells. RESULTS None of the 12 dogs developed a UTI. The bacteriostasis assay revealed no zone of inhibition for any urine samples. Bacterial adhesion was significantly reduced after culture with urine samples obtained at 30 and 60 days, compared with results for urine samples obtained before extract administration. Microscopic examination revealed that bacterial adherence to MDCK cells was significantly reduced after culture with urine samples obtained at 30 and 60 days, compared with results after culture with urine samples obtained before extract administration. CONCLUSIONS AND CLINICAL RELEVANCE Oral administration of cranberry extract prevented development of a UTI and prevented E coli adherence to MDCK cells, which may indicate it has benefit for preventing UTIs in dogs. PMID:27027843

  4. ToF-SIMS and laser-SNMS analysis of Madin-Darby canine kidney II cells with silver nanoparticles using an argon cluster ion beam.

    PubMed

    Nees, Ricarda; Pelster, Andreas; Körsgen, Martin; Jungnickel, Harald; Luch, Andreas; Galla, Hans-Joachim; Arlinghaus, Heinrich F

    2016-06-01

    The use of nanoparticles is one of the fastest expanding fields in industrial as well as in medical applications, owing to their remarkable characteristics. Silver nanoparticles (AgNPs) are among the most-commercialized nanoparticles because of their antibacterial effects. Laser postionization secondary neutral mass spectrometry (laser-SNMS) and time-of-flight secondary ion mass spectrometry in combination with argon cluster ion sputtering was used for the first time to investigate the effects of AgNPs on Madin-Darby canine kidney (MDCK) II cells. Depth profiles and high-resolution three dimensional (3D) images of nanoparticles and organic compounds from cells were obtained using an Ar cluster ion beam for sputtering and Bi3 (+) primary ions for the analysis. The 3D distribution of AgNPs and other organic compounds in MDCK II cells could be readily detected with very high efficiency, sensitivity, and submicron lateral resolution. The argon cluster ion beam is well suited for the sputtering of biological samples. It enables a high sample removal rate along with low molecular degradation. The outer membrane, the cytoplasm, and the nuclei of the cells could be clearly visualized using the signals PO(+) and C3H8N(+) or CN(+) and C3H8N(+). The laser-SNMS images showed unambiguously that AgNPs are incorporated by MDCK II cells and often form silver aggregates with a diameter of a few micrometers, mainly close to the outside of the cell nuclei. PMID:26671480

  5. Expression Profile of Drug and Nutrient Absorption Related Genes in Madin-Darby Canine Kidney (MDCK) Cells Grown under Differentiation Conditions

    PubMed Central

    Quan, Yong; Jin, Yisheng; Faria, Teresa N.; Tilford, Charles A.; He, Aiqing; Wall, Doris A.; Smith, Ronald L.; Vig, Balvinder S.

    2012-01-01

    The expression levels of genes involved in drug and nutrient absorption were evaluated in the Madin-Darby Canine Kidney (MDCK) in vitro drug absorption model. MDCK cells were grown on plastic surfaces (for 3 days) or on Transwell® membranes (for 3, 5, 7, and 9 days). The expression profile of genes including ABC transporters, SLC transporters, and cytochrome P450 (CYP) enzymes was determined using the Affymetrix® Canine GeneChip®. Expression of genes whose probe sets passed a stringent confirmation process was examined. Expression of a few transporter (MDR1, PEPT1 and PEPT2) genes in MDCK cells was confirmed by RT-PCR. The overall gene expression profile was strongly influenced by the type of support the cells were grown on. After 3 days of growth, expression of 28% of the genes was statistically different (1.5-fold cutoff, p < 0.05) between the cells grown on plastic and Transwell® membranes. When cells were differentiated on Transwell® membranes, large changes in gene expression profile were observed during the early stages, which then stabilized after 5–7 days. Only a small number of genes encoding drug absorption related SLC, ABC, and CYP were detected in MDCK cells, and most of them exhibited low hybridization signals. Results from this study provide valuable reference information on endogenous gene expression in MDCK cells that could assist in design of drug-transporter and/or drug-enzyme interaction studies, and help interpret the contributions of various transporters and metabolic enzymes in studies with MDCK cells. PMID:24300234

  6. Oncogenic H-Ras Reprograms Madin-Darby Canine Kidney (MDCK) Cell-derived Exosomal Proteins Following Epithelial-Mesenchymal Transition*

    PubMed Central

    Tauro, Bow J.; Mathias, Rommel A.; Greening, David W.; Gopal, Shashi K.; Ji, Hong; Kapp, Eugene A.; Coleman, Bradley M.; Hill, Andrew F.; Kusebauch, Ulrike; Hallows, Janice L.; Shteynberg, David; Moritz, Robert L.; Zhu, Hong-Jian; Simpson, Richard J.

    2013-01-01

    Epithelial-mesenchymal transition (EMT) is a highly conserved morphogenic process defined by the loss of epithelial characteristics and the acquisition of a mesenchymal phenotype. EMT is associated with increased aggressiveness, invasiveness, and metastatic potential in carcinoma cells. To assess the contribution of extracellular vesicles following EMT, we conducted a proteomic analysis of exosomes released from Madin-Darby canine kidney (MDCK) cells, and MDCK cells transformed with oncogenic H-Ras (21D1 cells). Exosomes are 40–100 nm membranous vesicles originating from the inward budding of late endosomes and multivesicular bodies and are released from cells on fusion of multivesicular bodies with the plasma membrane. Exosomes from MDCK cells (MDCK-Exos) and 21D1 cells (21D1-Exos) were purified from cell culture media using density gradient centrifugation (OptiPrep™), and protein content identified by GeLC-MS/MS proteomic profiling. Both MDCK- and 21D1-Exos populations were morphologically similar by cryo-electron microscopy and contained stereotypical exosome marker proteins such as TSG101, Alix, and CD63. In this study we show that the expression levels of typical EMT hallmark proteins seen in whole cells correlate with those observed in MDCK- and 21D1-Exos, i.e. reduction of characteristic inhibitor of angiogenesis, thrombospondin-1, and epithelial markers E-cadherin, and EpCAM, with a concomitant up-regulation of mesenchymal makers such as vimentin. Further, we reveal that 21D1-Exos are enriched with several proteases (e.g. MMP-1, -14, -19, ADAM-10, and ADAMTS1), and integrins (e.g. ITGB1, ITGA3, and ITGA6) that have been recently implicated in regulating the tumor microenvironment to promote metastatic progression. A salient finding of this study was the unique presence of key transcriptional regulators (e.g. the master transcriptional regulator YBX1) and core splicing complex components (e.g. SF3B1, SF3B3, and SFRS1) in mesenchymal 21D1-Exos. Taken

  7. Effects of oxalate exposure on Madin-Darby canine kidney cells in culture: renal prothrombin fragment-1 mRNA expression.

    PubMed

    Moryama, Manabu T; Domiki, Chizue; Miyazawa, Katsuhito; Tanaka, Tatsuro; Suzuki, Koji

    2005-12-01

    It has been suggested that renal tubular cell damage induced by oxalic acid, one of the components of urinary calculi, may be involved in a variety of ways in the development of urolithiasis. During our study on a calculus related protein, renal prothrombin fragment-1 (RPTF-1), we noted that this is an inflammation related substance that mediates an acute inflammatory reaction, one of the original roles of prothrombin. RPTF-1 is a part of prothrombin that is a coagulation factor known to be expressed in the renal tubule. We examined whether oxalic acid may cause cytotoxic effects on tubular epithelial cells and whether such chemical stimulation may promote the translation of RPTF-1 mRNA into RPTF-1 proteins. We used Madin-Darby canine kidney (MDCK) cells derived from the distal tubule of a dog kidney. In this study, the effects of oxalic acid in culture solution at different concentrations on cytotoxicity were assessed using a MTT assay. The location of active oxygen species was identified using dichlorofluorescein diacetate. After the prothrombin sequence of RPTF-1 was confirmed in MDCK cells, RPTF-1 mRNA expression was determined by RT-PCR. The gene sequence of the same promoter area was ligated, and a luciferase sequence was inserted downstream of the vector. The target sequence was transfected into MDCK cells and the relation between oxalic acid and prothrombin promoter was examined. In addition, the variable expression of RPTF-1 mRNA was quantitatively compared depending on oxalic acid concentrations using real-time PCR. When cytotoxicity was investigated, cells were not damaged but, by contrast, were stimulated and activated under oxalic acid below a certain concentration. The relation between cytotoxicity on the cultured MDCK cell membrane and active oxygen species was confirmed. Luminescence in MDCK cells containing the luciferase gene was detected by the addition of oxalic acid, which activated the prothrombin promoter. A part of the prothrombin gene

  8. Oncogenic H-ras reprograms Madin-Darby canine kidney (MDCK) cell-derived exosomal proteins following epithelial-mesenchymal transition.

    PubMed

    Tauro, Bow J; Mathias, Rommel A; Greening, David W; Gopal, Shashi K; Ji, Hong; Kapp, Eugene A; Coleman, Bradley M; Hill, Andrew F; Kusebauch, Ulrike; Hallows, Janice L; Shteynberg, David; Moritz, Robert L; Zhu, Hong-Jian; Simpson, Richard J

    2013-08-01

    Epithelial-mesenchymal transition (EMT) is a highly conserved morphogenic process defined by the loss of epithelial characteristics and the acquisition of a mesenchymal phenotype. EMT is associated with increased aggressiveness, invasiveness, and metastatic potential in carcinoma cells. To assess the contribution of extracellular vesicles following EMT, we conducted a proteomic analysis of exosomes released from Madin-Darby canine kidney (MDCK) cells, and MDCK cells transformed with oncogenic H-Ras (21D1 cells). Exosomes are 40-100 nm membranous vesicles originating from the inward budding of late endosomes and multivesicular bodies and are released from cells on fusion of multivesicular bodies with the plasma membrane. Exosomes from MDCK cells (MDCK-Exos) and 21D1 cells (21D1-Exos) were purified from cell culture media using density gradient centrifugation (OptiPrep™), and protein content identified by GeLC-MS/MS proteomic profiling. Both MDCK- and 21D1-Exos populations were morphologically similar by cryo-electron microscopy and contained stereotypical exosome marker proteins such as TSG101, Alix, and CD63. In this study we show that the expression levels of typical EMT hallmark proteins seen in whole cells correlate with those observed in MDCK- and 21D1-Exos, i.e. reduction of characteristic inhibitor of angiogenesis, thrombospondin-1, and epithelial markers E-cadherin, and EpCAM, with a concomitant up-regulation of mesenchymal makers such as vimentin. Further, we reveal that 21D1-Exos are enriched with several proteases (e.g. MMP-1, -14, -19, ADAM-10, and ADAMTS1), and integrins (e.g. ITGB1, ITGA3, and ITGA6) that have been recently implicated in regulating the tumor microenvironment to promote metastatic progression. A salient finding of this study was the unique presence of key transcriptional regulators (e.g. the master transcriptional regulator YBX1) and core splicing complex components (e.g. SF3B1, SF3B3, and SFRS1) in mesenchymal 21D1-Exos. Taken

  9. Apical expression of human full-length hCEACAM1-4L protein renders the Madin Darby Canine Kidney cells responsive to lipopolysaccharide leading to TLR4-dependent Erk1/2 and p38 MAPK signalling.

    PubMed

    Liévin-Le Moal, Vanessa; Beau, Isabelle; Rougeaux, Clémence; Kansau, Imad; Fabrega, Sylvie; Brice, Cédric; Korotkova, Natalia; Moseley, Steve L; Servin, Alain L

    2011-05-01

    CEACAM1 expressed by granulocytes and epithelial cells is recognized as a membrane-associated receptor by some Gram-negative pathogens. Here we report a previously unsuspected role of human CEACAM1-4L (hCEACAM1-4L) in polarized epithelial cells. We find that in contrast with non-transfected cells, Madin Darby Canine Kidney strain II (MDCK) engineered for the apical expression of the long cytoplasmic chain protein hCEACAM1-4L showed a serum-independent increase in the phosphorylation of the extracellular signal-regulated kinase 1/2 (Erk1/2) and p38 mitogen-activated protein kinases (MAPKs) after treatment with lipopolysaccharide (LPS) of wild-type, diffusely adhering Afa/Dr Escherichia coli (Afa/Dr DAEC) strain IH11128. Aggregates of FITC-LPS bind the apical domain of MDCK-hCEACAM1-4L cells colocalizing with the apically expressed hCEACAM1-4L protein and do not bind MDCK-pCEP cells, and surface plasmon resonance analysis shows that LPS binds to the extracellular domain of the CEACAM1-4L protein. We showed that cell polarization and lipid rafts positively control the LPS-IH11128-induced phosphorylation of Erk1/2 in MDCK-hCEACAM1-4L cells. Structure-function analysis using mutated hCEACAM1-4L protein shows that the cytoplasmic domain of the protein is needed for LPS-induced MAPK signalling, and that phosphorylation of Tyr-residues is not increased in association with MAPK signalling. The hCEACAM1-4L-dependent Erk1/2 phosphorylation develops in the presence of lipid A and does not develop in the presence of penta-acylated LPS. Finally, small interfering RNA (siRNA) silencing of canine TLR4 abolishes the hCEACAM1-4L-dependent, LPS-induced phosphorylation of Erk1/2. Collectively, our results support the notion that the apically expressed, full-length hCEACAM1-4L protein functions as a novel LPS-conveying molecule at the mucosal surface of polarized epithelial cells for subsequent MD-2/TLR4 receptor-dependent MAPK Erk1/2 and p38 signalling. PMID:21352462

  10. Transformation of Madin-Darby canine kidney (MDCK) epithelial cells by Epstein-Barr virus latent membrane protein 1 (LMP1) induces expression of Ets1 and invasive growth.

    PubMed

    Kim, K R; Yoshizaki, T; Miyamori, H; Hasegawa, K; Horikawa, T; Furukawa, M; Harada, S; Seiki, M; Sato, H

    2000-03-30

    The Epstein-Barr virus (EBV)-encoded latent membrane protein 1 (LMP1) has a significant role in initiating EBV-associated lymphoproliferative disease and EBV-related malignancies. In view of clinical features related to the type of EBV latency, LMP1 may influence invasiveness of EBV associated tumors categorized as types II and III as represented on nasopharyngeal carcinoma (NPC). To screen for genes associated with invasion of epithelial cells transformed by LMP1, Madin-Darby canine kidney (MDCK) epithelial cells were transformed by LMP1. Stable transfection of a LMP1 gene into MDCK cells induced morphological change from cobblestone to a long spindle-shape, reduced cell-cell adhesion and caused high cell motility. Parental MDCK cells, which form spherical cysts in three-dimensional collagen gel matrix, form branching tubules following exposure to hepatocyte growth factor (HGF). MDCK cells transformed by LMP1 showed invasive growth to form branching tubules into collagen gel without HGF-treatment. mRNA differential display and Northern hybridization identified plasminogen activator inhibitor-1 (PAI-1), urokinase type plasminogen activator (uPA) and ets1 as genes upregulated during transformation by LMP1. Expression of a dominant negative type of Etsl in LMP1-transformed cells downregulated uPA expression and cell motility. Deletion of LMP1 cytoplasmic carboxy-terminal activating region 1 (CTAR1) domain abolished transformation, but a deletion mutant lacking CTAR2 domain still retained transforming and uPA-inducing ability. Expression of Ets1 was immunolocalized in tumor cells of NPC tissue which frequently express LMP1. Taken together, it is suggested that LMP1 induces expression of Ets1 which may contribute to invasion of NPC by stimulating cell motility and uPA expression. PMID:10777210

  11. Solute Carrier Family of the Organic Anion-Transporting Polypeptides 1A2- Madin-Darby Canine Kidney II: A Promising In Vitro System to Understand the Role of Organic Anion-Transporting Polypeptide 1A2 in Blood-Brain Barrier Drug Penetration.

    PubMed

    Liu, Houfu; Yu, Na; Lu, Sijie; Ito, Sumito; Zhang, Xuan; Prasad, Bhagwat; He, Enuo; Lu, Xinyan; Li, Yang; Wang, Fei; Xu, Han; An, Gang; Unadkat, Jashvant D; Kusuhara, Hiroyuki; Sugiyama, Yuichi; Sahi, Jasminder

    2015-07-01

    Organic anion-transporting polypeptide (OATP) 1A2 has the potential to be a target for central nervous system drug delivery due to its luminal localization at the human blood-brain barrier and broad substrate specificity. We found OATP1A2 mRNA expression in the human brain to be comparable to breast cancer resistance protein and OATP2B1 and much higher than P-glycoprotein (P-gp), and confirmed greater expression in the brain relative to other tissues. The goal of this study was to establish a model system to explore OATP1A2-mediated transcellular transport of substrate drugs and the interplay with P-gp. In vitro (human embryonic kidney 293 cells stably expressing Oatp1a4, the closest murine isoform) and in vivo (naïve and Oatp1a4 knock-out mice) studies with OATP1A2 substrate triptan drugs demonstrated that these drugs were not Oatp1a4 substrates. This species difference demonstrates that the rodent is not a good model to investigate the active brain uptake of potential OATP1A2 substrates. Thus, we constructed a novel OATP1A2 expressing Madin-Darby canine kidney (MDCK) II wild type and an MDCKII-multidrug resistance protein 1 (MDR1) system using BacMam virus transduction. The spatial expression pattern of OATP1A2 after transduction in MDCKII-MDR1 cells was superimposed to P-gp, confirming apical membrane localization. OATP1A2-mediated uptake of zolmitriptan, rosuvastatin, and fexofenadine across monolayers increased with increasing OATP1A2 protein expression. OATP1A2 counteracted P-gp efflux for cosubstrates zolmitriptan and fexofenadine. A three-compartment model incorporating OATP1A2-mediated influx was used to quantitatively describe the time- and concentration-dependent apical-to-basolateral transcellular transport of rosuvastatin across OATP1A2 expressing the MDCKII monolayer. This novel, simple and versatile experimental system is useful for understanding the contribution of OATP1A2-mediated transcellular transport across barriers, such as the blood

  12. Zinc Supplementation Protects against Cadmium Accumulation and Cytotoxicity in Madin-Darby Bovine Kidney Cells

    PubMed Central

    Zhang, Ding; Liu, Jingying; Gao, Jianfeng; Shahzad, Muhammad; Han, Zhaoqing; Wang, Zhi; Li, Jiakui; Sjölinder, Hong

    2014-01-01

    Cadmium ions (Cd2+) have been reported to accumulate in bovine tissues, although Cd2+ cytotoxicity has not been investigated thoroughly in this species. Zinc ions (Zn2+) have been shown to antagonize the toxic effects of heavy metals such as Cd2+ in some systems. The present study investigated Cd2+ cytotoxicity in Madin-Darby bovine kidney (MDBK) epithelial cells, and explored whether this was modified by Zn2+. Exposure to Cd2+ led to a dose- and time-dependent increase in apoptotic cell death, with increased intracellular levels of reactive oxygen species and mitochondrial damage. Zn2+ supplementation alleviated Cd2+-induced cytotoxicity and this protective effect was more obvious when cells were exposed to a lower concentration of Cd2+ (10 μM), as compared to 50 μM Cd2+. This indicated that high levels of Cd2+ accumulation might induce irreversible damage in bovine kidney cells. Metallothioneins (MTs) are metal-binding proteins that play an essential role in heavy metal ion detoxification. We found that co-exposure to Zn2+ and Cd2+ synergistically enhanced RNA and protein expression of MT-1, MT-2, and the metal-regulatory transcription factor 1 in MDBK cells. Notably, addition of Zn2+ reduced the amounts of cytosolic Cd2+ detected following MDBK exposure to 10 μM Cd2+. These findings revealed a protective role of Zn2+ in counteracting Cd2+ uptake and toxicity in MDBK cells, indicating that this approach may provide a means to protect livestock from excessive Cd2+ accumulation. PMID:25105504

  13. Protein kinase C is involved in stimulation of arachidonic acid metabolism in Madin-Darby canine kidney (MDCK) cells

    SciTech Connect

    Parker, J.; Daniel, L.W.; Waite, M.

    1986-05-01

    The authors used 12-O-tetradecanoyl-phorbol-13-acetate (TPA) to directly stimulate protein kinase C (PKC) in order to examine the role of PKC in transduction of biological signals that increase metabolism of arachidonic acid. Release of radioactive arachidonic acid and prostaglandins from TPA-stimulated MDCK cells is inhibited by either of two PKC inhibitors: 1-(5-isoquinolinesulfonyl)piperazine and 1-octadecyl-2-methoxy-glycero-3-phosphocholine (ALP). ALP is unable to inhibit cyclooxygenase when added into an in vitro assay for this enzyme. Furthermore, TPA induces de novo synthesis of cyclooxygenase in MDCK cells but ALP fails to prevent this effect of TPA. Thus, cyclooxygenase activity appears to be independent of PKC and TPA can still induce de novo synthesis of cyclooxygenase even in the presence of the PKC inhibitor ALP. Also, ALP has no effect on the release of arachidonic acid which occurs upon addition of the calcium ionophore A23187 to MDCK cells suggesting that there are multiple mechanisms to mobilize arachidonic acid. Their data indicate that activation of PKC by TPA leads to increased release of arachidonic acid through regulation of phospholipase(s) by PKC.

  14. Microtubules and actin filaments are not critically involved in the biogenesis of epithelial cell surface polarity.

    PubMed

    Salas, P J; Misek, D E; Vega-Salas, D E; Gundersen, D; Cereijido, M; Rodriguez-Boulan, E

    1986-05-01

    We have studied the role of microtubules and actin filaments in the biogenesis of epithelial cell surface polarity, using influenza hemagglutinin and vesicular stomatitis G protein as model apical and basolateral proteins in infected Madin-Darby canine kidney cells. Addition of colchicine or nocodazole to confluent monolayers at concentrations sufficient to completely disassemble microtubules did not affect the asymmetric budding of influenza or vesicular stomatitis virus and only slightly reduced the typical asymmetric surface distribution of their envelope proteins, despite extensive cytoplasmic redistribution of the Golgi apparatus. Alteration of microtubular function by taxol or dissociation of actin filaments by cytochalasin D also failed to have a significant effect. Furthermore, neither colchicine nor cytochalasin D pretreatment blocked the ability of subconfluent Madin-Darby canine kidney cells to sustain polarized budding of influenza virus a few hours after attachment to the substrate. Our results indicate that domain-specific microtubule or actin filament "tracks" are not responsible for the vectorial delivery of apically or basolaterally directed transport vesicles. In conjunction with currently available evidence, they are compatible with a model in which receptors in the cytoplasmic aspect of apical or basolateral regions provide vectoriality to the transport of vesicles carrying plasma membrane proteins to their final surface localization. PMID:2871031

  15. 2,3,7,8-Tetrachlorodibenzo-p-dioxin increases Bovine Herpesvirus type-1 (BHV-1) replication in Madin-Darby Bovine Kidney (MDBK) cells in vitro.

    PubMed

    Fiorito, F; Pagnini, U; De Martino, L; Montagnaro, S; Ciarcia, R; Florio, S; Pacilio, M; Fucito, A; Rossi, A; Iovane, G; Giordano, A

    2008-01-01

    Dioxin-2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) is a common environmental toxin of current interest. In the last years, higher levels of TCDD than those permitted in UE [European Commission. 2002. European Commission Recommendation 2002/201/CE. Official Gazette, L 67/69] were detected in milk samples from cow, water buffalo, goat, and sheep raised on some areas of Campania Region (South Italy). Dioxin often causes immunosuppression and might render the animal liable to viral infections. In addition, viral infections are able to alter the pattern of dioxin distribution in different organs of the exposed animals. Bovine Herpesvirus type-1 (BHV-1) is a widespread pathogen, which causes infectious rhinotracheitis and infectious pustular vulvovaginitis in cattle. Herein, we have studied the effects of TCDD and BHV-1 infection, in Madin-Darby Bovine Kidney (MDBK) cells, alone as well as in association, so as cellular proliferation, apoptosis, and virus replication. We have observed an increase in cell viability of confluent monolayers at low TCDD concentrations. TCDD treated cells demonstrated increased viability compared to controls as evaluated by MTT test. TCDD exposure increased cell proliferation but induced no changes on apoptosis. Cells exposed to TCDD along with BHV-1 showed a dose-dependent increase in cytopathy, represented by ample syncytia formation with the elimination of the cellular sheets and increased viral titer. These results suggest that TCDD increases viral replication in MDBK cells while BHV-1 further decreases viability of TCDD exposed cells. Since very low concentrations (0.01 pg/ml) are sufficient to augment BHV-1 titer, TCDD may contribute to reactivate BHV-1 from latency, leading to recurrent disease and increase virus transmission. PMID:17516555

  16. Localization of GFP-tagged concentrative nucleoside transporters in a renal polarized epithelial cell line.

    PubMed

    Mangravite, L M; Lipschutz, J H; Mostov, K E; Giacomini, K M

    2001-05-01

    Many nucleosides undergo active reabsorption within the kidney, probably via nucleoside transporters. To date, two concentrative nucleoside transporters have been cloned, the sodium-dependent purine-selective nucleoside transporter (SPNT) and concentrative nucleoside transporter 1 (CNT1). We report the stable expression of green fluorescence protein (GFP)-tagged SPNT and CNT1 in Madin-Darby canine kidney (MDCK) cells, a polarized renal epithelial line. We demonstrate that the GFP tag does not alter the substrate selectivity and only modestly affects the kinetic activity of the transporters. By using confocal microscopy and functional studies, both SPNT and CNT1 are localized primarily to the apical membrane of MDCK and LLC-PK(1) cells. Apical localization of these transporters suggests a role in renal nucleoside reabsorption and regulation of tubular function via the adenosine pathway. PMID:11292631

  17. Changes in ganglioside content affect the binding of Clostridium perfringens epsilon-toxin to detergent-resistant membranes of Madin-Darby canine kidney cells.

    PubMed

    Shimamoto, Seiko; Tamai, Eiji; Matsushita, Osamu; Minami, Junzaburo; Okabe, Akinobu; Miyata, Shigeru

    2005-01-01

    Epsilon-toxin (ET) of Clostridium perfringens, which causes fatal enterotoxemia in ungulates, was previously shown to bind to and form a heptameric pore within the detergent-resistant membranes (DRMs) of MDCK cells. Depletion of cholesterol has also been shown to decrease the cytotoxicity of ET and its heptamerization. In this study, we investigated the effects of changes in sphingolipids, other DRM components of MDCK cells, on the cells' susceptibility to ET. Treatment with fumonisin B1 and PDMP, inhibitors of sphingolipid and glycosphingolipid syntheses, respectively, increased the susceptibility, while D609, a sphingomyelin synthesis inhibitor, had the opposite effect. The exogenous addition of ganglioside G(M1) dramatically decreased the ET binding, heptamerization and cytotoxicity. These effects were shown not to be due to ET binding to G(M1) or to denaturation of ET. We also found that the ET cytotoxicity towards MDCK cells decreased with an increase in culture time. In accordance with the resistance observed for prolonged cultured cells, G(M3), a major ganglioside component, increased and sialidase treatment increased their susceptibility. These results suggest that membrane-anchored sialic acid of G(M3) within DRMs inhibits ET binding, leading to prevention of the heptamerization of ET and cell death. It is also suggested that sialidase produced by this organism aids the targeting of ET to MDCK cells. PMID:15781998

  18. The pro-inflammatory cytokine tumor necrosis factor α stimulates expression of the carnitine transporter OCTN2 (novel organic cation transporter 2) and carnitine uptake via nuclear factor-κB in Madin-Darby bovine kidney cells.

    PubMed

    Zhou, X; Ringseis, R; Wen, G; Eder, K

    2015-06-01

    Carnitine uptake into tissues is mediated mainly by the novel organic cation transporter 2 (OCTN2), whose expression is upregulated in the liver of early-lactating dairy cows. It has been shown recently that pro-inflammatory cytokines, including tumor necrosis factor α (TNFα), stimulate OCTN2 expression and carnitine uptake in intestinal cells and inflamed intestinal mucosa. Given that many early-lactating dairy cows show typical signs of hepatic and systemic inflammation, such as elevated concentrations of circulating TNFα and activation of the key regulator of inflammation, nuclear factor κB (NF-κB), in tissues, it is possible that upregulation of OCTN2 and increase of carnitine uptake by TNFα is mediated by NF-κB, a mechanism that might contribute to the upregulation of OCNT2 in the liver of early-lactating dairy cows. Thus, in the present study, we tested the hypothesis that TNFα stimulates OCTN2 gene expression and carnitine uptake via NF-κB in the bovine Madin-Darby bovine kidney (MDBK) cell line. Treatment with TNFα caused activation of NF-κB, increased the mRNA and protein concentration of OCTN2, and stimulated the uptake of carnitine in MDBK cells. In contrast, combined treatment of MDBK cells with TNFα and the NF-κB inhibitor BAY 11-7085 completely blocked the effect of TNFα on OCTN2 mRNA and protein concentration and uptake of carnitine. These findings suggest that the bovine OCTN2 gene and carnitine uptake are regulated by NF-κB. Future studies are required to show the in vivo relevance of this regulatory mechanism in cattle. PMID:25892691

  19. Polarized trafficking of the sorting receptor SorLA in neurons and MDCK cells.

    PubMed

    Klinger, Stine C; Højland, Anne; Jain, Shweta; Kjolby, Mads; Madsen, Peder; Svendsen, Anna Dorst; Olivecrona, Gunilla; Bonifacino, Juan S; Nielsen, Morten S

    2016-07-01

    The sorting receptor SorLA is highly expressed in neurons and is also found in other polarized cells. The receptor has been reported to participate in the trafficking of several ligands, some of which are linked to human diseases, including the amyloid precursor protein, TrkB, and Lipoprotein Lipase (LpL). Despite this, only the trafficking in nonpolarized cells has been described so far. Due to the many differences between polarized and nonpolarized cells, we examined the localization and trafficking of SorLA in epithelial Madin-Darby canine kidney (MDCK) cells and rat hippocampal neurons. We show that SorLA is mainly found in sorting endosomes and on the basolateral surface of MDCK cells and in the somatodendritic domain of neurons. This polarized distribution of SorLA respectively depends on an acidic cluster and an extended version of this cluster and involves the cellular adaptor complex AP-1. Furthermore, we show that SorLA can mediate transcytosis across a tight cell layer. PMID:27192064

  20. Exocyst requirement for endocytic traffic directed toward the apical and basolateral poles of polarized MDCK cells.

    PubMed

    Oztan, Asli; Silvis, Mark; Weisz, Ora A; Bradbury, Neil A; Hsu, Shu-Chan; Goldenring, James R; Yeaman, Charles; Apodaca, Gerard

    2007-10-01

    The octameric exocyst complex is associated with the junctional complex and recycling endosomes and is proposed to selectively tether cargo vesicles directed toward the basolateral surface of polarized Madin-Darby canine kidney (MDCK) cells. We observed that the exocyst subunits Sec6, Sec8, and Exo70 were localized to early endosomes, transferrin-positive common recycling endosomes, and Rab11a-positive apical recycling endosomes of polarized MDCK cells. Consistent with its localization to multiple populations of endosomes, addition of function-blocking Sec8 antibodies to streptolysin-O-permeabilized cells revealed exocyst requirements for several endocytic pathways including basolateral recycling, apical recycling, and basolateral-to-apical transcytosis. The latter was selectively dependent on interactions between the small GTPase Rab11a and Sec15A and was inhibited by expression of the C-terminus of Sec15A or down-regulation of Sec15A expression using shRNA. These results indicate that the exocyst complex may be a multipurpose regulator of endocytic traffic directed toward both poles of polarized epithelial cells and that transcytotic traffic is likely to require Rab11a-dependent recruitment and modulation of exocyst function, likely through interactions with Sec15A. PMID:17686995

  1. Pak1 Regulates the Orientation of Apical Polarization and Lumen Formation by Distinct Pathways

    PubMed Central

    Smits, Jos; ter Beest, Martin B.; Zegers, Mirjam M.

    2012-01-01

    The development of the basic architecture of branching tubules enclosing a central lumen that characterizes most epithelial organs crucially depends on the apico-basolateral polarization of epithelial cells. Signals from the extracellular matrix control the orientation of the apical surface, so that it faces the lumen interior, opposite to cell-matrix adhesion sites. This orientation of the apical surface is thought to be intrinsically linked to the formation of single lumens. We previously demonstrated in three-dimensional cyst cultures of Madin-Darby canine kidney (MDCK) cells that signaling by β1 integrins regulates the orientation of the apical surface, via a mechanism that depends on the activity of the small GTPase Rac1. Here, we investigated whether the Rac1 effector Pak1 is a downstream effector in this pathway. Expression of constitutive active Pak1 phenocopies the effect of β1 integrin inhibition in that it misorients the apical surface and induces a multilumen phenotype. The misorientation of apical surfaces depends on the interaction of active Pak1 with PIX proteins and is linked to defects in basement membrane assembly. In contrast, the multilumen phenotype was independent of PIX and the basement membrane. Therefore, Pak1 likely regulates apical polarization and lumen formation by two distinct pathways. PMID:22815903

  2. A Novel Human Cytomegalovirus Glycoprotein, gpUS9, Which Promotes Cell-to-Cell Spread in Polarized Epithelial Cells, Colocalizes with the Cytoskeletal Proteins E-Cadherin and F-Actin

    PubMed Central

    Maidji, Ekaterina; Tugizov, Sharof; Abenes, Gerardo; Jones, Thomas; Pereira, Lenore

    1998-01-01

    Processes by which human herpesviruses penetrate and are released from polarized epithelial cells, which have distinct apical and basolateral membrane domains differing in protein and lipid content, are poorly understood. We recently reported that human cytomegalovirus (CMV) mutants with deletions of the gene US9 formed wild-type plaques in cultures of human fibroblasts but were impaired in the capacity for cell-to-cell spread in polarized human retinal pigment epithelial cells. Unlike the glycoproteins that are required for infection, the protein encoded by CMV US9 plays an accessory role by promoting dissemination of virus across cell-cell junctions of polarized epithelial cells. To identify the product and investigate its specialized functions, we selected Madine-Darby canine kidney II (MDCK) epithelial cells that constitutively express CMV US9 or, as a control, US8. The gene products, designated gpUS9 and gpUS8, were glycosylated proteins of comparable molecular masses but differed considerably in intracellular distribution and solubility. Immunofluorescence laser scanning confocal microscopy indicated that, like gpUS8, gpUS9 was present in the endoplasmic reticulum and Golgi compartments of nonpolarized cells. In polarized epithelial cells, gpUS9 also accumulated along lateral membranes, colocalizing with cadherin and actin, and was insoluble in Triton X-100, a property shared with proteins that associate with the cytoskeleton. We hypothesize that gpUS9 may enhance the dissemination of CMV in infected epithelial tissues by associating with the cytoskeletal matrix. PMID:9621030

  3. p21-Activated kinase 2 (PAK2) inhibits TGF-β signaling in Madin-Darby canine kidney (MDCK) epithelial cells by interfering with the receptor-Smad interaction.

    PubMed

    Yan, Xiaohua; Zhang, Junyu; Sun, Qinyu; Tuazon, Polygena T; Wu, Xiaoping; Traugh, Jolinda A; Chen, Ye-Guang

    2012-04-20

    TGF-β (transforming growth factor β) plays a variety of cellular functions mainly through the Smad pathway. Phosphorylation of the carboxyl SXS motif in R-Smads (Smad2 and Smad3) by the type I receptor TβRI is a key step for their activation. It has been reported that the serine/threonine kinase PAK2 (p21-activated kinase 2) can mediate TGF-β signaling in mesenchymal cells. Here, we report that PAK2 restricts TGF-β-induced Smad2/3 activation and transcriptional responsiveness in MDCK epithelial cells. Mechanistically, PAK2 associates with Smad2 and Smad3 in a kinase activity-dependent manner and blocks their activation. PAK2 phosphorylates Smad2 at Ser-417, which is adjacent to the L3 loop that contributes to the TβRI-R-Smad association. Consistently, substitution of Ser-417 with glutamic acid attenuates the interaction of Smad2 with TβRI. Together, our results indicate that PAK2 negatively modulate TGF-β signaling by attenuating the receptor-Smad interaction and thus Smad activation. PMID:22393057

  4. Infection of polarized MDCK cells with herpes simplex virus 1: two asymmetrically distributed cell receptors interact with different viral proteins.

    PubMed Central

    Sears, A E; McGwire, B S; Roizman, B

    1991-01-01

    Herpes simplex virus 1 attaches to at least two cell surface receptors. In polarized epithelial (Madin-Darby canine kidney; MDCK) cells one receptor is located in the apical surface and attachment to the cells requires the presence of glycoprotein C in the virus. The second receptor is located in the basal surface and does not require the presence of glycoprotein C. Exposure of MDCK cells at either the apical or basal surface to wild-type virus yields plaques and viral products whereas infection by a glycoprotein C-negative mutant yields identical results only after exposure of MDCK cells to virus at the basal surface. Multiple receptors for viral entry into cells expand the host range of the virus. The observation that glycoprotein C-negative mutants are infectious in many nonpolarized cell lines suggests that cells in culture may express more than one receptor and explains why genes that specify the viral proteins that recognize redundant receptors, like glycoprotein C, are expendable. Images PMID:1647025

  5. Infection of Polarized MDCK Cells with Herpes Simplex Virus 1: Two Asymmetrically Distributed Cell Receptors Interact with Different Viral Proteins

    NASA Astrophysics Data System (ADS)

    Sears, Amy E.; McGwire, Bradford S.; Roizman, Bernard

    1991-06-01

    Herpes simplex virus 1 attaches to at least two cell surface receptors. In polarized epithelial (Madin-Darby canine kidney; MDCK) cells one receptor is located in the apical surface and attachment to the cells requires the presence of glycoprotein C in the virus. The second receptor is located in the basal surface and does not require the presence of glycoprotein C. Exposure of MDCK cells at either the apical or basal surface to wild-type virus yields plaques and viral products whereas infection by a glycoprotein C-negative mutant yields identical results only after exposure of MDCK cells to virus at the basal surface. Multiple receptors for viral entry into cells expand the host range of the virus. The observation that glycoprotein C-negative mutants are infectious in many nonpolarized cell lines suggests that cells in culture may express more than one receptor and explains why genes that specify the viral proteins that recognize redundant receptors, like glycoprotein C, are expendable.

  6. Na,K-ATPase β-Subunit Is Required for Epithelial Polarization, Suppression of Invasion, and Cell Motility

    PubMed Central

    Rajasekaran, Sigrid A.; Palmer, Lawrence G.; Quan, Karina; Harper, Jeffrey F.; Ball, William J.; Bander, Neil H.; Soler, Alejandro Peralta; Rajasekaran, Ayyappan K.

    2001-01-01

    The cell adhesion molecule E-cadherin has been implicated in maintaining the polarized phenotype of epithelial cells and suppression of invasiveness and motility of carcinoma cells. Na,K-ATPase, consisting of an α- and β-subunit, maintains the sodium gradient across the plasma membrane. A functional relationship between E-cadherin and Na,K-ATPase has not previously been described. We present evidence that the Na,K-ATPase plays a crucial role in E-cadherin–mediated development of epithelial polarity, and suppression of invasiveness and motility of carcinoma cells. Moloney sarcoma virus-transformed Madin-Darby canine kidney cells (MSV-MDCK) have highly reduced levels of E-cadherin and β1-subunit of Na,K-ATPase. Forced expression of E-cadherin in MSV-MDCK cells did not reestablish epithelial polarity or inhibit the invasiveness and motility of these cells. In contrast, expression of E-cadherin and Na,K-ATPase β1-subunit induced epithelial polarization, including the formation of tight junctions and desmosomes, abolished invasiveness, and reduced cell motility in MSV-MDCK cells. Our results suggest that E-cadherin–mediated cell-cell adhesion requires the Na,K-ATPase β-subunit's function to induce epithelial polarization and suppress invasiveness and motility of carcinoma cells. Involvement of the β1-subunit of Na,K-ATPase in the polarized phenotype of epithelial cells reveals a novel link between the structural organization and vectorial ion transport function of epithelial cells. PMID:11179415

  7. Human cytomegalovirus glycoprotein B contains autonomous determinants for vectorial targeting to apical membranes of polarized epithelial cells.

    PubMed

    Tugizov, S; Maidji, E; Xiao, J; Zheng, Z; Pereira, L

    1998-09-01

    We previously reported that human cytomegalovirus (CMV) glycoprotein B (gB) is vectorially transported to apical membranes of CMV-infected polarized human retinal pigment epithelial cells propagated on permeable filter supports and that virions egress predominantly from the apical membrane domain. In the present study, we investigated whether gB itself contains autonomous information for apical transport by expressing the molecule in stably transfected Madine-Darby canine kidney (MDCK) cells grown on permeable filter supports. Laser scanning confocal immunofluorescence microscopy and domain-selective biotinylation of surface membrane domains showed that CMV gB was transported to apical membranes independently of other envelope glycoproteins and that it colocalized with proteins in transport vesicles of the biosynthetic and endocytic pathways. Determinants for trafficking to apical membranes were located by evaluating the targeting of gB derivatives with deletions in the lumen, transmembrane (TM) anchor, and carboxyl terminus. Derivative gB(Delta717-747), with an internal deletion in the luminal juxtamembrane sequence that preserved the N- and O-glycosylation sites, retained vectorial transport to apical membranes. In contrast, derivatives that lacked the TM anchor and cytosolic domain (gBDelta646-906) or the TM anchor alone (gBDelta751-771) underwent considerable basolateral targeting. Likewise, derivatives lacking the entire cytosolic domain (gBDelta772-906) or the last 73 amino acids (gBDelta834-906) showed disrupted apical transport. Site-specific mutations that deleted or altered the cluster of acidic residues with a casein kinase II phosphorylation site at the extreme carboxyl terminus, which can serve as an internalization signal, caused partial missorting of gB to basolateral membranes. Our studies indicate that CMV gB contains autonomous information for apical targeting in luminal, TM anchor, and cytosolic domain sequences, forming distinct structural

  8. Human Cytomegalovirus Glycoprotein B Contains Autonomous Determinants for Vectorial Targeting to Apical Membranes of Polarized Epithelial Cells

    PubMed Central

    Tugizov, Sharof; Maidji, Ekaterina; Xiao, Jianqiao; Zheng, Zhenwei; Pereira, Lenore

    1998-01-01

    We previously reported that human cytomegalovirus (CMV) glycoprotein B (gB) is vectorially transported to apical membranes of CMV-infected polarized human retinal pigment epithelial cells propagated on permeable filter supports and that virions egress predominantly from the apical membrane domain. In the present study, we investigated whether gB itself contains autonomous information for apical transport by expressing the molecule in stably transfected Madine-Darby canine kidney (MDCK) cells grown on permeable filter supports. Laser scanning confocal immunofluorescence microscopy and domain-selective biotinylation of surface membrane domains showed that CMV gB was transported to apical membranes independently of other envelope glycoproteins and that it colocalized with proteins in transport vesicles of the biosynthetic and endocytic pathways. Determinants for trafficking to apical membranes were located by evaluating the targeting of gB derivatives with deletions in the lumen, transmembrane (TM) anchor, and carboxyl terminus. Derivative gB(Δ717-747), with an internal deletion in the luminal juxtamembrane sequence that preserved the N- and O-glycosylation sites, retained vectorial transport to apical membranes. In contrast, derivatives that lacked the TM anchor and cytosolic domain (gBΔ646-906) or the TM anchor alone (gBΔ751-771) underwent considerable basolateral targeting. Likewise, derivatives lacking the entire cytosolic domain (gBΔ772-906) or the last 73 amino acids (gBΔ834-906) showed disrupted apical transport. Site-specific mutations that deleted or altered the cluster of acidic residues with a casein kinase II phosphorylation site at the extreme carboxyl terminus, which can serve as an internalization signal, caused partial missorting of gB to basolateral membranes. Our studies indicate that CMV gB contains autonomous information for apical targeting in luminal, TM anchor, and cytosolic domain sequences, forming distinct structural elements that

  9. Glycoprotein E of Varicella-Zoster Virus Enhances Cell-Cell Contact in Polarized Epithelial Cells

    PubMed Central

    Mo, Chengjun; Schneeberger, Eveline E.; Arvin, Ann M.

    2000-01-01

    Varicella-zoster virus (VZV) infection involves the cell-cell spread of virions, but how viral proteins interact with the host cell membranes that comprise intercellular junctions is not known. Madin-Darby canine kidney (MDCK) cells were constructed to express the glycoproteins gE, gI, or gE/gI constitutively and were used to examine the effects of these VZV glycoproteins in polarized epithelial cells. At low cell density, VZV gE induced partial tight junction (TJ) formation under low-calcium conditions, whether expressed alone or with gI. Although most VZV gE was intracellular, gE was also shown to colocalize with the TJ protein ZO-1 with or without concomitant expression of gI. Freeze fracture electron microscopy revealed normal TJ strand morphology in gE-expressing MDCK cells. Functionally, the expression of gE was associated with a marked acceleration in the establishment of maximum transepithelial electrical resistance (TER) in MDCK-gE cells; MDCK-gI and MDCK-gE/gI cells exhibited a similar pattern of early TER compared to MDCK cells, although peak resistances were lower than those of gE alone. VZV gE expression altered F-actin organization and lipid distribution, but coexpression of gI modulated these effects. Two regions of the gE ectodomain, amino acids (aa) 278 to 355 and aa 467 to 498, although lacking Ca2+ binding motifs, exhibit similarities with corresponding regions of the cell adhesion molecules, E-cadherin and desmocollin. These observations suggest that VZV gE and gE/gI may contribute to viral pathogenesis by facilitating epithelial cell-cell contacts. PMID:11070038

  10. The Sec6/8 complex in mammalian cells: characterization of mammalian Sec3, subunit interactions, and expression of subunits in polarized cells.

    PubMed

    Matern, H T; Yeaman, C; Nelson, W J; Scheller, R H

    2001-08-14

    The yeast exocyst complex (also called Sec6/8 complex in higher eukaryotes) is a multiprotein complex essential for targeting exocytic vesicles to specific docking sites on the plasma membrane. It is composed of eight proteins (Sec3, -5, -6, -8, -10, and -15, and Exo70 and -84), with molecular weights ranging from 70 to 144 kDa. Mammalian orthologues for seven of these proteins have been described and here we report the cloning and initial characterization of the remaining subunit, Sec3. Human Sec3 (hSec3) shares 17% sequence identity with yeast Sec3p, interacts in the two-hybrid system with other subunits of the complex (Sec5 and Sec8), and is expressed in almost all tissues tested. In yeast, Sec3p has been proposed to be a spatial landmark for polarized secretion (1), and its localization depends on its interaction with Rho1p (2). We demonstrate here that hSec3 lacks the potential Rho1-binding site and GFP-fusions of hSec3 are cytosolic. Green fluorescent protein (GFP)-fusions of nearly every subunit of the mammalian Sec6/8 complex were expressed in Madin-Darby canine kidney (MDCK) cells, but they failed to assemble into a complex with endogenous proteins and localized in the cytosol. Of the subunits tested, only GFP-Exo70 localized to lateral membrane sites of cell-cell contact when expressed in MDCK cells. Cells overexpressing GFP-Exo70 fail to form a tight monolayer, suggesting the Exo70 targeting interaction is critical for normal development of polarized epithelial cells. PMID:11493706

  11. Epoxyeicosatrienoic acids affect electrolyte transport in renal tubular epithelial cells: dependence on cyclooxygenase and cell polarity.

    PubMed

    Nüsing, Rolf M; Schweer, Horst; Fleming, Ingrid; Zeldin, Darryl C; Wegmann, Markus

    2007-07-01

    We investigated the effects of epoxyeicosatrienoic acids (EETs) on ion transport in the polarized renal distal tubular cell line, Madin-Darby canine kidney (MDCK) C7. Of the four EET regioisomers (5,6-EET, 8,9-EET, 11,12-EET, and 14,15-EET) studied, only apical, but not basolateral, application of 5,6-EET increased short-circuit current (I(sc)) with kinetics similar to those of arachidonic acid. The ion transport was blocked by preincubation with the cyclooxygenase inhibitor indomethacin or with the chloride channel blocker NPPB. Furthermore, both a Cl(-)-free bath solution and the Ca(2+) antagonist verapamil blocked 5,6-EET-induced ion transport. Although the presence of the PGE(2) receptors EP2, EP3, and EP4 was demonstrated, apically added PGE(2) was ineffective and basolaterally added PGE(2) caused a different kinetics in ion transport compared with 5,6-EET. Moreover, PGE(2) synthesis in MDCK C7 cells was unaffected by 5,6-EET treatment. GC/MS/MS analysis of cell supernatants revealed the presence of the biologically inactive 5,6-dihydroxy-PGE(1) in 5,6-EET-treated cells, but not in control cells. Indomethacin suppressed the formation of 5,6-dihydroxy-PGE(1). 5,6-Epoxy-PGE(1), the precursor of 5,6-dihydroxy-PGE(1), caused a similar ion transport as 5,6-EET. Cytochrome P-450 enzymes homolog to human CYP2C8, CYP2C9, and CYP2J2 protein were detected immunologically in the MDCK C7 cells. Our findings suggest that 5,6-EET affects Cl(-) transport in renal distal tubular cells independent of PGE(2) but by a mechanism, dependent on its conversion to 5,6-epoxy-PGE(1) by cyclooxygenase. We suggest a role for this P450 epoxygenase product in the regulation of electrolyte transport, especially as a saluretic compound acting from the luminal side of tubular cells in the mammalian kidney. PMID:17494091

  12. Metabolic flux profiling of MDCK cells during growth and canine adenovirus vector production

    PubMed Central

    Carinhas, Nuno; Pais, Daniel A. M.; Koshkin, Alexey; Fernandes, Paulo; Coroadinha, Ana S.; Carrondo, Manuel J. T.; Alves, Paula M.; Teixeira, Ana P.

    2016-01-01

    Canine adenovirus vector type 2 (CAV2) represents an alternative to human adenovirus vectors for certain gene therapy applications, particularly neurodegenerative diseases. However, more efficient production processes, assisted by a greater understanding of the effect of infection on producer cells, are required. Combining [1,2-13C]glucose and [U-13C]glutamine, we apply for the first time 13C-Metabolic flux analysis (13C-MFA) to study E1-transformed Madin-Darby Canine Kidney (MDCK) cells metabolism during growth and CAV2 production. MDCK cells displayed a marked glycolytic and ammoniagenic metabolism, and 13C data revealed a large fraction of glutamine-derived labelling in TCA cycle intermediates, emphasizing the role of glutamine anaplerosis. 13C-MFA demonstrated the importance of pyruvate cycling in balancing glycolytic and TCA cycle activities, as well as occurrence of reductive alphaketoglutarate (AKG) carboxylation. By turn, CAV2 infection significantly upregulated fluxes through most central metabolism, including glycolysis, pentose-phosphate pathway, glutamine anaplerosis and, more prominently, reductive AKG carboxylation and cytosolic acetyl-coenzyme A formation, suggestive of increased lipogenesis. Based on these results, we suggest culture supplementation strategies to stimulate nucleic acid and lipid biosynthesis for improved canine adenoviral vector production. PMID:27004747

  13. A simplified system for generating recombinant E3-deleted canine adenovirus-2.

    PubMed

    Yu, Zuo; Jiang, Qian; Liu, Jiasen; Guo, Dongchun; Quan, Chuansong; Li, Botao; Qu, Liandong

    2015-01-01

    Canine adenovirus type 2 (CAV-2) has been used extensively as a vector for studying gene therapy and vaccine applications. We describe a simple strategy for generating a replication-competent recombinant CAV-2 using a backbone vector and a shuttle vector. The backbone plasmid containing the full-length CAV-2 genome was constructed by homologous recombination in Escherichia coli strain BJ5183. The shuttle plasmid, which has a deletion of 1478 bp in the nonessential E3 viral genome region, was generated by subcloning a fusion fragment containing the flanking sequences of the CAV-2 E3 region and expression cassette sequences from pcDNA3.1(+) into modified pUC18. To determine system effectiveness, a gene for enhanced green fluorescent protein (EGFP) was inserted into the shuttle plasmid and cloned into the backbone plasmid using two unique NruI and SalI sites. Transfection of Madin-Darby canine kidney (MDCK) cells with the recombinant adenovirus genome containing the EGFP expression cassette resulted in infectious viral particles. This strategy provides a solid foundation for developing candidate vaccines using CAV-2 as a delivery vector. PMID:25450764

  14. 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

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

    1994-05-01

    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

  15. Kinetic analysis of human and canine P-glycoprotein-mediated drug transport in MDR1-MDCK cell model: approaches to reduce false-negative substrate classification.

    PubMed

    Li, Jibin; Wang, Ying; Hidalgo, Ismael J

    2013-09-01

    Madin-Darby canine kidney (MDCK) cells transfected with the multidrug resistance 1 (MDR1) gene, MDR1-MDCK, are widely used as an in vitro model to classify compounds as human P-glycoprotein (hPgp) substrates or nonsubstrates. Because MDCK cells express endogenous canine Pgp (cPgp), which is prone to downregulation after transfection with hPgp, this situation could lead to false-negative classification of hPgp substrates. The aim of this study was to investigate factors that influence hPgp substrate classification in MDR1-MDCK model and to seek ways to reduce false classification. Three-compartment models were used to derive flux equations describing the drug transport processes; factors influencing hPgp substrate classification were evaluated by simulations. Pgp functionality was assessed by determining the bidirectional permeability of a series of test compounds. Expressions of hPgp and cPgp were measured by quantitative polymerase chain reaction (qPCR). Kinetic model analysis revealed that the current net flux ratio calculation for hPgp substrate classification is influenced by endogenous cPgp expression as well as hPgp-cPgp expression ratio; the effect was more pronounced in low hPgp-cPgp region and diminished in high ratio region. On the basis of kinetic considerations, this study provides a rational experimental approach and appropriate mathematical corrections to minimize the potential occurrence of false-negative classification of new molecular entities. PMID:23558561

  16. Photoinduced cytotoxicity of a photochromic diarylethene via caspase cascade activation.

    PubMed

    Okuda, Jun-ya; Tanaka, Yukimi; Kodama, Ryuhei; Sumaru, Kimio; Morishita, Kana; Kanamori, Toshiyuki; Yamazoe, Seiji; Hyodo, Kengo; Yamazaki, Shohei; Miyatake, Tomohiro; Yokojima, Satoshi; Nakamura, Sinichiro; Uchida, Kingo

    2015-07-11

    The photo-generated closed-ring isomer of bis(5-methyl-2-phenylthiazoyl)perfluorocyclopentene shows cytotoxicity to Madin-Darby canine kidney (MDCK) cells through a caspase cascade and induces apoptosis of cells. PMID:26063471

  17. Production of canine adenovirus type 2 in serum-free suspension cultures of MDCK cells.

    PubMed

    Castro, R; Fernandes, P; Laske, T; Sousa, M F Q; Genzel, Y; Scharfenberg, K; Alves, P M; Coroadinha, A S

    2015-09-01

    The potential of adherent Madin Darby Canine Kidney (MDCK) cells for the production of influenza viruses and canine adenovirus type 2 (CAV-2) for vaccines or gene therapy approaches has been shown. Recently, a new MDCK cell line (MDCK.SUS2) that was able to grow in suspension in a fully defined system was established. In this work, we investigated whether the new MDCK.SUS2 suspension cell line is suitable for the amplification of CAV-2 under serum-free culture conditions. Cell growth performance and CAV-2 production were evaluated in three serum-free media: AEM, SMIF8, and EXCELL MDCK. CAV-2 production in shake flasks was maximal when AEM medium was used, resulting in an amplification ratio of infectious particles (IP) of 142 IP out/IP in and volumetric and cell-specific productivities of 2.1 × 10(8) IP/mL and 482 IP/cell, respectively. CAV-2 production was further improved when cells were cultivated in a 0.5-L stirred tank bioreactor. To monitor infection and virus production, cells were analyzed by flow cytometry. A correlation between the side scatter measurement and CAV-2 productivity was found, which represents a key feature to determine the best harvesting time during process development of gene therapy vectors that do not express reporter genes. This work demonstrates that MDCK.SUS2 is a suitable cell substrate for CAV-2 production, constituting a step forward in developing a production process transferable to industrial scales. This could allow for the production of high CAV-2 titers either for vaccination or for gene therapy purposes. PMID:25994255

  18. Diagnostic application of H3N8-specific equine influenza real-time reverse transcription polymerase chain reaction assays for the detection of Canine influenza virus in clinical specimens.

    PubMed

    Lu, Zhengchun; Dubovi, Edward J; Zylich, Nancy C; Crawford, P Cynda; Sells, Stephen; Go, Yun Young; Loynachan, Alan T; Timoney, Peter J; Chambers, Thomas M; Balasuriya, Udeni B R

    2010-11-01

    The objective of the current study was to determine the capability of 3 recently described one-step TaqMan real-time reverse transcription polymerase chain reaction (real-time RT-PCR) assays targeting the nucleoprotein (NP), matrix (M), and hemagglutinin (HA) genes of H3N8 Equine influenza virus (EIV NP, EIV M, and EIV HA3 assays, respectively) to detect Canine influenza virus (CIV). The assays were initially evaluated with nucleic acid extracted from tissue culture fluid (TCF) containing the A/canine/FL/43/04 strain of Influenza A virus associated with the 2004 canine influenza outbreak in Florida. The EIV NP, EIV M, and EIV HA3 assays could detect CIV nucleic acid at threshold cycle (Ct) values of 16.31, 23.71, and 15.28, respectively. Three assays using TCF or allantoic fluid (AF) samples containing CIV (n  =  13) and archived canine nasal swab samples (n  =  20) originally submitted for laboratory diagnosis of CIV were further evaluated. All TCF and AF samples, together with 10 nasal swab samples that previously tested positive for virus by attempted isolation in embryonated hens' eggs or Madin-Darby canine kidney cells, were positive in all 3 real-time RT-PCR assays. None of the 3 assays detected the H1N1 Swine influenza virus strain in current circulation. These findings demonstrate that previously described real-time RT-PCR assays targeting NP, M, and H3 HA gene segments of H3N8 EIV are also valuable for the diagnosis of CIV infection in dogs. The assays could expedite the detection and identification of CIV. PMID:21088179

  19. Enhanced replication of avian-origin H3N2 canine influenza virus in eggs, cell cultures and mice by a two-amino acid insertion in neuraminidase stalk.

    PubMed

    Lin, Yan; Xie, Xing; Zhao, Yanbing; Kalhoro, Dildar Hussain; Lu, Chengping; Liu, Yongjie

    2016-01-01

    Canine influenza virus (CIV) is a newly identified, highly contagious respiratory pathogen in dogs. Recent studies indicate that avian-origin H3N2 CIV are circulating in Chinese dogs. To investigate the effects of a two-amino acid (2-aa) insertion naturally occurring at the distal end of the neuraminidase (NA) stalk found in Chinese isolates since 2010 on virus replication and virulence, we rescued the CIV strain, A/canine/Jiangsu/06/2011(H3N2) and its NA mutant without the 2-aa insertion using reverse genetics. The NA stalk length affected virus growth in cell culture. Compared to the short stalk strain (without 2-aa insertion), the long stalk strain (with 2-aa insertion) exhibited higher peak titers and greater yields in Madin-Darby canine kidney (MDCK) cells, chicken embryo fibroblasts and canine bronchiolar epithelial cells, as well as much larger plaques in MDCK cell monolayers. Furthermore, mice inoculated with the long stalk strain showed more severe pathologic damage in lung and higher proportion of detectable viral RNA in tissues. The long stalk strain induced local IFN-γ production with faster kinetics and higher levels in mice. However, in chickens, the two viral strains showed no significant difference with nearly the same proportion of detectable viral RNA loads in tissues. These observations suggest that the 2-aa insertion in the NA stalk acquired by avian-origin H3N2 CIV helps to enhance viral replication and is likely a result of adaptive evolution in canine hosts. PMID:27160077

  20. Characteristics of taurine transport in cultured renal epithelial cell lines: asymmetric polarity of proximal and distal cell lines.

    PubMed

    Jones, D P; Miller, L A; Budreau, A; Chesney, R W

    1992-01-01

    Taurine transport was determined in two continuous, renal epithelial cell lines: LLC-PK1 derived from the proximal tubule of the pig, and the Madin-Darby canine kidney cell (MDCK) from the distal tubule of the dog. In LLC-PK1, taurine transport is maximal at the apical surface, whereas in MDCK cells, transport is greatest at the basolateral surface. Transport is highly dependent on both sodium and chloride in the external medium, and is specific for beta-amino acids. The apical and basolateral surfaces of both cell lines show an adaptive response to extracellular taurine concentration, but only the basolateral surface of the MDCK cell responds to hyperosomolality by increased taurine accumulation. Thus, differential control of the beta-amino acid transport system by substrate and external tonicity exists. The role of the beta-amino acid transport system may differ according to the origin of the cell: in the proximal renal tubular cell, net transepithelial reabsorption of filtered taurine increases the body pool. By contrast, taurine accumulation by distal tubular cells may form a mechanism of cell volume regulation in response to osmotic stress. PMID:1509959

  1. Transepithelial pressure pulses induce nucleotide release in polarized MDCK cells.

    PubMed

    Praetorius, H A; Frøkiaer, J; Leipziger, J

    2005-01-01

    The release of nucleotides is involved in mechanosensation in various epithelial cells. Intriguingly, kidney epithelial cells are absolutely dependent on the primary cilium to sense changes in apical laminar flow. During fluid passage, the renal epithelial cells are subjected to various mechanical stimuli in addition to changes in the laminar flow rate. In the distal part of the collecting duct, the epithelial cells are exposed to pressure changes and possibly distension during papillary contractions. The aim of the present study was to determine whether nucleotide release contributes to mechanosensation in kidney epithelial cells, thereby establishing whether pressure changes are sufficient to produce nucleotide-mediated responses. Madin-Darby canine kidney (MDCK) cells grown on permeable supports were mounted in a closed double perfusion chamber on an inverted microscope. The intracellular Ca(2+) concentration ([Ca(2+)](i)) was monitored with the Ca(2+)-sensitive fluorescence probe fluo 4. Transepithelial pressure pulses of 30-80 mm Hg produced a transient increase in [Ca(2+)](i) of MDCK cells. This response is independent of the primary cilium, since it is readily observed in immature cells that do not yet express primary cilia. The amplitudes of the pressure-induced Ca(2+) transients varied with the applied chamber pressure in a quantity-dependent manner. The ATPase apyrase and the P2Y antagonist suramin significantly reduced the pressure-induced Ca(2+) transients. Applying apyrase or suramin to both sides of the preparation simultaneously nearly abolished the pressure-induced Ca(2+) response. In conclusion, these observations suggest that rapid pressure changes induce both apical and basolateral nucleotide release that contribute to mechanosensation in kidney epithelial cells. PMID:15367389

  2. Galectin-7 modulates the length of the primary cilia and wound repair in polarized kidney epithelial cells.

    PubMed

    Rondanino, Christine; Poland, Paul A; Kinlough, Carol L; Li, Hui; Rbaibi, Youssef; Myerburg, Michael M; Al-bataineh, Mohammad M; Kashlan, Ossama B; Pastor-Soler, Nuria M; Hallows, Kenneth R; Weisz, Ora A; Apodaca, Gerard; Hughey, Rebecca P

    2011-09-01

    Galectins (Gal) are β-galactoside-binding proteins that function in epithelial development and homeostasis. An overlapping role for Gal-3 and Gal-7 in wound repair was reported in stratified epithelia. Although Gal-7 was thought absent in simple epithelia, it was reported in a proteomic analysis of cilia isolated from cultured human airway, and we recently identified Gal-7 transcripts in Madin-Darby canine kidney (MDCK) cells (Poland PA, Rondanino C, Kinlough CL, Heimburg-Molinaro J, Arthur CM, Stowell SR, Smith DF, Hughey RP. J Biol Chem 286: 6780-6790, 2011). We now report that Gal-7 is localized exclusively on the primary cilium of MDCK, LLC-PK(1) (pig kidney), and mpkCCD(c14) (mouse kidney) cells as well as on cilia in the rat renal proximal tubule. Gal-7 is also present on most cilia of multiciliated cells in human airway epithelia primary cultures. Interestingly, exogenous glutathione S-transferase (GST)-Gal-7 bound the MDCK apical plasma membrane as well as the cilium, while the lectin Ulex europeaus agglutinin, with glycan preferences similar to Gal-7, bound the basolateral plasma membrane as well as the cilium. In pull-down assays, β1-integrin isolated from either the basolateral or apical/cilia membranes of MDCK cells was similarly bound by GST-Gal-7. Selective localization of Gal-7 to cilia despite the presence of binding sites on all cell surfaces suggests that intracellular Gal-7 is specifically delivered to cilia rather than simply binding to surface glycoconjugates after generalized secretion. Moreover, depletion of Gal-7 using tetracycline-induced short-hairpin RNA in mpkCCD(c14) cells significantly reduced cilia length and slowed wound healing in a scratch assay. We conclude that Gal-7 is selectively targeted to cilia and plays a key role in surface stabilization of glycoconjugates responsible for integrating cilia function with epithelial repair. PMID:21677144

  3. A critical synopsis: Continuous growth of proximal tubular kidney epithelial cells in hormone-supplemented serum-free medium

    NASA Technical Reports Server (NTRS)

    Chuman, L. M.; FINE; COHEN; Saier, M. H.

    1985-01-01

    The kidney forms urine and reabsorbs electrolytes and water. Kidney cell lines and hormone supplemented serum free medium were used for growth. The hormones were insulin, transferrin, vasopressin, cholesterol, prostaglandins, hydrocortisone, and triidothyronine. Epithelial cell lines are polar and form hemicysts. The Madin-Darby canine kidney(MDCK) cell line used is distal tubulelike. LLC-PK sub 1 cells are derived from pig kidneys and have the properties of different kidney segments. The LLC-PK sub 1 cells with proximal tubule properties were maintained in hormone-supplemented serum free medium. Seven factors (the aforementioned homrones and selenium) were needed for growth. Hormone-defined medium supported LLC-PK sub 1 cell growth, allowed transport (as seen by hemicyst formation), and influenced cell morphology. Vasopressin (used for growth and morphology) could be partially replaced by isobutylmethylxanthine or dibutyryl cAMP. The defined medium was used to isolate rabbit proximal tubule kidney epithelial cells free of fibroblasts.

  4. Basic science track. Entry and release of canine coronavirus from polarized epithelial cells.

    PubMed

    Pratelli, Annamaria

    2011-01-01

    Virus entry into and release from epithelial cells are polarized as a result of the distribution of the viral receptors. In order to establish the polarity of entry and release of CCoV from epithelial cells, the interactions of the virus with A72 and CrFK cells grown on permeable supports was evaluated, and the amount of infective virus in the apical and in the basolateral media was determined and compared. Infection of A72 cells after different times post seeding demonstrated that CCoV grow after infection from both apical and basolateral sides. In CrFK cells, CCoV was observed in both compartments only in the later phase of the infection. To establish the reciprocal binding of CCoV on plasma membrane, A72 cells on a permeable support were preincubated with a mAb specific for CCoV. Infection from the apical side was blocked by mAb applied to that side; in contrast, such treatment on the basolateral side had no effect on the infectious process. Similarly, the low levels of CCoV observed after basolateral exposure to virus was abolished following mAb treatment of that side. The identification of CCoV into the basolateral medium could play an important role in the viral pathogenesis. PMID:21344143

  5. Dynamin Binding Protein (Tuba) Deficiency Inhibits Ciliogenesis and Nephrogenesis in Vitro and in Vivo.

    PubMed

    Baek, Jeong-In; Kwon, Sang-Ho; Zuo, Xiaofeng; Choi, Soo Young; Kim, Seok-Hyung; Lipschutz, Joshua H

    2016-04-15

    Dysfunction of renal primary cilia leads to polycystic kidney disease. We previously showed that the exocyst, a protein trafficking complex, is essential for ciliogenesis and regulated by multiple Rho and Rab family GTPases, such as Cdc42. Cdc42 deficiency resulted in a disruption of renal ciliogenesis and a polycystic kidney disease phenotype in zebrafish and mice. Here we investigate the role of Dynamin binding protein (also known as Tuba), a Cdc42-specific guanine nucleotide exchange factor, in ciliogenesis and nephrogenesis using Tuba knockdown Madin-Darby canine kidney cells and tuba knockdown in zebrafish. Tuba depletion resulted in an absence of cilia, with impaired apical polarization and inhibition of hepatocyte growth factor-induced tubulogenesis in Tuba knockdown Madin-Darby canine kidney cell cysts cultured in a collagen gel. In zebrafish, tuba was expressed in multiple ciliated organs, and, accordingly, tuba start and splice site morphants showed various ciliary mutant phenotypes in these organs. Co-injection of tuba and cdc42 morpholinos at low doses, which alone had no effect, resulted in genetic synergy and led to abnormal kidney development with highly disorganized pronephric duct cilia. Morpholinos targeting two other guanine nucleotide exchange factors not known to be in the Cdc42/ciliogenesis pathway and a scrambled control morpholino showed no phenotypic effect. Given the molecular nature of Cdc42 and Tuba, our data strongly suggest that tuba and cdc42 act in the same ciliogenesis pathway. Our study demonstrates that Tuba deficiency causes an abnormal renal ciliary and morphogenetic phenotype. Tuba most likely plays a critical role in ciliogenesis and nephrogenesis by regulating Cdc42 activity. PMID:26895965

  6. The ectopic expression of Snail in MDBK cells does not induce epithelial-mesenchymal transition

    PubMed Central

    IZAWA, GENYA; KOBAYASHI, WAKAKO; HARAGUCHI, MISAKO; SUDO, AKIHARU; OZAWA, MASAYUKI

    2015-01-01

    Epithelial-mesenchymal transition (EMT), a key process in the tumor metastatic cascade, is characterized by the loss of cell-cell junctions and cell polarity, as well as by the acquisition of migratory and invasive properties. However, the precise molecular events that initiate this complex EMT process are poorly understood. Snail expression induces EMT in Madin-Darby canine kidney (MDCK) cells and the human epidermoid carcinoma cell line, A431. Snail is a zinc finger transcription factor and triggers EMT by suppressing E-cadherin expression. In the present study, to broaden our knowledge of Snail-induced EMT, we generated stable Snail transfectants using Madin-Darby bovine kidney (MDBK) cells. Contrary to the MDCK or A431 cells examined in our previous studies, the MDBK cells transfected with the Snail construct maintained an epithelial morphology and showed no sign of reduced cell-cell adhesiveness compared to the control cells. Consistent with these observations, the down-regulation of epithelial marker proteins, e.g. E-cadherin and desmoglein, and the upregulation of mesenchymal marker proteins, e.g., N-cadherin and fibronectin, were not detected. Furthermore, the E-cadherin promoter was not methylated. Therefore, in the MDBK cells, the ectopic expression of Snail failed to induce EMT. As previously demonstrated, in MDCK cells, Snail expression is accompanied by the increased expression of other EMT-inducing transcription factors, e.g., Slug and zinc finger E-box-binding homeobox 1 (ZEB1). However, the MDBK cells transfected with the Snail construct did not exhibit an increased expression of these factors. Thus, it is possible that the failure to upregulate other EMT-related transcription factors may explain the lack of Snail-mediated induction of EMT in MDBK cells. PMID:25998899

  7. Canine Distemper

    MedlinePlus

    Although this brochure provides basic information about canine distemper, your veterinarian is always your best source of health information. Consult your veterinarian for more information about canine distemper and its prevention. ...

  8. Three-dimensional analysis of post-Golgi carrier exocytosis in epithelial cells.

    PubMed

    Kreitzer, Geri; Schmoranzer, Jan; Low, Seng Hui; Li, Xin; Gan, Yunbo; Weimbs, Thomas; Simon, Sanford M; Rodriguez-Boulan, Enrique

    2003-02-01

    Targeted delivery of proteins to distinct plasma membrane domains is critical to the development and maintenance of polarity in epithelial cells. We used confocal and time-lapse total internal reflection fluorescence microscopy (TIR-FM) to study changes in localization and exocytic sites of post-Golgi transport intermediates (PGTIs) carrying GFP-tagged apical or basolateral membrane proteins during epithelial polarization. In non-polarized Madin Darby Canine Kidney (MDCK) cells, apical and basolateral PGTIs were present throughout the cytoplasm and were observed to fuse with the basal domain of the plasma membrane. During polarization, apical and basolateral PGTIs were restricted to different regions of the cytoplasm and their fusion with the basal membrane was completely abrogated. Quantitative analysis suggested that basolateral, but not apical, PGTIs fused with the lateral membrane in polarized cells, correlating with the restricted localization of Syntaxins 4 and 3 to lateral and apical membrane domains, respectively. Microtubule disruption induced Syntaxin 3 depolarization and fusion of apical PGTIs with the basal membrane, but affected neither the lateral localization of Syntaxin 4 or Sec6, nor promoted fusion of basolateral PGTIs with the basal membrane. PMID:12545172

  9. 4.1N binding regions of inositol 1,4,5-trisphosphate receptor type 1.

    PubMed

    Fukatsu, Kazumi; Bannai, Hiroko; Inoue, Takafumi; Mikoshiba, Katsuhiko

    2006-04-01

    Zhang et al. and Maximov et al. [S. Zhang, A. Mizutani, C. Hisatsune, T. Higo, H. Bannai, T. Nakayama, M. Hattori, and K. Mikoshiba, Protein 4.1N is required for translocation of inositol 1,4,5-trisphosphate receptor type 1 to the basolateral membrane domain in polarized Madin-Darby canine kidney cells, J. Biol. Chem. 278 (2003) 4048-4056; A. Maximov, T. S. Tang, and I. Bezprozvanny, Association of the type 1 inositol (1,4,5)-trisphosphate receptor with 4.1N protein in neurons, Mol. Cell. Neurosci. 22 (2003) 271-283.] reported that 4.1N is a binding partner of inositol 1,4,5-trisphosphate receptor type 1 (IP(3)R1), however the binding site of IP(3)R1 differed: the former determined the C-terminal 14 amino acids of the cytoplasmic tail (CTT14aa) as the binding site, while the latter assigned another segment, cytoplasmic tail middle 1 (CTM1). To solve this discrepancy, we performed immunoprecipitation and found that both the segments had binding activity to 4.1N. Both segments also interfered the 4.1N-regulated IP(3)R1 diffusion in neuronal dendrites. However, IP(3)R1 lacking the CTT14aa (IP(3)R1-DeltaCTT14aa) does not bind to 4.1N [S. Zhang, A. Mizutani, C. Hisatsune, T. Higo, H. Bannai, T. Nakayama, M. Hattori, and K. Mikoshiba, Protein 4.1N is required for translocation of inositol 1,4,5-trisphosphate receptor type 1 to the basolateral membrane domain in polarized Madin-Darby canine kidney cells, J. Biol. Chem. 278 (2003) 4048-4056.] and its diffusion constant is larger than that of IP(3)R1 full-length in neuronal dendrites [K. Fukatsu, H. Bannai, S. Zhang, H. Nakamura, T. Inoue, and K. Mikoshiba, Lateral diffusion of inositol 1,4,5-trisphosphate receptor type 1 is regulated by actin filaments and 4.1N in neuronal dendrites, J. Biol. Chem. 279 (2004) 48976-48982.]. We conclude that both the CTT14aa and CTM1 sequences can bind to 4.1N in peptide fragment forms. However, we propose that the responsible binding site for 4.1N binding in full-length tetramer form of IP

  10. Ouabain modulates ciliogenesis in epithelial cells

    PubMed Central

    Larre, Isabel; Castillo, Aida; Flores-Maldonado, Catalina; Contreras, Ruben G.; Galvan, Ivan; Muñoz-Estrada, Jesus; Cereijido, Marcelino

    2011-01-01

    The exchange of substances between higher organisms and the environment occurs across transporting epithelia whose basic features are tight junctions (TJs) that seal the intercellular space, and polarity, which enables cells to transport substances vectorially. In a previous study, we demonstrated that 10 nM ouabain modulates TJs, and we now show that it controls polarity as well. We gauge polarity through the development of a cilium at the apical domain of Madin-Darby canine kidney cells (MDCK, epithelial dog kidney). Ouabain accelerates ciliogenesis in an ERK1/2-dependent manner. Claudin-2, a molecule responsible for the Na+ and H2O permeability of the TJs, is also present at the cilium, as it colocalizes and coprecipitates with acetylated α-tubulin. Ouabain modulates claudin-2 localization at the cilium through ERK1/2. Comparing wild-type and ouabain-resistant MDCK cells, we show that ouabain acts through Na+,K+-ATPase. Taken together, our previous and present results support the possibility that ouabain constitutes a hormone that modulates the transporting epithelial phenotype, thereby playing a crucial role in metazoan life. PMID:22143774

  11. Canine Parvovirus

    MedlinePlus

    Finally, do not let your puppy or adult dog to come into contact with the fecal waste of other dogs while walking or playing outdoors. Prompt and proper ... advisable as a way to limit spread of canine parvovirus infection as well as other diseases that ...

  12. VIP17/MAL expression modulates epithelial cyst formation and ciliogenesis

    PubMed Central

    Takiar, Vinita; Mistry, Kavita; Carmosino, Monica; Schaeren-Wiemers, Nicole

    2012-01-01

    The polarized organization of epithelial cells is required for vectorial solute transport and may be altered in renal cystic diseases. Vesicle integral protein of 17 kDa (VIP17/MAL) is involved in apical vesicle transport. VIP17/MAL overexpression in vivo results in renal cystogenesis of unknown etiology. Renal cystogenesis can occur as a consequence of defects of the primary cilium. To explore the role of VIP17/MAL in renal cystogenesis and ciliogenesis, we examined the polarization and ciliary morphology of wild-type and VIP17/MAL overexpressing Madin-Darby canine kidney renal epithelial cells grown in two-dimensional (2D) and three-dimensional (3D) cyst culture. VIP17/MAL is apically localized when expressed in cells maintained in 2D and 3D culture. VIP17/MAL overexpressing cells produce more multilumen cysts compared with controls. While the distributions of basolateral markers are not affected, VIP17/MAL expression results in aberrant sorting of the apical marker gp135 to the primary cilium. VIP17/MAL overexpression is also associated with shortened or absent cilia. Immunofluorescence analysis performed on kidney sections from VIP17/MAL transgenic mice also demonstrates fewer and shortened cilia within dilated lumens (P < 0.01). These studies demonstrate that VIP17/MAL overexpression results in abnormal cilium and cyst development, in vitro and in vivo, suggesting that VIP17/MAL overexpressing mice may develop cysts secondary to a ciliary defect. PMID:22895261

  13. Two distinct oncornaviruses harbor an intracytoplasmic tyrosine-based basolateral targeting signal in their viral envelope glycoprotein.

    PubMed Central

    Lodge, R; Delamarre, L; Lalonde, J P; Alvarado, J; Sanders, D A; Dokhélar, M C; Cohen, E A; Lemay, G

    1997-01-01

    It has been clearly established that the budding of the human immunodeficiency virus (HIV-1), a lentivirus, occurs specifically through the basolateral membrane in polarized epithelial cells. More recently, the signal was assigned to a tyrosine-based motif located in the intracytoplasmic domain of the envelope glycoprotein, as previously observed on various other viral and cellular basolateral proteins. In the present study, expression of human T-cell leukemia virus type 1 (HTLV-1) or Moloney murine leukemia virus envelope glycoproteins was used for trans-complementation of an envelope-negative HIV-1. This demonstrated the potential of oncornaviral retrovirus envelope glycoproteins to confer polarized basolateral budding in epithelial Madin-Darby canine kidney cells (MDCK cells). Site-directed mutagenesis confirmed the importance of a common motif encompassing at least one crucial membrane-proximal intracytoplasmic tyrosine residue. The conservation of a similar basolateral maturation signal in different retroviruses further supports its importance in the biology of this group of viruses. PMID:9188652

  14. Canine leishmaniosis.

    PubMed

    Sapierzyński, R

    2008-01-01

    Canine visceral leishmaniosis (CVL) is an infectious disease of zoonotic potential, caused by protozoan parasite of the genus Leishmania. Common clinical manifestations of canine visceral leishmaniosis include decrease of appetite, progressive weight loss, exercise intolerance, peripheral lymph node and spleen enlargement, chronic renal and liver disease, muscle, atrophy, polyarthritis and others. Because the Polish literature in the field contains no information on leishmaniosis in animals the recognised case of this disease is presented. Homeless mongrel, intact female dog, 3 years of age was brought to a veterinary clinic because of apathy, and generalised dermatologic lesions to perform routine examination. Because therapeutic effect of primarily recognised scabies was unsatisfactory, the skin samples from ear margins, trunk and lesion of the area of the left gluteal region for histopatologic examination were taken. Due to suspicion of leishmaniosis, fine-needle aspiration biopsy of lymph nodes, skin lesions, ocular discharge and imprint samples from skin lesion were performed, and tissue collected were examined under optical microscopy for identification of Leishmania amastigotes. To confirm cytologic diagnosis, blood samples for serological tests (enzyme-linked immunoabsorbent assay-ELISA; indirect immunofluorescence assay test-IFAT) were taken. Based on physical examination, histopatology, cytopathology and serology, canine visceral leishmaniosis was finally diagnosed. PMID:18683546

  15. Glycogen Synthase Kinase 3 (GSK-3) influences epithelial barrier function by regulating Occludin, Claudin-1 and E-cadherin expression

    SciTech Connect

    Severson, Eric A.; Kwon, Mike; Hilgarth, Roland S.; Parkos, Charles A.; Nusrat, Asma

    2010-07-02

    The Apical Junctional Complex (AJC) encompassing the tight junction (TJ) and adherens junction (AJ) plays a pivotal role in regulating epithelial barrier function and epithelial cell proliferative processes through signaling events that remain poorly characterized. A potential regulator of AJC protein expression is Glycogen Synthase Kinase-3 (GSK-3). GSK-3 is a constitutively active kinase that is repressed during epithelial-mesenchymal transition (EMT). In the present study, we report that GSK-3 activity regulates the structure and function of the AJC in polarized model intestinal (SK-CO15) and kidney (Madin-Darby Canine Kidney (MDCK)) epithelial cells. Reduction of GSK-3 activity, either by small molecule inhibitors or siRNA targeting GSK-3 alpha and beta mRNA, resulted in increased permeability to both ions and bulk solutes. Immunofluorescence labeling and immunoblot analyses revealed that the barrier defects correlated with decreased protein expression of AJC transmembrane proteins Occludin, Claudin-1 and E-cadherin without influencing other TJ proteins, Zonula Occludens-1 (ZO-1) and Junctional Adhesion Molecule A (JAM-A). The decrease in Occludin and E-cadherin protein expression correlated with downregulation of the corresponding mRNA levels for these respective proteins following GSK-3 inhibition. These observations implicate an important role of GSK-3 in the regulation of the structure and function of the AJC that is mediated by differential modulation of mRNA transcription of key AJC proteins, Occludin, Claudin-1 and E-cadherin.

  16. AMP-activated protein kinase induces actin cytoskeleton reorganization in epithelial cells

    SciTech Connect

    Miranda, Lisa; Carpentier, Sarah; Platek, Anna; Hussain, Nusrat; Gueuning, Marie-Agnes; Vertommen, Didier; Ozkan, Yurda; Sid, Brice; Hue, Louis; Courtoy, Pierre J.; Rider, Mark H.; Horman, Sandrine

    2010-06-04

    AMP-activated protein kinase (AMPK), a known regulator of cellular and systemic energy balance, is now recognized to control cell division, cell polarity and cell migration, all of which depend on the actin cytoskeleton. Here we report the effects of A769662, a pharmacological activator of AMPK, on cytoskeletal organization and signalling in epithelial Madin-Darby canine kidney (MDCK) cells. We show that AMPK activation induced shortening or radiation of stress fibers, uncoupling from paxillin and predominance of cortical F-actin. In parallel, Rho-kinase downstream targets, namely myosin regulatory light chain and cofilin, were phosphorylated. These effects resembled the morphological changes in MDCK cells exposed to hyperosmotic shock, which led to Ca{sup 2+}-dependent AMPK activation via calmodulin-dependent protein kinase kinase-{beta}(CaMKK{beta}), a known upstream kinase of AMPK. Indeed, hypertonicity-induced AMPK activation was markedly reduced by the STO-609 CaMKK{beta} inhibitor, as was the increase in MLC and cofilin phosphorylation. We suggest that AMPK links osmotic stress to the reorganization of the actin cytoskeleton.

  17. Cell confinement controls centrosome positioning and lumen initiation during epithelial morphogenesis

    PubMed Central

    Rodríguez-Fraticelli, Alejo E.; Auzan, Muriel; Alonso, Miguel A.; Bornens, Michel

    2012-01-01

    Epithelial organ morphogenesis involves sequential acquisition of apicobasal polarity by epithelial cells and development of a functional lumen. In vivo, cells perceive signals from components of the extracellular matrix (ECM), such as laminin and collagens, as well as sense physical conditions, such as matrix stiffness and cell confinement. Alteration of the mechanical properties of the ECM has been shown to promote cell migration and invasion in cancer cells, but the effects on epithelial morphogenesis have not been characterized. We analyzed the effects of cell confinement on lumen morphogenesis using a novel, micropatterned, three-dimensional (3D) Madin-Darby canine kidney cell culture method. We show that cell confinement, by controlling cell spreading, limits peripheral actin contractility and promotes centrosome positioning and lumen initiation after the first cell division. In addition, peripheral actin contractility is mediated by master kinase Par-4/LKB1 via the RhoA–Rho kinase–myosin II pathway, and inhibition of this pathway restores lumen initiation in minimally confined cells. We conclude that cell confinement controls nuclear–centrosomal orientation and lumen initiation during 3D epithelial morphogenesis. PMID:22965908

  18. Mitogen-stimulated and rapamycin-sensitive glucose transporter 12 targeting and functional glucose transport in renal epithelial cells.

    PubMed

    Wilson-O'Brien, Amy L; Dehaan, Carrie L; Rogers, Suzanne

    2008-03-01

    We hypothesized that glucose transporter 12 (GLUT12) is involved in regulation of glucose flux in distal renal tubules in response to elevated glucose. We used the Madin-Darby canine kidney polarized epithelial cell model and neutralizing antibodies to analyze GLUT12 targeting and directional GLUT12-mediated glucose transport. At physiological glucose concentrations, GLUT12 was localized to a perinuclear position. High glucose and serum treatment resulted in GLUT12 localization to the apical membrane. This mitogen-stimulated targeting of GLUT12 was inhibited by rapamycin, the specific inhibitor of mammalian target of rapamycin (mTOR). The functional role of GLUT12 was also examined. We constructed a GLUT12 cDNA containing a c-Myc epitope tag in the fifth exofacial loop. Assays of glucose transport at the apical membrane were performed using Transwell filters. By comparing transport assays in the presence of neutralizing anti-c-Myc monoclonal antibody, we specifically measured GLUT12-mediated glucose transport at the apical surface. GLUT12-mediated glucose transport was mitogen dependent and rapamycin sensitive. Our results implicate mTOR signaling in a novel pathway of glucose transporter protein targeting and glucose transport. Activity of the mTOR pathway has been associated with diabetic kidney disease. Our results provide evidence for a link between GLUT12 protein trafficking, glucose transport and signaling molecules central to the control of metabolic disease processes. PMID:18039784

  19. Soybean extracts increase cell surface ZIP4 abundance and cellular zinc levels: a potential novel strategy to enhance zinc absorption by ZIP4 targeting.

    PubMed

    Hashimoto, Ayako; Ohkura, Katsuma; Takahashi, Masakazu; Kizu, Kumiko; Narita, Hiroshi; Enomoto, Shuichi; Miyamae, Yusaku; Masuda, Seiji; Nagao, Masaya; Irie, Kazuhiro; Ohigashi, Hajime; Andrews, Glen K; Kambe, Taiho

    2015-12-01

    Dietary zinc deficiency puts human health at risk, so we explored strategies for enhancing zinc absorption. In the small intestine, the zinc transporter ZIP4 functions as an essential component of zinc absorption. Overexpression of ZIP4 protein increases zinc uptake and thereby cellular zinc levels, suggesting that food components with the ability to increase ZIP4 could potentially enhance zinc absorption via the intestine. In the present study, we used mouse Hepa cells, which regulate mouse Zip4 (mZip4) in a manner indistinguishable from that in intestinal enterocytes, to screen for suitable food components that can increase the abundance of ZIP4. Using this ZIP4-targeting strategy, two such soybean extracts were identified that were specifically able to decrease mZip4 endocytosis in response to zinc. These soybean extracts also effectively increased the abundance of apically localized mZip4 in transfected polarized Caco2 and Madin-Darby canine kidney cells and, moreover, two apically localized mZip4 acrodermatitis enteropathica mutants. Soybean components were purified from one extract and soyasaponin Bb was identified as an active component that increased both mZip4 protein abundance and zinc levels in Hepa cells. Finally, we confirmed that soyasaponin Bb is capable of enhancing cell surface endogenous human ZIP4 in human cells. Our results suggest that ZIP4 targeting may represent a new strategy to improve zinc absorption in humans. PMID:26385990

  20. Cd{sup 2+} versus Zn{sup 2+} uptake by the ZIP8 HCO{sub 3}{sup -}-dependent symporter: Kinetics, electrogenicity and trafficking

    SciTech Connect

    Liu Zhiwei; Li Hong; Soleimani, Manoocher; Girijashanker, Kuppuswami; Reed, Jodie M.; He Lei; Dalton, Timothy P.; Nebert, Daniel W.

    2008-01-25

    The mouse Slc39a8 gene encodes the ZIP8 transporter, which has been shown to be a divalent cation/HCO{sub 3}{sup -} symporter. Using ZIP8 cRNA-injected Xenopus oocyte cultures, we show herein that: [a] ZIP8-mediated cadmium (Cd{sup 2+}) and zinc (Zn{sup 2+}) uptake have V{sub max} values of 1.8 {+-} 0.08 and 1.0 {+-} 0.08 pmol/oocyte/h, and K{sub m} values of 0.48 {+-} 0.08 and 0.26 {+-} 0.09 {mu}M, respectively; [b] ZIP8-mediated Cd{sup 2+} uptake is most inhibited by Zn{sup 2+}, second-best inhibited by Cu{sup 2+}, Pb{sup 2+} and Hg{sup 2+}, and not inhibited by Mn{sup 2+} or Fe{sup 2+}; and [c] electrogenicity studies demonstrate an influx of two HCO{sub 3}{sup -} anions per one Cd{sup 2+} (or one Zn{sup 2+}) cation, i.e. electroneutral complexes. Using Madin-Darby canine kidney (MDCK) polarized epithelial cells retrovirally infected with ZIP8 cDNA and tagged with hemagglutinin at the C-terminus, we show that-similar to ZIP4-the ZIP8 eight-transmembrane protein is largely internalized during Zn{sup 2+} homeostasis, but moves predominantly to the cell surface membrane (trafficking) under conditions of Zn{sup 2+} depletion.

  1. FSGS3/CD2AP is a barbed-end capping protein that stabilizes actin and strengthens adherens junctions

    PubMed Central

    Brieher, William M.

    2013-01-01

    By combining in vitro reconstitution biochemistry with a cross-linking approach, we have identified focal segmental glomerulosclerosis 3/CD2-associated protein (FSGS3/CD2AP) as a novel actin barbed-end capping protein responsible for actin stability at the adherens junction. FSGS3/CD2AP colocalizes with E-cadherin and α-actinin-4 at the apical junction in polarized Madin-Darby canine kidney (MDCK) cells. Knockdown of FSGS3/CD2AP compromised actin stability and decreased actin accumulation at the adherens junction. Using a novel apparatus to apply mechanical stress to cell–cell junctions, we showed that knockdown of FSGS3/CD2AP compromised adhesive strength, resulting in tearing between cells and disruption of barrier function. Our results reveal a novel function of FSGS3/CD2AP and a previously unrecognized role of barbed-end capping in junctional actin dynamics. Our study underscores the complexity of actin regulation at cell–cell contacts that involves actin activators, inhibitors, and stabilizers to control adhesive strength, epithelial behavior, and permeability barrier integrity. PMID:24322428

  2. Epithelial cell invasion by bovine septicemic Escherichia coli.

    PubMed Central

    Korth, M J; Lara, J C; Moseley, S L

    1994-01-01

    Little is known regarding the pathogenesis of Escherichia coli-induced septicemic colibacillosis of calves. To understand the mechanism by which these strains penetrate the intestinal epithelium and gain access to the bloodstream, we examined the potential of bovine septicemic E. coli to invade cultured epithelial cells. By using a gentamicin survival assay, we demonstrated bacterial invasion of Madin-Darby canine kidney (MDCK) cells. Transcytosis of polarized MDCK cell monolayers was also observed, but only when bacteria were added to the basolateral surface. Electron microscopy confirmed the presence of intracellular organisms which appeared to be within membrane-bound vacuoles. The bovine septicemic isolate used in this study expressed the fimbrial adhesion CS31A. To examine the role of CS31A-mediated adherence in invasion and transcytosis of MDCK cell monolayers, a CS31A-deficient mutant was constructed by suicide vector-mediated insertional mutagenesis. Although nonadherent, the mutant showed a level of invasion similar to that of the wild-type parent. E. coli DH5 alpha carrying the cloned CS31A determinant was noninvasive. These findings suggest that expression of CS31A is neither required nor sufficient to mediate invasion. Images PMID:7903284

  3. BigA is a novel adhesin of Brucella that mediates adhesion to epithelial cells.

    PubMed

    Czibener, Cecilia; Merwaiss, Fernando; Guaimas, Francisco; Del Giudice, Mariela Giselda; Serantes, Diego Armando Rey; Spera, Juan Manuel; Ugalde, Juan Esteban

    2016-04-01

    Adhesion to cells is the initial step in the infectious cycle of basically all pathogenic bacteria, and to do so, microorganisms have evolved surface molecules that target different cellular receptors. Brucella is an intracellular pathogen that infects a wide range of mammals whose virulence is completely dependent on the capacity to replicate in phagocytes. Although much has been done to elucidate how Brucella multiplies in macrophages, we still do not understand how bacteria invade epithelial cells to perform a replicative cycle or what adhesion molecules are involved in the process. We report the identification in Brucella abortus of a novel adhesin that harbours a bacterial immunoglobulin-like domain and demonstrate that this protein is involved in the adhesion to polarized epithelial cells such as the Caco-2 and Madin-Darby canine kidney models targeting the bacteria to the cell-cell interaction membrane. While deletion of the gene significantly reduced adhesion, over-expression dramatically increased it. Addition of the recombinant protein to cells induced cytoskeleton rearrangements and showed that this adhesin targets proteins of the cell-cell interaction membrane in confluent cultures. PMID:26400021

  4. Canine lymphoma

    SciTech Connect

    Weller, R.E.

    1986-10-01

    Canine lymphoma has served as the ''workhorse'' for the development of veterinary oncology and as an important animal model for human non-Hodgkins lymphomas. Significant advances have been achieved in understanding the biological behavior of the disease and in its treatment. Although it is unlikely that a cure for lymphoma will be achieved, owners should be encouraged to treat their pets, provided they understand that only prolonged remissions and survivals are likely to result. Cooperative studies, employing large numbers of dogs, are needed to optimize and refine the classification scheme to provide a system with diagnostic and prognostic correlates and derive maximum benefit from therapeutic regimens. Such studies need to be prospective in nature, with a solid statistical base incorporated into their design. Rather than being content with what we have accomplished to date in treatment of canine lymphoma, the opportunity exists for the veterinary profession to make further significant contributions to the understanding and treatment of lymphoma in the dog. 10 refs., 4 tabs.

  5. Canine hyperlipidaemia.

    PubMed

    Xenoulis, P G; Steiner, J M

    2015-10-01

    Hyperlipidaemia refers to an increased concentration of lipids in the blood. Hyperlipidaemia is common in dogs and has recently emerged as an important clinical condition that requires a systematic diagnostic approach and appropriate treatment. Hyperlipidaemia can be either primary or secondary to other diseases. Secondary hyperlipidaemia is the most common form in dogs, and it can be a result of endocrine disorders, pancreatitis, cholestasis, protein-losing nephropathy, obesity, as well as other conditions and the use of certain drugs. Primary hyperlipidaemia is less common in the general canine population but it can be very common within certain breeds. Hypertriglyceridaemia of Miniature Schnauzers is the most common form of primary hyperlipidaemia in dogs but other breeds are also affected. Possible complications of hyperlipidaemia in dogs include pancreatitis, liver disease, atherosclerosis, ocular disease and seizures. Management of primary hyperlipidaemia in dogs is achieved by administration of ultra low-fat diets with or without the administration of lipid lowering drugs such as omega-3 fatty acids, fibrates, niacin and statins. PMID:26456868

  6. A statistical approach to determining criticality of residual host cell DNA.

    PubMed

    Yang, Harry; Wei, Ziping; Schenerman, Mark

    2015-01-01

    We propose a method for determining the criticality of residual host cell DNA, which is characterized through two attributes, namely the size and amount of residual DNA in biopharmaceutical product. By applying a mechanistic modeling approach to the problem, we establish the linkage between residual DNA and product safety measured in terms of immunogenicity, oncogenicity, and infectivity. Such a link makes it possible to establish acceptable ranges of residual DNA size and amount. Application of the method is illustrated through two real-life examples related to a vaccine manufactured in Madin Darby Canine Kidney cell line and a monoclonal antibody using Chinese hamster ovary (CHO) cell line as host cells. PMID:25358029

  7. Cysteine mutagenesis to study the structure of claudin-2 paracellular pores.

    PubMed

    Angelow, Susanne; Yu, Alan S L

    2009-05-01

    The structure and transport mechanism of paracellular pores are only poorly understood. Here we describe for the first time how the substituted cysteine accessibility method (SCAM), previously developed to study transmembrane transport, can be applied to analyze the pathway of paracellular ion permeation. Using stable transfected Madin Darby canine kidney type I cells, induced to express claudin-2, we show that paracellular cation transport can be blocked by sulfhydryl-specific methanethiosulfonate (MTS) and that SCAM can be used to identify residues that line paracellular pores. PMID:19538299

  8. Cell cytoskeletal conformation under reversible thermal control

    NASA Astrophysics Data System (ADS)

    Chang, Ting-Ya; Yang, Chung-Yao; Liao, Kai-Wei; Andrew Yeh, J.; Cheng, Chao-Min

    2013-12-01

    In order to assess the role of cytoskeletal structure in modulating cell surface topography during cell transformation, we investigated cytoskeletal organization of Madin-Darby canine kidney (MDCK) epithelial cells at different thermal gradients. Specifically, we examined actin polymerization as a function of temperature in a controlled thermal environment. After applying an increase in temperature of 5 °C, we observed fewer actin filaments in the network, as these molecular polymers depolymerized. Partial stress fibers of MDCK cells could be rearranged, but some of them were disrupted irreversibly after a second thermal treatment, and MDCK cells underwent apoptosis at higher temperatures as well.

  9. Dielectric spectroscopy for non-invasive monitoring of epithelial cell differentiation within three-dimensional scaffolds

    NASA Astrophysics Data System (ADS)

    Daoud, Jamal; Asami, Koji; Rosenberg, Lawrence; Tabrizian, Maryam

    2012-08-01

    In this study, we introduce a cellular differentiation cellular model based on dielectric spectroscopy that characterizes epithelial differentiation processes. Non-invasive cellular monitoring was achieved within a three-dimensional microenvironment consisting of a cell-containing collagen I gel seeded onto microfabricated scaffolds. In this proof-of-concept investigation, Madin-Darby canine kidney cells were cultured within microfabricated, geometrically controlled scaffolds and allowed us to differentiate to hollow cyst-like structures. This transformation within the three-dimensional environment is monitored and characterized through dielectric spectroscopy while maintaining cell culture in vitro.

  10. Generalized Klein-Kramers equation: solution and application

    NASA Astrophysics Data System (ADS)

    Sau Fa, Kwok; Wang, K. G.

    2013-09-01

    A generalized Klein-Kramers equation based on the continuous time random walk model is investigated. The equation generalizes the ordinary and fractional Klein-Kramers equations. Analytic solutions for the probability density and first two moments (for the force-free case) are obtained, and their dynamic behaviors are investigated in detail. The model is used to describe the cell migration of two migrating transformed renal epithelial Madin-Darby canine kidney (MDCK-F) cell strains: wild-type (NHE+) and NHE-deficient (NHE-) cells. Our theoretical predictions are in good agreement with experimental work in the paper (Dieterich et al 2008 Proc. Nat. Acad. Sci. USA 105 459).

  11. Localization of Impacted Canines

    PubMed Central

    Mehrotra, Praveen; Bhagchandani, Jitendra; Singh, Ashish; Garg, Aarti; Kumar, Snehi; Sharma, Ashish; Yadav, Harsh

    2015-01-01

    Impaction of maxillary canines is a frequently encountered clinical problem. The impaction of canine can be prevented in some situationsif the canine displacement is diagnosed in the early mixed dentition period and this would be extremely useful for the clinician. Hence,it is very important to focus on the means of early diagnosis and interception of this clinical situation. In the present article, the differentmodalities used to diagnose the impacted canine are reviewed with an insight into current 3-D modalities. PMID:25738100

  12. EB1-recruited microtubule +TIP complexes coordinate protrusion dynamics during 3D epithelial remodeling

    PubMed Central

    Gierke, Sarah; Wittmann, Torsten

    2012-01-01

    SUMMARY Background Epithelial remodeling, in which apical-basal polarized cells switch to a migratory phenotype, plays a central role in development and disease of multicellular organisms. Although dynamic microtubules (MTs) are required for directed migration on flat surfaces, how MT dynamics are controlled or contribute to epithelial remodeling in a more physiological three-dimensional (3D) environment is not understood. We use confocal live cell imaging to analyze MT function and dynamics during 3D epithelial morphogenesis and remodeling of polarized Madin-Darby canine kidney (MDCK) epithelial cells that undergo partial epithelial-to-mesenchymal transition (EMT) in response to hepatocyte growth factor (HGF). Results We find that HGF treatment increases MT growth rate before morphological changes are evident, and that large numbers of MTs grow into HGF-induced cell extensions independent of centrosome reorientation. Using lentivirus-mediated shRNA, we demonstrate that EB1, an adaptor protein that mediates recruitment of numerous other +TIP proteins to growing MT plus ends, is required for this HGF-induced MT reorganization. We further show that protrusion and adhesion dynamics are disorganized, and that vesicular trafficking to the tip of HGF-induced cell extensions is disrupted in EB1-depleted cells. Conclusions We conclude that EB1-mediated interactions with growing MTs are important to coordinate cell shape changes and directed migration into the surrounding extracellular matrix during epithelial remodeling in a physiological 3D environment. In contrast, EB1 is not required for the establishment or maintenance of apical-basal cell polarity, suggesting different functions of +TIPs and MTs in different types of cell polarity. PMID:22483942

  13. The exocyst gene Sec10 regulates renal epithelial monolayer homeostasis and apoptotic sensitivity.

    PubMed

    Polgar, Noemi; Lee, Amanda J; Lui, Vanessa H; Napoli, Josephine A; Fogelgren, Ben

    2015-08-01

    The highly conserved exocyst protein complex regulates polarized exocytosis of subsets of secretory vesicles. A previous study reported that shRNA knockdown of an exocyst central subunit, Sec10 (Sec10-KD) in Madin-Darby canine kidney (MDCK) cells disrupted primary cilia assembly and 3D cyst formation. We used three-dimensional collagen cultures of MDCK cells to further investigate the mechanisms by which Sec10 and the exocyst regulate epithelial polarity, morphogenesis, and homeostasis. Sec10-KD cysts initially demonstrated undisturbed lumen formation although later displayed significantly fewer and shorter primary cilia than controls. Later in cystogenesis, control cells maintained normal homeostasis, while Sec10-KD cysts displayed numerous apoptotic cells extruded basally into the collagen matrix. Sec10-KD MDCK cells were also more sensitive to apoptotic triggers than controls. These phenotypes were reversed by restoring Sec10 expression with shRNA-resistant human Sec10. Apico-basal polarity appeared normal in Sec10-KD cysts, whereas mitotic spindle angles differed significantly from controls, suggesting a planar cell polarity defect. In addition, analysis of renal tubules in a newly generated kidney-specific Sec10-knockout mouse model revealed significant defects in primary cilia assembly and in the targeted renal tubules; abnormal epithelial cell extrusion was also observed, supporting our in vitro results. We hypothesize that, in Sec10-KD cells, the disrupted exocyst activity results in increased apoptotic sensitivity through defective primary cilia signaling and that, in combination with an increased basal cell extrusion rate, it affects epithelial barrier integrity and homeostasis. PMID:26040895

  14. Core-glycosylated mucin-like repeats from MUC1 are an apical targeting signal.

    PubMed

    Kinlough, Carol L; Poland, Paul A; Gendler, Sandra J; Mattila, Polly E; Mo, Di; Weisz, Ora A; Hughey, Rebecca P

    2011-11-11

    MUC1 is efficiently delivered to the apical surface of polarized Madin-Darby canine kidney (MDCK) cells by transit through apical recycling endosomes, a route associated with delivery of apical proteins with glycan-dependent targeting signals. However, a role for glycans in MUC1 sorting has not been established. A key feature of MUC1 is a heavily O-glycosylated mucin-like domain with a variable number of nearly perfect tandem repeats and adjacent imperfect repeats. Metabolic labeling, cell surface biotinylation, immobilized lectins, and confocal immunofluorescence microscopy were used to characterize the polarized delivery of MUC1 mutants and chimeras in MDCK cells to identify the apical targeting signal. Both the interleukin-2 receptor α subunit (Tac) and a chimera where the Tac ectodomain replaced that of MUC1 were delivered primarily to the basolateral surface. Attachment of the MUC1 mucin-like domain to the N terminus of Tac enhanced apical but not basolateral delivery when compared with Tac. Conversely, deletions within the mucin-like domain in MUC1 reduced apical but not basolateral delivery when compared with MUC1. In pull-down assays with lectins, we found a notable difference in the presence of core 1 O-glycans, but not poly-N-acetyllactosamine, in apically targeted MUC1 and chimeras when compared with Tac. Consistent with these data, we found no effect on MUC1 targeting when galectin-3, with preference for poly-N-acetyllactosamine, was depleted from polarized MDCK cells. However, we did block the apical targeting activity of the mucin-like repeats when we overexpressed CMP-Neu5Ac:GalNAc-Rα2,6-sialyltransferase-1 to block core O-glycan synthesis. The cumulative data indicate that the core-glycosylated mucin-like repeats of MUC1 constitute an apical targeting signal. PMID:21937430

  15. An acidic cluster in the cytosolic domain of human cytomegalovirus glycoprotein B is a signal for endocytosis from the plasma membrane.

    PubMed

    Tugizov, S; Maidji, E; Xiao, J; Pereira, L

    1999-10-01

    We previously reported that human cytomegalovirus (CMV) glycoprotein B (gB) is transported to apical membranes in CMV-infected polarized retinal pigment epithelial (ARPE-19) cells and in Madin-Darby canine kidney (MDCK) epithelial cells constitutively expressing gB. The cytosolic domain of gB contains a cluster of acidic amino acids, a motif that plays a pivotal role in vectorial trafficking in polarized epithelial cells and may also function as a signal for entry into the endocytic pathway. Here we compared gB internalization and recycling to the plasma membrane in CMV-infected human fibroblasts (HF) and ARPE-19 cells by using antibody-internalization experiments. Immunofluorescence and quantitative assays showed that gB was internalized from the cell surface into clathrin-coated transport vesicles and then recycled to the plasma membrane. gB colocalized with clathrin-coated vesicles containing the transferrin receptor in the early endocytic/recycling pathway, indicating that gB traffics in this pathway. The specific role of the acidic cluster in regulating the sorting of gB-containing vesicles in the early endocytic/recycling pathway was examined in MDCK cells expressing mutated gB derivatives. Immunofluorescence assays showed that derivatives lacking the acidic cluster were impaired in internalization and failed to recycle. These findings, together with our earlier observation that the acidic cluster is a key determinant for targeting gB molecules to apical membranes in epithelial cells, establish that this signal is recognized by cellular proteins that participate in polarized sorting and transport in the early endocytic/recycling pathway. PMID:10482621

  16. An Acidic Cluster in the Cytosolic Domain of Human Cytomegalovirus Glycoprotein B Is a Signal for Endocytosis from the Plasma Membrane

    PubMed Central

    Tugizov, Sharof; Maidji, Ekaterina; Xiao, Jianqiao; Pereira, Lenore

    1999-01-01

    We previously reported that human cytomegalovirus (CMV) glycoprotein B (gB) is transported to apical membranes in CMV-infected polarized retinal pigment epithelial (ARPE-19) cells and in Madin-Darby canine kidney (MDCK) epithelial cells constitutively expressing gB. The cytosolic domain of gB contains a cluster of acidic amino acids, a motif that plays a pivotal role in vectorial trafficking in polarized epithelial cells and may also function as a signal for entry into the endocytic pathway. Here we compared gB internalization and recycling to the plasma membrane in CMV-infected human fibroblasts (HF) and ARPE-19 cells by using antibody-internalization experiments. Immunofluorescence and quantitative assays showed that gB was internalized from the cell surface into clathrin-coated transport vesicles and then recycled to the plasma membrane. gB colocalized with clathrin-coated vesicles containing the transferrin receptor in the early endocytic/recycling pathway, indicating that gB traffics in this pathway. The specific role of the acidic cluster in regulating the sorting of gB-containing vesicles in the early endocytic/recycling pathway was examined in MDCK cells expressing mutated gB derivatives. Immunofluorescence assays showed that derivatives lacking the acidic cluster were impaired in internalization and failed to recycle. These findings, together with our earlier observation that the acidic cluster is a key determinant for targeting gB molecules to apical membranes in epithelial cells, establish that this signal is recognized by cellular proteins that participate in polarized sorting and transport in the early endocytic/recycling pathway. PMID:10482621

  17. A single amino acid in the PB2 gene of influenza A virus is a determinant of host range.

    PubMed Central

    Subbarao, E K; London, W; Murphy, B R

    1993-01-01

    The single gene reassortant virus that derives its PB2 gene from the avian influenza A/Mallard/NY/78 virus and remaining genes from the human influenza A/Los Angeles/2/87 virus exhibits a host range restriction (hr) phenotype characterized by efficient replication in avian tissue and failure to produce plaques in mammalian Madin-Darby canine kidney cells. The hr phenotype is associated with restriction of viral replication in the respiratory tract of squirrel monkeys and humans. To identify the genetic basis of the hr phenotype, we isolated four phenotypic hr mutant viruses that acquired the ability to replicate efficiently in mammalian tissue. Segregational analysis indicated that the loss of the hr phenotype was due to a mutation in the PB2 gene itself. The nucleotide sequences of the PB2 gene of each of the four hr mutants revealed that a single amino acid substitution at position 627 (Glu-->Lys) was responsible for the restoration of the ability of the PB2 single gene reassortant to replicate in Madin-Darby canine kidney cells. Interestingly, the amino acid at position 627 in every avian influenza A virus PB2 protein analyzed to date is glutamic acid, and in every human influenza A virus PB2 protein, it is lysine. Thus, the amino acid at residue 627 of PB2 is an important determinant of host range of influenza A viruses. PMID:8445709

  18. Real-time monitoring of NKCC2 endocytosis by total internal reflection fluorescence (TIRF) microscopy.

    PubMed

    Jaykumar, Ankita Bachhawat; Caceres, Paulo S; Sablaban, Ibrahim; Tannous, Bakhos A; Ortiz, Pablo A

    2016-01-15

    The apical Na-K-2Cl cotransporter (NKCC2) mediates NaCl reabsorption by the thick ascending limb (TAL). The amount of NKCC2 at the apical membrane of TAL cells is determined by exocytic delivery, recycling, and endocytosis. Surface biotinylation allows measurement of NKCC2 endocytosis, but it has low time resolution and does not allow imaging of the dynamic process of endocytosis. We hypothesized that total internal reflection fluorescence (TIRF) microscopy imaging of labeled NKCC2 would allow monitoring of NKCC2 endocytosis in polarized Madin-Darby canine kidney (MDCK) and TAL cells. Thus we generated a NKCC2 construct containing a biotin acceptor domain (BAD) sequence between the transmembrane domains 5 and 6. Once expressed in polarized MDCK or TAL cells, surface NKCC2 was specifically biotinylated by exogenous biotin ligase (BirA). We also demonstrate that expression of a secretory form of BirA in TAL cells induces metabolic biotinylation of NKCC2. Labeling biotinylated surface NKCC2 with fluorescent streptavidin showed that most apical NKCC2 was located within small discrete domains or clusters referred to as "puncta" on the TIRF field. NKCC2 puncta were observed to disappear from the TIRF field, indicating an endocytic event which led to a decrease in the number of surface puncta at a rate of 1.18 ± 0.16%/min in MDCK cells, and a rate 1.09 ± 0.08%/min in TAL cells (n = 5). Treating cells with a cholesterol-chelating agent (methyl-β-cyclodextrin) completely blocked NKCC2 endocytosis. We conclude that TIRF microscopy of labeled NKCC2 allows the dynamic imaging of individual endocytic events at the apical membrane of TAL cells. PMID:26538436

  19. Molecular Mechanism of Renal Tubular Secretion of the Antimalarial Drug Chloroquine ▿

    PubMed Central

    Müller, Fabian; König, Jörg; Glaeser, Hartmut; Schmidt, Ingrid; Zolk, Oliver; Fromm, Martin F.; Maas, Renke

    2011-01-01

    The antimalarial drug chloroquine is eliminated to a significant extent by renal tubular secretion. The molecular mechanism of renal chloroquine secretion remains unknown. We hypothesized that organic cation transporter 2 (OCT2) and multidrug and toxin extrusion protein 1 (MATE1), localized in the basolateral and luminal membranes of proximal tubule cells, respectively, are involved in chloroquine transport. The interaction of chloroquine with both transporters was investigated using single-transfected human embryonic kidney 293 (HEK293)-MATE1 cells in uptake experiments and single-transfected Madin-Darby canine kidney II (MDCK)-OCT2 and MDCK-MATE1 cells as well as double-transfected MDCK-OCT2-MATE1 cells grown as polarized monolayers on transwell filters. In HEK293-MATE1 cells, chloroquine competitively inhibited MATE1-mediated metformin uptake (Ki = 2.8 μM). Cellular accumulation of chloroquine was significantly lower (P < 0.001) and transcellular chloroquine transport was significantly increased (P < 0.001) in MDCK-MATE1 and MDCK-OCT2-MATE1 cells compared to vector control cells after basal addition of chloroquine (0.1 to 10 μM). In contrast, no difference in cellular accumulation or transcellular transport of chloroquine was observed between MDCK-OCT2 and vector control cells. In line with an oppositely directed proton gradient acting as a driving force for MATE1, basal-to-apical transport of chloroquine by MDCK-OCT2-MATE1 cells increased with decreasing apical pH from 7.8 to 6.0. Transcellular transport of chloroquine by MDCK-OCT2-MATE1 cells was inhibited by cimetidine, trimethoprim, and amitriptyline. Our data demonstrate that chloroquine is a substrate and potent competitive inhibitor of MATE1, whereas OCT2 seems to play no role in chloroquine uptake. Concomitantly administered MATE1 inhibitors are likely to modify the renal secretion of chloroquine. PMID:21518836

  20. Identification and Functional Characterization of the First Nucleobase Transporter in Mammals

    PubMed Central

    Yamamoto, Syunsuke; Inoue, Katsuhisa; Murata, Tomoaki; Kamigaso, Syunsuke; Yasujima, Tomoya; Maeda, Jun-ya; Yoshida, Yukihiro; Ohta, Kin-ya; Yuasa, Hiroaki

    2010-01-01

    Nucleobases are important compounds that constitute nucleosides and nucleic acids. Although it has long been suggested that specific transporters are involved in their intestinal absorption and uptake in other tissues, none of their molecular entities have been identified in mammals to date. Here we describe identification of rat Slc23a4 as the first sodium-dependent nucleobase transporter (rSNBT1). The mRNA of rSNBT1 was expressed highly and only in the small intestine. When transiently expressed in HEK293 cells, rSNBT1 could transport uracil most efficiently. The transport of uracil mediated by rSNBT1 was sodium-dependent and saturable with a Michaelis constant of 21.2 μm. Thymine, guanine, hypoxanthine, and xanthine were also transported, but adenine was not. It was also suggested by studies of the inhibitory effect on rSNBT1-mediated uracil transport that several nucleobase analogs such as 5-fluorouracil are recognized by rSNBT1, but cytosine and nucleosides are not or only poorly recognized. Furthermore, rSNBT1 fused with green fluorescent protein was mainly localized at the apical membrane, when stably expressed in polarized Madin-Darby canine kidney II cells. These characteristics of rSNBT1 were almost fully in agreement with those of the carrier-mediated transport system involved in intestinal uracil uptake. Therefore, it is likely that rSNBT1 is its molecular entity or at least in part responsible for that. It was also found that the gene orthologous to the rSNBT1 gene is genetically defective in humans. This may have a biological and evolutional meaning in the transport and metabolism of nucleobases. The present study provides novel insights into the specific transport and metabolism of nucleobases and their analogs for therapeutic use. PMID:20042597

  1. VAMP7 Modulates Ciliary Biogenesis in Kidney Cells

    PubMed Central

    Szalinski, Christina M.; Labilloy, Anatália; Bruns, Jennifer R.; Weisz, Ora A.

    2014-01-01

    Epithelial cells elaborate specialized domains that have distinct protein and lipid compositions, including the apical and basolateral surfaces and primary cilia. Maintaining the identity of these domains is required for proper cell function, and requires the efficient and selective SNARE-mediated fusion of vesicles containing newly synthesized and recycling proteins with the proper target membrane. Multiple pathways exist to deliver newly synthesized proteins to the apical surface of kidney cells, and the post-Golgi SNAREs, or VAMPs, involved in these distinct pathways have not been identified. VAMP7 has been implicated in apical protein delivery in other cell types, and we hypothesized that this SNARE would have differential effects on the trafficking of apical proteins known to take distinct routes to the apical surface in kidney cells. VAMP7 expressed in polarized Madin Darby canine kidney cells colocalized primarily with LAMP2-positive compartments, and siRNA-mediated knockdown modulated lysosome size, consistent with the known function of VAMP7 in lysosomal delivery. Surprisingly, VAMP7 knockdown had no effect on apical delivery of numerous cargoes tested, but did decrease the length and frequency of primary cilia. Additionally, VAMP7 knockdown disrupted cystogenesis in cells grown in a three-dimensional basement membrane matrix. The effects of VAMP7 depletion on ciliogenesis and cystogenesis are not directly linked to the disruption of lysosomal function, as cilia lengths and cyst morphology were unaffected in an MDCK lysosomal storage disorder model. Together, our data suggest that VAMP7 plays an essential role in ciliogenesis and lumen formation. To our knowledge, this is the first study implicating an R-SNARE in ciliogenesis and cystogenesis. PMID:24466086

  2. The ABCG2 Efflux Transporter in the Mammary Gland Mediates Veterinary Drug Secretion across the Blood-Milk Barrier into Milk of Dairy Cows.

    PubMed

    Mahnke, Hanna; Ballent, Mariana; Baumann, Sven; Imperiale, Fernanda; von Bergen, Martin; Lanusse, Carlos; Lifschitz, Adrian L; Honscha, Walther; Halwachs, Sandra

    2016-05-01

    In human and mice ATP-binding cassette efflux transporter ABCG2 represents the main route for active drug transport into milk. However, there is no detailed information on the role of ABCG2 in drug secretion and accumulation in milk of dairy animals. We therefore examined ABCG2-mediated drug transport in the bovine mammary gland by parallel pharmacokinetic studies in lactating Jersey cows and in vitro flux studies using the anthelmintic drug monepantel (MNP) as representative bovine ABCG2 (bABCG2) drug substrate. Animals received MNP (Zolvix, Novartis Animal Health Inc.) once (2.5 mg/kg per os) and the concentrations of MNP and the active MNP metabolite MNPSO2were assessed by high-performance liquid chromatography. Compared with the parent drug MNP, we detected higher MNPSO2plasma concentrations (expressed as area under the concentration-versus-time curve). Moreover, we observed MNPSO2excretion into milk of dairy cows with a high milk-to-plasma ratio of 6.75. In mechanistic flux assays, we determined a preferential time-dependent basolateral-to-apical (B > A) MNPSO2transport across polarized Madin-Darby canine kidney II cells-bABCG2 monolayers using liquid chromatography coupled with tandem mass spectrometry analysis. The B > A MNPSO2transport was significantly inhibited by the ABCG2 inhibitor fumitremorgin C in bABCG2- but not in mock-transduced MDCKII cells. Additionally, the antibiotic drug enrofloxacin, the benzimidazole anthelmintic oxfendazole and the macrocyclic lactone anthelmintic moxidectin caused a reduction in the MNPSO2(B > A) net efflux. Altogether, this study indicated that therapeutically relevant drugs like the anthelmintic MNP represent substrates of the bovine mammary ABCG2 transporter and may thereby be actively concentrated in dairy milk. PMID:26956640

  3. Evaluation of the bioavailability of major withanolides of Withania somnifera using an in vitro absorption model system.

    PubMed

    Devkar, Santosh T; Kandhare, Amit D; Sloley, Brian D; Jagtap, Suresh D; Lin, James; Tam, Yun K; Katyare, Surendra S; Bodhankar, Subhash L; Hegde, Mahabaleshwar V

    2015-01-01

    Withania somnifera (L.) Dunal, shows several pharmacological properties which are attributed mainly to the withanolides present in the root. The efficacy of medicinally active withanolides constituents depends on the absorption and transportation through the intestinal epithelium. We examined these characteristics by employing the Sino-Veda Madin-Darby canine kidney cells culture system, which under in vitro condition shows the absorption characteristics similar to the human intestinal epithelium. Thus, the aim of the present investigation was to assess the bioavailability of individual withanolides. Withanolides were diluted in Hank's buffered saline at a concentration of 2 μg/ml were tested for permeability studies carried out for 1 h duration. Permeability was measured in terms of efflux pump (P eff) in cm/s. P eff values of withanolide A (WN A), withanone (WNN), 1,2-deoxywithastramonolide (1,2 DWM), withanolide B (WN B), withanoside IV-V (WS IV-V), and withaferin A were 4.05 × 10(-5), 2.06 × 10(-5), 1.97 × 10(-5), 1.80 × 10(-5), 3.19 × 10(-6), 3.03 × 10(-6) and 3.30 × 10(-7) respectively. In conclusion, the nonpolar and low molecular weight compounds (WN A, WNN, 1,2 DWM, and WN B) were highly permeable. As against this, the glycosylated and polar WS IV and WS V showed low permeability. Surprisingly and paradoxically, the highly biologically active withaferin A was completely impermeable, suggesting that further studies possibly using human epithelial colorectal adenocarcinoma (Caco-2) cells may be needed to delineate the absorption characteristics of withanolides, especially withaferin A. PMID:26605156

  4. The inwardly rectifying potassium channel Kir1.1: development of functional assays to identify and characterize channel inhibitors.

    PubMed

    Felix, John P; Priest, Birgit T; Solly, Kelli; Bailey, Timothy; Brochu, Richard M; Liu, Chou J; Kohler, Martin G; Kiss, Laszlo; Alonso-Galicia, Magdalena; Tang, Haifeng; Pasternak, Alexander; Kaczorowski, Gregory J; Garcia, Maria L

    2012-10-01

    The renal outer medullary potassium (ROMK) channel is a member of the inwardly rectifying family of potassium (Kir) channels. ROMK (Kir1.1) is predominantly expressed in kidney where it plays a major role in the salt reabsorption process. Loss-of-function mutations in the human Kir1.1 channel are associated with antenatal Bartter's syndrome type II, a life-threatening salt and water balance disorder. Heterozygous carriers of Kir1.1 mutations associated with antenatal Bartter's syndrome have reduced blood pressure and a decreased risk of developing hypertension by age 60. These data suggest that Kir1.1 inhibitors could represent novel diuretics for the treatment of hypertension. Because little is known about the molecular pharmacology of Kir1.1 channels, assays that provide a robust, reliable readout of channel activity-while operating in high-capacity mode-are needed. In the present study, we describe high-capacity, 384- and 1,536-well plate, functional thallium flux, and IonWorks electrophysiology assays for the Kir1.1 channel that fulfill these criteria. In addition, 96-well (86)Rb(+) flux assays were established that can operate in the presence of 100% serum, and can provide an indication of the effect of a serum shift on compound potencies. The ability to grow Madin-Darby canine kidney cells expressing Kir1.1 in Transwell supports provides a polarized cell system that can be used to study the mechanism of Kir1.1 inhibition by different agents. All these functional Kir1.1 assays together can play an important role in supporting different aspects of drug development efforts during lead identification and/or optimization. PMID:22881347

  5. Regulation of Epithelial-to-Mesenchymal Transition Using Biomimetic Fibrous Scaffolds.

    PubMed

    Ravikrishnan, Anitha; Ozdemir, Tugba; Bah, Mohamed; Baskerville, Karen A; Shah, S Ismat; Rajasekaran, Ayyappan K; Jia, Xinqiao

    2016-07-20

    Epithelial-to-mesenchymal transition (EMT) is a well-studied biological process that takes place during embryogenesis, carcinogenesis, and tissue fibrosis. During EMT, the polarized epithelial cells with a cuboidal architecture adopt an elongated fibroblast-like morphology. This process is accompanied by the expression of many EMT-specific molecular markers. Although the molecular mechanism leading to EMT has been well-established, the effects of matrix topography and microstructure have not been clearly elucidated. Synthetic scaffolds mimicking the meshlike structure of the basement membrane with an average fiber diameter of 0.5 and 5 μm were produced via the electrospinning of poly(ε-caprolactone) (PCL) and were used to test the significance of fiber diameter on EMT. Cell-adhesive peptide motifs were conjugated to the fiber surface to facilitate cell attachment. Madin-Darby Canine Kidney (MDCK) cells grown on these substrates showed distinct phenotypes. On 0.5 μm substrates, cells grew as compact colonies with an epithelial phenotype. On 5 μm scaffolds, cells were more individually dispersed and appeared more fibroblastic. Upon the addition of hepatocyte growth factor (HGF), an EMT inducer, cells grown on the 0.5 μm scaffold underwent pronounced scattering, as evidenced by the alteration of cell morphology, localization of focal adhesion complex, weakening of cell-cell adhesion, and up-regulation of mesenchymal markers. In contrast, HGF did not induce a pronounced scattering of MDCK cells cultured on the 5.0 μm scaffold. Collectively, our results show that the alteration of the fiber diameter of proteins found in the basement membrane may create enough disturbances in epithelial organization and scattering that might have important implications in disease progression. PMID:27322677

  6. Regulation of the Axillary Osmidrosis-Associated ABCC11 Protein Stability by N-Linked Glycosylation: Effect of Glucose Condition

    PubMed Central

    Toyoda, Yu; Takada, Tappei; Miyata, Hiroshi; Ishikawa, Toshihisa; Suzuki, Hiroshi

    2016-01-01

    ATP-binding cassette C11 (ABCC11) is a plasma membrane protein involved in the transport of a variety of lipophilic anions. ABCC11 wild-type is responsible for the high-secretion phenotypes in human apocrine glands, such as that of wet-type ear wax, and the risk of axillary osmidrosis. We have previously reported that mature ABCC11 is a glycoprotein containing two N-linked glycans at Asn838 and Asn844. However, little is known about the role of N-linked glycosylation in the regulation of ABCC11 protein. In the current study, we investigated the effects of N-linked glycosylation on the protein level and localization of ABCC11 using polarized Madin-Darby canine kidney II cells. When the N-linked glycosylation in ABCC11-expressing cells was chemically inhibited by tunicamycin treatment, the maturation of ABCC11 was suppressed and its protein level was significantly decreased. Immunoblotting analyses demonstrated that the protein level of the N-linked glycosylation-deficient mutant (N838Q and N844Q: Q838/844) was about half of the ABCC11 wild-type level. Further biochemical studies with the Q838/844 mutant showed that this glycosylation-deficient ABCC11 was degraded faster than wild-type probably due to the enhancement of the MG132-sensitive protein degradation pathway. Moreover, the incubation of ABCC11 wild-type-expressing cells in a low-glucose condition decreased mature, glycosylated ABCC11, compared with the high-glucose condition. On the other hand, the protein level of the Q838/844 mutant was not affected by glucose condition. These results suggest that N-linked glycosylation is important for the protein stability of ABCC11, and physiological alteration in glucose may affect the ABCC11 protein level and ABCC11-related phenotypes in humans, such as axillary osmidrosis. PMID:27281343

  7. Involvement of intestinal uptake transporters in the absorption of azithromycin and clarithromycin in the rat.

    PubMed

    Garver, Eric; Hugger, Erin D; Shearn, Shawn P; Rao, Anuradha; Dawson, Paul A; Davis, Charles B; Han, Chao

    2008-12-01

    Macrolide antibiotics azithromycin (AZI) and clarithromycin (CLARI) are large molecular weight compounds and are substrates for apically polarized efflux transporters such as P-glycoprotein, which can potentially restrict intestinal absorption. However, despite these undesired physicochemical and biopharmaceutical properties, AZI and CLARI exhibit moderate to excellent p.o. bioavailability in preclinical species and humans. Intestinal uptake transporters, such as organic anion transporting polypeptides (OATPs), can facilitate the uptake of drugs that are substrates and hence increase p.o. absorption. The present study was designed to determine whether the intestinal Oatps are involved in absorption of these macrolides. AZI or CLARI was dosed p.o. to Sprague-Dawley rats after p.o. administration with vehicle or rifamycin SV (RIF), an OATP inhibitor. The p.o. exposures of AZI and CLARI were reduced 65 and 45%, respectively, when coadministered with an optimized RIF regimen. The p.o. RIF had no affect on the total blood clearance of these macrolides and most likely did not cause induction of metabolizing enzymes and/or transporters. Therefore, the results suggest that inhibition of an RIF-sensitive uptake transporter such as Oatp along the rat gastrointestinal tract was responsible for reduced p.o. exposure of AZI and CLARI. In addition, AZI and CLARI caused inhibition of taurocholate uptake in rat Oatp1a5-transfected Madin-Darby canine kidney cell monolayers. The in vitro and in vivo results suggest that the intestinal Oatps are involved in the p.o. absorption of AZI and CLARI in the rat. PMID:18755851

  8. Syntaxin specificity of aquaporins in the inner medullary collecting duct.

    PubMed

    Mistry, Abinash C; Mallick, Rickta; Klein, Janet D; Weimbs, Thomas; Sands, Jeff M; Fröhlich, Otto

    2009-08-01

    Proper targeting of the aquaporin-2 (AQP2) water channel to the collecting duct apical plasma membrane is critical for the urine concentrating mechanism and body water homeostasis. However, the trafficking mechanisms that recruit AQP2 to the plasma membrane are still unclear. Snapin is emerging as an important mediator in the initial interaction of trafficked proteins with target soluble N-ethylmaleimide-sensitive factor attachment protein (SNAP) receptor (t-SNARE) proteins, and this interaction is functionally important for AQP2 regulation. We show that in AQP2-Madin-Darby canine kidney cells subjected to adenoviral-mediated expression of both snapin and syntaxins, the association of AQP2 with both syntaxin-3 and syntaxin-4 is highly enhanced by the presence of snapin. In pull-down studies, snapin detected AQP2, syntaxin-3, syntaxin-4, and SNAP23 from the inner medullary collecting duct. AQP2 transport activity, as probed by AQP2's urea permeability, was greatly enhanced in oocytes that were coinjected with cRNAs of SNARE components (snapin+syntaxin-3+SNAP23) over those injected with AQP2 cRNA alone. It was not enhanced when syntaxin-3 was replaced by syntaxin-4 (snapin+syntaxin-4+SNAP23). On the other hand, the latter combination significantly enhanced the transport activity of the related AQP3 water channel while the presence of syntaxin-3 did not. This AQP-syntaxin interaction agrees with the polarity of these proteins' expression in the inner medullary collecting duct epithelium. Thus our findings suggest a selectivity of interactions between different aquaporin and syntaxin isoforms, and thus in the regulation of AQP2 and AQP3 activities in the plasma membrane. Snapin plays an important role as a linker between the water channel and the t-SNARE complex, leading to the fusion event, and the pairing with specific t-SNAREs is essential for the specificity of membrane recognition and fusion. PMID:19515809

  9. Aqueous extract of Costus arabicus inhibits calcium oxalate crystal growth and adhesion to renal epithelial cells.

    PubMed

    de Cógáin, Mitra R; Linnes, Michael P; Lee, Hyo Jung; Krambeck, Amy E; de Mendonça Uchôa, Julio Cezar; Kim, Sung-Hoon; Lieske, John C

    2015-04-01

    Costus arabicus L. (C. arabicus) is a plant used in Brazilian folk medicine to treat urolithiasis; however, its mechanism of action is unclear. The interaction between calcium oxalate (CaOx) crystals and the renal epithelium is important in calculogenesis, and compounds that modulate this process represent candidate therapeutic agents for stone prevention. Therefore, we assessed the inhibitory activity of C. arabicus on CaOx crystallization and the interaction of CaOx crystals with the renal epithelium. A seeded CaOx monohydrate (COM) crystallization system was used to study the effect of C. arabicus on crystal growth. Madin Darby canine kidney (MDCK) cells were used to study [(14)C] COM crystal adhesion in the presence and absence of an aqueous extract of C. arabicus. Cytotoxicity was assessed using a tetrazolium (MTS) cell proliferation assay. Aqueous extracts of C. arabicus decreased crystal growth in a concentration-dependent fashion. Precoating crystals with C. arabicus extract prevented their adhesion to MDCK cells, while pretreating cells did not show any effect. The extract was non-cytotoxic in concentrations of at least 1 mg/ml, which is likely above concentrations achievable in the urine following oral ingestion and excretion. No inhibitory activity was found in hexane, methyl chloride, n-butanol and ethyl acetate fractions of an ethanol extract of the herb. An aqueous extract of C. arabicus may disrupt calculogenesis by interacting with CaOx crystal surfaces. Activity was present in the aqueous extract; therefore, this agent may be bioavailable when administered orally. Fractionation results suggest that the active agent might be a polar polysaccharide. Further identification and characterization along these lines may be warranted. PMID:25652357

  10. Mutations in the Cytoplasmic Domain of the Newcastle Disease Virus Fusion Protein Confer Hyperfusogenic Phenotypes Modulating Viral Replication and Pathogenicity

    PubMed Central

    Samal, Sweety; Khattar, Sunil K.; Paldurai, Anandan; Palaniyandi, Senthilkumar; Zhu, Xiaoping; Collins, Peter L.

    2013-01-01

    The Newcastle disease virus (NDV) fusion protein (F) mediates fusion of viral and host cell membranes and is a major determinant of NDV pathogenicity. In the present study, we demonstrate the effects of functional properties of F cytoplasmic tail (CT) amino acids on virus replication and pathogenesis. Out of a series of C-terminal deletions in the CT, we were able to rescue mutant viruses lacking two or four residues (rΔ2 and rΔ4). We further rescued viral mutants with individual amino acid substitutions at each of these four terminal residues (rM553A, rK552A, rT551A, and rT550A). In addition, the NDV F CT has two conserved tyrosine residues (Y524 and Y527) and a dileucine motif (LL536-537). In other paramyxoviruses, these residues were shown to affect fusion activity and are central elements in basolateral targeting. The deletion of 2 and 4 CT amino acids and single tyrosine substitution resulted in hyperfusogenic phenotypes and increased viral replication and pathogenesis. We further found that in rY524A and rY527A viruses, disruption of the targeting signals did not reduce the expression on the apical or basolateral surface in polarized Madin-Darby canine kidney cells, whereas in double tyrosine mutant, it was reduced on both the apical and basolateral surfaces. Interestingly, in rL536A and rL537A mutants, the F protein expression was more on the apical than on the basolateral surface, and this effect was more pronounced in the rL537A mutant. We conclude that these wild-type residues in the NDV F CT have an effect on regulating F protein biological functions and thus modulating viral replication and pathogenesis. PMID:23843643

  11. Regulation of the Axillary Osmidrosis-Associated ABCC11 Protein Stability by N-Linked Glycosylation: Effect of Glucose Condition.

    PubMed

    Toyoda, Yu; Takada, Tappei; Miyata, Hiroshi; Ishikawa, Toshihisa; Suzuki, Hiroshi

    2016-01-01

    ATP-binding cassette C11 (ABCC11) is a plasma membrane protein involved in the transport of a variety of lipophilic anions. ABCC11 wild-type is responsible for the high-secretion phenotypes in human apocrine glands, such as that of wet-type ear wax, and the risk of axillary osmidrosis. We have previously reported that mature ABCC11 is a glycoprotein containing two N-linked glycans at Asn838 and Asn844. However, little is known about the role of N-linked glycosylation in the regulation of ABCC11 protein. In the current study, we investigated the effects of N-linked glycosylation on the protein level and localization of ABCC11 using polarized Madin-Darby canine kidney II cells. When the N-linked glycosylation in ABCC11-expressing cells was chemically inhibited by tunicamycin treatment, the maturation of ABCC11 was suppressed and its protein level was significantly decreased. Immunoblotting analyses demonstrated that the protein level of the N-linked glycosylation-deficient mutant (N838Q and N844Q: Q838/844) was about half of the ABCC11 wild-type level. Further biochemical studies with the Q838/844 mutant showed that this glycosylation-deficient ABCC11 was degraded faster than wild-type probably due to the enhancement of the MG132-sensitive protein degradation pathway. Moreover, the incubation of ABCC11 wild-type-expressing cells in a low-glucose condition decreased mature, glycosylated ABCC11, compared with the high-glucose condition. On the other hand, the protein level of the Q838/844 mutant was not affected by glucose condition. These results suggest that N-linked glycosylation is important for the protein stability of ABCC11, and physiological alteration in glucose may affect the ABCC11 protein level and ABCC11-related phenotypes in humans, such as axillary osmidrosis. PMID:27281343

  12. Evaluation of the bioavailability of major withanolides of Withania somnifera using an in vitro absorption model system

    PubMed Central

    Devkar, Santosh T.; Kandhare, Amit D.; Sloley, Brian D.; Jagtap, Suresh D.; Lin, James; Tam, Yun K.; Katyare, Surendra S.; Bodhankar, Subhash L.; Hegde, Mahabaleshwar V.

    2015-01-01

    Withania somnifera (L.) Dunal, shows several pharmacological properties which are attributed mainly to the withanolides present in the root. The efficacy of medicinally active withanolides constituents depends on the absorption and transportation through the intestinal epithelium. We examined these characteristics by employing the Sino-Veda Madin-Darby canine kidney cells culture system, which under in vitro condition shows the absorption characteristics similar to the human intestinal epithelium. Thus, the aim of the present investigation was to assess the bioavailability of individual withanolides. Withanolides were diluted in Hank's buffered saline at a concentration of 2 μg/ml were tested for permeability studies carried out for 1 h duration. Permeability was measured in terms of efflux pump (Peff) in cm/s. Peff values of withanolide A (WN A), withanone (WNN), 1,2-deoxywithastramonolide (1,2 DWM), withanolide B (WN B), withanoside IV-V (WS IV-V), and withaferin A were 4.05 × 10−5, 2.06 × 10−5, 1.97 × 10−5, 1.80 × 10−5, 3.19 × 10−6, 3.03 × 10−6 and 3.30 × 10−7 respectively. In conclusion, the nonpolar and low molecular weight compounds (WN A, WNN, 1,2 DWM, and WN B) were highly permeable. As against this, the glycosylated and polar WS IV and WS V showed low permeability. Surprisingly and paradoxically, the highly biologically active withaferin A was completely impermeable, suggesting that further studies possibly using human epithelial colorectal adenocarcinoma (Caco-2) cells may be needed to delineate the absorption characteristics of withanolides, especially withaferin A. PMID:26605156

  13. Lymphocytes accelerate epithelial tight junction assembly: role of AMP-activated protein kinase (AMPK).

    PubMed

    Tang, Xiao Xiao; Chen, Hao; Yu, Sidney; Zhang, Li; Caplan, Michael J; Chan, Hsiao Chang

    2010-01-01

    The tight junctions (TJs), characteristically located at the apicolateral borders of adjacent epithelial cells, are required for the proper formation of epithelial cell polarity as well as for sustaining the mucosal barrier to the external environment. The observation that lymphocytes are recruited by epithelial cells to the sites of infection [1] suggests that they may play a role in the modulation of epithelial barrier function and thus contribute to host defense. To test the ability of lymphocytes to modulate tight junction assembly in epithelial cells, we set up a lymphocyte-epithelial cell co-culture system, in which Madin-Darby canine kidney (MDCK) cells, a well-established model cell line for studying epithelial TJ assembly [2], were co-cultured with mouse lymphocytes to mimic an infection state. In a typical calcium switch experiment, the TJ assembly in co-culture was found to be accelerated compared to that in MDCK cells alone. This accelaration was found to be mediated by AMP-activated protein kinase (AMPK). AMPK activation was independent of changes in cellular ATP levels but it was found to be activated by the pro-inflammatory cytokine TNF-alpha. Forced suppression of AMPK, either with a chemical inhibitor or by knockdown, abrogated the accelerating effect of lymphocytes on TJ formation. Similar results were also observed in a co-culture with lymphocytes and Calu-3 human airway epithelial cells, suggesting that the activation of AMPK may be a general mechanism underlying lymphocyte-accelerated TJ assembly in different epithelia. These results suggest that signals from lymphocytes, such as cytokines, facilitate TJ assembly in epithelial cells via the activation of AMPK. PMID:20808811

  14. The microenvironmental determinants for kidney epithelial cyst morphogenesis

    PubMed Central

    Guo, Qiusha; Xia, Bing; Moshiach, Simon; Xu, Congfeng; Jiang, Yongde; Chen, Yuanjian; Sun, Yao; Lahti, Jill M.; Zhang, Xin A.

    2011-01-01

    Although epithelial morphogenesis is tightly controlled by intrinsic genetic programs, the microenvironment in which epithelial cells proliferate and differentiate also contributes to the morphogenetic process. The roles of the physical microenvironment in epithelial morphogenesis, however, have not been well dissected. In this study, we assessed the impact of the microenvironment on epithelial cyst formation, which often marks the beginning or end step of morphogenesis of epithelial tissues and the pathological characteristic of some diseases. Previous studies have demonstrated that Madin-Darby canine kidney (MDCK) epithelial cells form cysts when grown in a three-dimensional (3D) extracellullar matrix (ECM) environment. We have now further demonstrated that the presence of ECM in the 3D scaffold is required for the formation of properly polarized cysts. Also, we have found that the full interface of epithelial cells with the ECM environment (in-3D) is not essential for cyst formation, since partial contact (on-3D) is sufficient to induce cystogenesis. In addition, we have defined the minimal ECM environment or the physical threshold for cystogenesis under the on-3D condition. Only above the threshold can the morphological cues from the ECM environment induce cyst formation. Moreover, cyst formation under the on-3D condition described in this study actually defines a novel and more feasible model to analyze in vitro morphogenesis. Finally, we have found that, during cystogenesis, MDCK cells generate basal microprotrusions and produce vesicle-like structures to the basal extracellular space, which are specific to and correlated with cyst formation. For the first time, we have systematically elucidated the microenvironmental determinants for epithelial cystogenesis. PMID:18191498

  15. Structural determinants of peripheral O-arylcarbamate FAAH inhibitors render them dual substrates for Abcb1 and Abcg2 and restrict their access to the brain

    PubMed Central

    Moreno-Sanz, Guillermo; Barrera, Borja; Armirotti, Andrea; Bertozzi, Sine M.; Scarpelli, Rita; Bandiera, Tiziano; Prieto, Julio G.; Duranti, Andrea; Tarzia, Giorgio; Merino, Gracia

    2014-01-01

    The blood-brain barrier (BBB) is the main entry route for chemicals into the mammalian central nervous system (CNS). Two transmembrane transporters of the ATP-binding cassette (ABC) family – Breast Cancer Resistance Protein (ABCG2 in humans, Abcg2 in rodents) and P-glycoprotein (ABCB1 in humans, Abcb1 in rodents) – play a key role in mediating this process. Pharmacological and genetic evidence suggests that Abcg2 prevents CNS access to a group of highly potent and selective O-arylcarbamate fatty-acid amidohydrolase (FAAH) inhibitors, which include the compound URB937 (cyclohexylcarbamic acid 3′-carbamoyl-6-hydroxybiphenyl-3-yl ester). To define structure-activity relationships of the interaction of these molecules with Abcg2, in the present study we tested various peripherally restricted and non-restricted O-arylcarbamate FAAH inhibitors for their ability to serve as transport substrates in monolayer cultures of Madin-Darby Canine Kidney-II (MDCKII) cells over-expressing Abcg2. Surprisingly, we found that the majority of compounds tested – even those able to enter the CNS in vivo – were substrates for Abcg2 in vitro. Additional experiments in MDCKII cells overexpressing ABCB1 revealed that only those compounds that were dual substrates for ABCB1 and Abcg2 in vitro were also peripherally restricted in vivo. The extent of such restriction seems to depend upon other physicochemical features of the compounds, in particular the polar surface area. Consistent with these in vitro results, we found that URB937 readily enters the brain in dual knockout mice lacking both Abcg2 and Abcb1, whereas it is either partially or completely excluded from the brain of mice lacking either transporter alone. The results suggest that Abcg2 and Abcb1 act together to restrict the access of URB937 to the CNS. PMID:24993496

  16. Vaccines for Canine Leishmaniasis

    PubMed Central

    Foroughi-Parvar, Faeze; Hatam, Gholamreza

    2014-01-01

    Leishmania infantum is the obligatory intracellular parasite of mammalian macrophages and causes zoonotic visceral leishmaniasis (ZVL). The presence of infected dogs as the main reservoir host of ZVL is regarded as the most important potential risk for human infection. Thus the prevention of canine visceral leishmaniasis (CVL) is essential to stop the current increase of the Mediterranean visceral leishmaniasis. Recently considerable advances in achieving protective immunization of dogs and several important attempts for achieving an effective vaccine against CVL lead to attracting the scientists trust in its important role for eradication of ZVL. This paper highlights the recent advances in vaccination against canine visceral leishmaniasis from 2007 until now. PMID:25628897

  17. 9 CFR 113.305 - Canine Hepatitis and Canine Adenovirus Type 2 Vaccine.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 9 Animals and Animal Products 1 2011-01-01 2011-01-01 false Canine Hepatitis and Canine Adenovirus... STANDARD REQUIREMENTS Live Virus Vaccines § 113.305 Canine Hepatitis and Canine Adenovirus Type 2 Vaccine. Canine Hepatitis Vaccine and Canine Adenovirus Type 2 Vaccine shall be prepared from virus-bearing...

  18. 9 CFR 113.202 - Canine Hepatitis and Canine Adenovirus Type 2 Vaccine, Killed Virus.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 9 Animals and Animal Products 1 2010-01-01 2010-01-01 false Canine Hepatitis and Canine Adenovirus...; ORGANISMS AND VECTORS STANDARD REQUIREMENTS Killed Virus Vaccines § 113.202 Canine Hepatitis and Canine Adenovirus Type 2 Vaccine, Killed Virus. Canine Hepatitis and Canine Adenovirus Type 2 Vaccine, Killed...

  19. 9 CFR 113.305 - Canine Hepatitis and Canine Adenovirus Type 2 Vaccine.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 9 Animals and Animal Products 1 2010-01-01 2010-01-01 false Canine Hepatitis and Canine Adenovirus... STANDARD REQUIREMENTS Live Virus Vaccines § 113.305 Canine Hepatitis and Canine Adenovirus Type 2 Vaccine. Canine Hepatitis Vaccine and Canine Adenovirus Type 2 Vaccine shall be prepared from virus-bearing...

  20. 9 CFR 113.202 - Canine Hepatitis and Canine Adenovirus Type 2 Vaccine, Killed Virus.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 9 Animals and Animal Products 1 2011-01-01 2011-01-01 false Canine Hepatitis and Canine Adenovirus...; ORGANISMS AND VECTORS STANDARD REQUIREMENTS Killed Virus Vaccines § 113.202 Canine Hepatitis and Canine Adenovirus Type 2 Vaccine, Killed Virus. Canine Hepatitis and Canine Adenovirus Type 2 Vaccine, Killed...

  1. Vaccines for Canine Leishmaniasis

    PubMed Central

    Palatnik-de-Sousa, Clarisa B.

    2012-01-01

    Leishmaniasis is the third most important vector-borne disease worldwide. Visceral leishmaniasis (VL) is a severe and frequently lethal protozoan disease of increasing incidence and severity due to infected human and dog migration, new geographical distribution of the insect due to global warming, coinfection with immunosuppressive diseases, and poverty. The disease is an anthroponosis in India and Central Africa and a canid zoonosis (ZVL) in the Americas, the Middle East, Central Asia, China, and the Mediterranean. The ZVL epidemic has been controlled by one or more measures including the culling of infected dogs, treatment of human cases, and insecticidal treatment of homes and dogs. However, the use of vaccines is considered the most cost–effective control tool for human and canine disease. Since the severity of the disease is related to the generation of T-cell immunosuppression, effective vaccines should be capable of sustaining or enhancing the T-cell immunity. In this review we summarize the clinical and parasitological characteristics of ZVL with special focus on the cellular and humoral canine immune response and review state-of-the-art vaccine development against human and canine VL. Experimental vaccination against leishmaniasis has evolved from the practice of leishmanization with living parasites to vaccination with crude lysates, native parasite extracts to recombinant and DNA vaccination. Although more than 30 defined vaccines have been studied in laboratory models no human formulation has been licensed so far; however three second-generation canine vaccines have already been registered. As expected for a zoonotic disease, the recent preventive vaccination of dogs in Brazil has led to a reduction in the incidence of canine and human disease. The recent identification of several Leishmania proteins with T-cell epitopes anticipates development of a multiprotein vaccine that will be capable of protecting both humans and dogs against VL. PMID:22566950

  2. The Canine Oral Microbiome

    PubMed Central

    Dewhirst, Floyd E.; Klein, Erin A.; Thompson, Emily C.; Blanton, Jessica M.; Chen, Tsute; Milella, Lisa; Buckley, Catherine M. F.; Davis, Ian J.; Bennett, Marie-Lousie; Marshall-Jones, Zoe V.

    2012-01-01

    Determining the bacterial composition of the canine oral microbiome is of interest for two primary reasons. First, while the human oral microbiome has been well studied using molecular techniques, the oral microbiomes of other mammals have not been studied in equal depth using culture independent methods. This study allows a comparison of the number of bacterial taxa, based on 16S rRNA-gene sequence comparison, shared between humans and dogs, two divergent mammalian species. Second, canine oral bacteria are of interest to veterinary and human medical communities for understanding their roles in health and infectious diseases. The bacteria involved are mostly unnamed and not linked by 16S rRNA-gene sequence identity to a taxonomic scheme. This manuscript describes the analysis of 5,958 16S rRNA-gene sequences from 65 clone libraries. Full length 16S rRNA reference sequences have been obtained for 353 canine bacterial taxa, which were placed in 14 bacterial phyla, 23 classes, 37 orders, 66 families, and 148 genera. Eighty percent of the taxa are currently unnamed. The bacterial taxa identified in dogs are markedly different from those of humans with only 16.4% of oral taxa are shared between dogs and humans based on a 98.5% 16S rRNA sequence similarity cutoff. This indicates that there is a large divergence in the bacteria comprising the oral microbiomes of divergent mammalian species. The historic practice of identifying animal associated bacteria based on phenotypic similarities to human bacteria is generally invalid. This report describes the diversity of the canine oral microbiome and provides a provisional 16S rRNA based taxonomic scheme for naming and identifying unnamed canine bacterial taxa. PMID:22558330

  3. Canine degenerative myelopathy.

    PubMed

    Coates, Joan R; Wininger, Fred A

    2010-09-01

    Canine degenerative myelopathy (DM) is an adult-onset fatal neurodegenerative disease that occurs in many breeds. The initial upper motor neuron spastic paraparesis and general proprioceptive ataxia in the pelvic limbs progress to a flaccid lower motor neuron tetraparesis. Recently, a missense mutation in the superoxide dismutase 1 (SOD1) gene was found to be a risk factor for DM, suggesting that DM is similar to some forms of human amyotrophic lateral sclerosis (ALS or Lou Gehrig's disease). This article reviews the current knowledge of canine DM with regard to its signalment, clinical spectrum, diagnostic approach, and treatment. The implications of the SOD1 mutation on both diseases are discussed, comparing pathogenic mechanisms while conveying perspectives to translational medicine. PMID:20732599

  4. Control of canine distemper.

    PubMed

    Chappuis, G

    1995-05-01

    Control of canine distemper can realistically only be achieved by the use of vaccination. The types of vaccine in current use are described, together with some of the problems encountered such as interference by maternal antibodies, and usage in species other than dogs. Modified live viral vaccines, as used for more than thirty years, have proved very effective. Nevertheless there is scope for some improvement in vaccine efficacy and recent developments in genetic recombinant methods are described. PMID:8588329

  5. Canine ehrlichiosis in Connecticut.

    PubMed Central

    Magnarelli, L A; Litwin, H J; Holland, C J; Anderson, J F; Ristic, M

    1990-01-01

    The first case of canine ehrlichiosis in Connecticut is reported. A female Brittany spaniel from Milford presented with lethargy, anorexia, fever, petechiae, splenomegaly, thrombocytopenia, anemia, elevated serum alkaline phosphatase, lymphopenia, and hypoalbuminemia. Serologic analysis revealed antibodies to Ehrlichia canis (titer, 1:2,560). This documents a more northern geographic distribution in the United States for this infectious agent than had previously been suspected. PMID:2312682

  6. American canine hepatozoonosis.

    PubMed

    Panciera, R J; Ewing, S A

    2003-06-01

    American canine hepatozoonosis is an emerging, tick-transmitted infection of domestic dogs caused by a recently recognized species of apicomplexan parasite, Hepatozoon americanum. The known definitive host of the protozoan is the Gulf Coast tick, Amblyomma maculatum. Presently recognized intermediate hosts include the domestic dog and the coyote, Canis latrans. Laboratory-reared larval or nymphal A. maculatum can be infected readily by feeding to repletion on a parasitemic intermediate host; sporogony requires 35-40 days. Transmission of infection to the dog has been produced experimentally by oral administration of mature oocysts or oocyst-containing ticks. Canine disease follows experimental exposure in 4-6 weeks and is characterized by systemic illness, extreme neutrophilic leukocytosis, muscle and bone pain, and proliferation of periosteal bone. Histopathological findings include multifocal skeletal and cardiac myositis associated with escape of mature merozoites from within the host-cell environment. There is also rapid onset of periosteal activation and osteogenesis and, less frequently, glomerulopathy and amyloidosis. Sequential stages of development of H. americanum in both the dog and the tick have been elucidated. Gamonts potentially infectious to ticks have been observed in peripheral blood leukocytes of the dog in as few as 28 days after exposure to oocysts. Young coyotes experimentally exposed to a canine strain of H. americanum acquired disease indistinguishable from that of similarly exposed young dogs. PMID:12885206

  7. Canine mast cell tumors.

    PubMed

    Macy, D W

    1985-07-01

    Despite the fact that the mast cell tumor is a common neoplasm of the dog, we still have only a meager understanding of its etiology and biologic behavior. Many of the published recommendations for treatment are based on opinion rather than facts derived from careful studies and should be viewed with some skepticism. Because of the infrequent occurrence of this tumor in man, only a limited amount of help can be expected from human oncologists; therefore, burden of responsibility for progress in predicting behavior and developing treatment effective for canine mast cell tumors must fall on the shoulders of the veterinary profession. PMID:3929444

  8. Brazilian canine hepatozoonosis.

    PubMed

    O'Dwyer, Lucia Helena

    2011-01-01

    The genus Hepatozoon includes hundreds of species that infect birds, reptiles, amphibians and mammals, in all continents with tropical and subtropical climates. Two species have been described in domestic dogs: H. canis, reported in Europe, Asia, Africa, South America and the United States; and H. americanum, which so far has only been diagnosed in the United States. In Brazil, the only species found infecting dogs is H. canis. The objective of this review was to detail some aspects of canine hepatozoonosis, caused by H. canis, and the main points of its biology, transmission, pathogenicity, symptoms, epidemiology and diagnostic methods, with emphasis on research developed in Brazil. PMID:21961746

  9. Absence of canine papillomavirus sequences in canine mammary tumours.

    PubMed

    Sardon, D; Blundell, R; Burrai, G P; Alberti, A; Tore, G; Passino, E Sanna; Antuofermo, E

    2015-01-01

    Human papillomaviruses (PVs) are found in human breast cancer tissue; however, it remains controversial as to whether these viruses play a role in the aetiology of this tumour. There has been minimal study of whether PVs are found in normal or abnormal mammary glands of animals. The present study investigated whether a PV sequence could be found in the mammary glands of 33 female dogs by rolling circle amplification and polymerase chain reaction. No PV DNA was found in normal or neoplastic canine mammary tissues, suggesting that canine PVs are probably not involved in the pathogenesis of canine mammary neoplasia. PMID:25435511

  10. Canine rickettsial infections.

    PubMed

    Stiles, J

    2000-09-01

    Dogs that live in tick-infested areas are at risk for contracting rickettsial infections. Clinical signs associated with ehrlichiosis or Rocky Mountain spotted fever may be dramatic or mild. Clinicians must consider the possibility of rickettsial diseases to request laboratory tests that will permit a proper diagnosis. Specific antimicrobial therapy usually brings about clinical improvement, although some dogs may not be cleared of rickettsial organisms, even with prolonged treatment. A small percentage of dogs die of rickettsial infections, either in the acute stage or owing to chronic bone marrow suppression and generalized debilitation. Ocular lesions are an important clinical sign in canine rickettsial infections and may aid the clinician in making a diagnosis and monitoring response to therapy. PMID:11033879

  11. Canine cutaneous leishmaniasis.

    PubMed

    Sasani, F; Javanbakht, J; Samani, R; Shirani, D

    2016-03-01

    Canine cutaneous leishmaniasis (CCL) is a significant veterinary problem. Infected dogs also serve as parasite reservoirs and contribute to human transmission of cutaneous leishmaniasis. Histologically, the lesions were nodular to diffuse interstitial granulomatous dermatitis with histiocytic pseudorosettes together with numerous amastigotes within macrophages and occasionally within the interstitium. Organisms were often contained within clear and intracellular vacuoles. The other inflammatory cells, which were present in the biopsies of the Leishmania-infected dog, were lymphocytes and plasma cells. The histopathology results emphasized the role of dog, particularly asymptomatic dog, as reservoirs for CCL because of the high cutaneous parasite loads. These results may help to explain the maintenance of high transmission rates and numbers of CCL cases in endemic urban regions. PMID:27065598

  12. Bactericidal effects of antimicrobial agents on epithelial cell-associated Pseudomonas aeruginosa.

    PubMed

    Hirakata, Yoichi; Yano, Hisakazu; Arai, Kazuaki; Kitagawa, Miho; Hatta, Masumitsu; Kunishima, Hiroyuki; Kaku, Mitsuo

    2012-06-01

    It is not clear whether antipseudomonal agents can kill cell-associated bacteria within a short time. Madin-Darby canine kidney (MDCK) and A549 cells were infected with Pseudomonas aeruginosa ATCC 27853 and PAO1 and the bactericidal activity of ceftazidime, imipenem, meropenem, gentamicin, and ciprofloxacin against the organisms was investigated. In both MDCK and A549 cells, β-lactams could not kill epithelial cell-associated bacteria within 2 h. Gentamicin at concentrations ≤32 μg/ml killed more than 99% of epithelial cell-associated bacteria. Ciprofloxacin at 0.5 μg/ml killed more than 99.9% of MDCK cell-associated bacteria. Ciprofloxacin has the strongest and most rapid bactericidal activity against epithelial cell-associated bacteria, which may be explained by the combination of potent in-vitro bactericidal activity and high penetration ability into epithelial cells. PMID:22116462

  13. Synthesis and antiviral activity of 5'-deoxypyrazofurin.

    PubMed

    Chen, X; Schneller, S W; Ikeda, S; Snoeck, R; Andrei, G; Balzarini, J; De Clercq, E

    1993-11-12

    In searching for derivatives of pyrazofurin that could display antiviral properties by means that do not require C-5' phosphorylation, 5'-deoxypyrazofurin (3) has been synthesized in six steps from methyl5-deoxy-2,3-O-isopropylidene-beta-D-ribofuranoside (4). Compound 3 was evaluated for antiviral activity against a large number of viruses including herpes-, pox-, myxo-, toga-, arena-, rhabdo-, picorna-,reo-, and retroviruses. Compound 3 proved active against respiratory syncytial virus (in HeLa cells), vaccinia virus (in embryonic skin-muscle fibroblast cells), vesicular stomatitis virus (in HeLa cells), and influenza A virus (in Madin-Darby canine kidney cells) at concentrations (ranging from 4 to 20 micrograms/mL) that were nontoxic to the confluent host cell cultures. PMID:8246242

  14. Short communication: antiviral activity of subcritical water extract of Brassica juncea against influenza virus A/H1N1 in nonfat milk.

    PubMed

    Lee, N-K; Lee, J-H; Lim, S-M; Lee, K A; Kim, Y B; Chang, P-S; Paik, H-D

    2014-09-01

    Subcritical water extract (SWE) of Brassica juncea was studied for antiviral effects against influenza virus A/H1N1 and for the possibility of application as a nonfat milk supplement for use as an "antiviral food." At maximum nontoxic concentrations, SWE had higher antiviral activity against influenza virus A/H1N1 than n-hexane, ethanol, or hot water (80°C) extracts. Addition of 0.5mg/mL of B. juncea SWE to culture medium led to 50.35% cell viability (% antiviral activity) for Madin-Darby canine kidney cells infected with influenza virus A/H1N1. Nonfat milk supplemented with 0.28mg/mL of B. juncea SWE showed 39.62% antiviral activity against influenza virus A/H1N1. Thus, the use of B. juncea SWE as a food supplement might aid in protection from influenza viral infection. PMID:25022686

  15. Comparison of large-scale mammalian cell culture systems with egg culture for the production of influenza virus A vaccine strains.

    PubMed

    Tree, J A; Richardson, C; Fooks, A R; Clegg, J C; Looby, D

    2001-05-14

    Different types of microcarriers were assessed for the large-scale culture of influenza virus in the Madin-Darby canine kidney (MDCK) cells. Both porous and solid carriers were examined. A higher titre of influenza A/PR8/34 virus was recovered from cultures using solid (1.3x10(9) PFU per ml) rather than porous carriers (4.0x10(8) PFU per ml). High titres of virus (1.0x10(9) PFU per ml) were also obtained from roller bottle cultures of MDCK cells and the traditional culture technique using embryonated hens' eggs (3.9x10(9) PFU per ml). We found that solid carriers composed of dextran with a positive charge are the most suitable carriers for the large-scale growth of influenza A virus in MDCK cells using serum-free media. PMID:11348709

  16. Knockdown of Sec6 improves cell-cell adhesion by increasing α-E-catenin in oral cancer cells.

    PubMed

    Tanaka, Toshiaki; Iino, Mituyoshi; Goto, Kaoru

    2012-03-23

    The Sec6/8 complex is essential for specific exocytic sites on the plasma membrane and contributes to membrane growth in mammalian cells. In Madin-Darby canine kidney (MDCK) cells, E-cadherin and nectin-based adhesion complexes recruit the Sec6/8 complex to intercellular contacts. However, in cancer cells, the relationship between the Sec6/8 complex and the cell-cell adhesion proteins remains obscure. We demonstrate that the expression of α-E-catenin is increased by Sec6 siRNAs, and E-cadherin and β-catenin localize mainly at the cell-cell contact region in HSC3 cells, which were transfected with Sec6 siRNA. PMID:22381337

  17. Anti-H5N1 virus flavonoids from Capparis sinaica Veill.

    PubMed

    Ibrahim, Amany K; Youssef, Ahmed I; Arafa, Abdel Satar; Ahmed, Safwat A

    2013-01-01

    Methanolic extract of Capparis sinaica Veill was tested for its in vitro antiviral activity against highly pathogenic avian influenza strain H5N1 using plaque inhibition assay in Madin-Darby canine kidney. The results indicated that the extract possessed potent antiviral activity (100% inhibition at the concentration of 1 μg/ml). Based on this result, C. sinaica Veill was selected for further study by applying bioactivity-guided fractionation to isolate its antiviral principles. The fractions eluted with EtOAc and 25% MeOH in EtOAc were found to hold the antiviral activity. Further chromatographic separation of the fractions holding the antiviral activity led to the isolation of quercetin (1), isoquercetin (2) and rutin (3) for the first time from this species. The isolates showed reduction in the virus titre by 68.13%, 79.66% and 73.22% inhibition at a concentration of 1 ng/ml, respectively. PMID:23651316

  18. Resistance of ovine, caprine and bovine endothelial cells to Clostridium perfringens type D epsilon toxin in vitro.

    PubMed

    Uzal, F A; Rolfe, B E; Smith, N J; Thomas, A C; Kelly, W R

    1999-08-01

    Ovine, caprine and bovine endothelial cells were grown in vitro and challenged with Clostridium perfringens type D epsilon toxin to compare their susceptibility to this toxin. Madin Darby canine kidney (MDCK) cells, which are known to be susceptible to epsilon toxin, were used as a positive control. No morphological alterations were observed in any of the endothelial cell cultures tested, even after challenging with doses as high as 1200 MLD50/ml of epsilon toxin. MDCK cells showed contour rounding and nuclear condensation as early as 30 min after exposure to 100 MLD50/ml of epsilon toxin and after 60 min of exposure to 12.5 MLD50/ml of the same toxin. All the MDCK cells were dead after 3 h of exposure to all concentrations of epsilon toxin. The results indicate that ovine, caprine and bovine endothelial cells are not morphologically responsive to the action of epsilon toxin in vitro. PMID:10493114

  19. Inhibitors of influenza viruses replication: a patent evaluation (WO2013019828).

    PubMed

    Xie, Yuanchao; Song, Weiguo; Xiao, Weidong; Gu, Changjuan; Xu, Wenfang

    2013-11-01

    A series of compounds incorporating two aromatic heterocycles were prepared as inhibitors of influenza virus replication in the patent. Some of them presented potent activity against influenza virus in Madin-Darby canine kidney (MDCK) cells and in influenza therapeutic mouse model. These compounds in the patent were also defined to be pharmaceutically acceptable salts and pharmaceutical compositions that were claimed to be useful for treating influenza. In view of the threat of influenza pandemic, it is necessary to discover new anti-influenza drugs. Although there is a lack of essential biological data and the molecular mechanisms are not clear, these compounds with potent antiviral activity stand for a new type of anti-influenza agents and deserve further studies. PMID:23967861

  20. Canine leishmaniosis in South America

    PubMed Central

    Dantas-Torres, Filipe

    2009-01-01

    Canine leishmaniosis is widespread in South America, where a number of Leishmania species have been isolated or molecularly characterised from dogs. Most cases of canine leishmaniosis are caused by Leishmania infantum (syn. Leishmania chagasi) and Leishmania braziliensis. The only well-established vector of Leishmania parasites to dogs in South America is Lutzomyia longipalpis, the main vector of L. infantum, but many other phlebotomine sandfly species might be involved. For quite some time, canine leishmaniosis has been regarded as a rural disease, but nowadays it is well-established in large urbanised areas. Serological investigations reveal that the prevalence of anti-Leishmania antibodies in dogs might reach more than 50%, being as high as 75% in highly endemic foci. Many aspects related to the epidemiology of canine leishmaniosis (e.g., factors increasing the risk disease development) in some South American countries other than Brazil are poorly understood and should be further studied. A better understanding of the epidemiology of canine leishmaniosis in South America would be helpful to design sustainable control and prevention strategies against Leishmania infection in both dogs and humans. PMID:19426440

  1. BRAF Mutations in Canine Cancers

    PubMed Central

    Mochizuki, Hiroyuki; Kennedy, Katherine; Shapiro, Susan G.; Breen, Matthew

    2015-01-01

    Activating mutations of the BRAF gene lead to constitutive activation of the MAPK pathway. Although many human cancers carry the mutated BRAF gene, this mutation has not yet been characterized in canine cancers. As human and canine cancers share molecular abnormalities, we hypothesized that BRAF gene mutations also exist in canine cancers. To test this hypothesis, we sequenced the exon 15 of BRAF, mutation hot spot of the gene, in 667 canine primary tumors and 38 control tissues. Sequencing analysis revealed that a single nucleotide T to A transversion at nucleotide 1349 occurred in 64 primary tumors (9.6%), with particularly high frequency in prostatic carcinoma (20/25, 80%) and urothelial carcinoma (30/45, 67%). This mutation results in the amino acid substitution of glutamic acid for valine at codon 450 (V450E) of canine BRAF, corresponding to the most common BRAF mutation in human cancer, V600E. The evolutional conservation of the BRAF V600E mutation highlights the importance of MAPK pathway activation in neoplasia and may offer opportunity for molecular diagnostics and targeted therapeutics for dogs bearing BRAF-mutated cancers. PMID:26053201

  2. Sodium-Calcium Exchanger 1 Regulates Epithelial Cell Migration via Calcium-dependent Extracellular Signal-regulated Kinase Signaling*

    PubMed Central

    Balasubramaniam, Sona Lakshme; Gopalakrishnapillai, Anilkumar; Gangadharan, Vimal; Duncan, Randall L.; Barwe, Sonali P.

    2015-01-01

    Na+/Ca2+ exchanger-1 (NCX1) is a major calcium extrusion mechanism in renal epithelial cells enabling the efflux of one Ca2+ ion and the influx of three Na+ ions. The gradient for this exchange activity is provided by Na,K-ATPase, a hetero-oligomer consisting of a catalytic α-subunit and a regulatory β-subunit (Na,K-β) that also functions as a motility and tumor suppressor. We showed earlier that mice with heart-specific ablation (KO) of Na,K-β had a specific reduction in NCX1 protein and were ouabain-insensitive. Here, we demonstrate that Na,K-β associates with NCX1 and regulates its localization to the cell surface. Madin-Darby canine kidney cells with Na,K-β knockdown have reduced NCX1 protein and function accompanied by 2.1-fold increase in free intracellular calcium and a corresponding increase in the rate of cell migration. Increased intracellular calcium up-regulated ERK1/2 via calmodulin-dependent activation of PI3K. Both myosin light chain kinase and Rho-associated kinase acted as mediators of ERK1/2-dependent migration. Restoring NCX1 expression in β-KD cells reduced migration rate and ERK1/2 activation, suggesting that NCX1 functions downstream of Na,K-β in regulating cell migration. In parallel, inhibition of NCX1 by KB-R7943 in Madin-Darby canine kidney cells, LLC-PK1, and human primary renal epithelial cells (HREpiC) increased ERK1/2 activation and cell migration. This increased migration was associated with high myosin light chain phosphorylation by PI3K/ERK-dependent mechanism in HREpiC cells. These data confirm the role of NCX1 activity in regulating renal epithelial cell migration. PMID:25770213

  3. Inhibition of A/Human/Hubei/3/2005 (H3N2) influenza virus infection by silver nanoparticles in vitro and in vivo

    PubMed Central

    Xiang, Dongxi; Zheng, Yang; Duan, Wei; Li, Xiujing; Yin, Jianjian; Shigdar, Sarah; O’Connor, Michael Liam; Marappan, Manju; Zhao, Xiaojuan; Miao, Yingqiu; Xiang, Bin; Zheng, Conglong

    2013-01-01

    Silver nanoparticles (AgNPs) have attracted much attention as antimicrobial agents and have demonstrated efficient inhibitory activity against various viruses, including human immunodeficiency virus, hepatitis B virus, and Tacaribe virus. In this study, we investigated if AgNPs could have antiviral and preventive effects in A/Human/Hubei/3/2005 (H3N2) influenza virus infection. Madin-Darby canine kidney cells infected with AgNP-treated H3N2 influenza virus showed better viability (P<0.05 versus influenza virus control) and no obvious cytopathic effects compared with an influenza virus control group and a group treated with the solvent used for preparation of the AgNPs. Hemagglutination assay indicated that AgNPs could significantly inhibit growth of the influenza virus in Madin-Darby canine kidney cells (P<0.01 versus the influenza virus control). AgNPs significantly reduced cell apoptosis induced by H3N2 influenza virus at three different treatment pathways (P<0.05 versus influenza virus control). H3N2 influenza viruses treated with AgNPs were analyzed by transmission electron microscopy and found to interact with each other, resulting in destruction of morphologic viral structures in a time-dependent manner in a time range of 30 minutes to 2 hours. In addition, intranasal AgNP administration in mice significantly enhanced survival after infection with the H3N2 influenza virus. Mice treated with AgNPs showed lower lung viral titer levels and minor pathologic lesions in lung tissue, and had a marked survival benefit during secondary intranasal passage in vivo. These results provide evidence that AgNPs have beneficial effects in preventing H3N2 influenza virus infection both in vitro and in vivo, and demonstrate that AgNPs can be used as potential therapeutics for inhibiting outbreaks of influenza. PMID:24204140

  4. Molecular cloning and characterization of canine ICOS.

    PubMed

    Lee, Je-Hwan; Joo, Young-Don; Yim, Daesong; Lee, Richard; Ostrander, Elaine A; Loretz, Carol; Little, Marie-Térèse; Storb, Rainer; Kuhr, Christian S

    2004-10-01

    Inducible costimulatory receptor (ICOS) is one recently identified member of the CD28 family of costimulatory molecules. Evidence suggests ICOS functions as a critical immune regulator and, to evaluate these effects, we employed the canine model system that has been used to develop strategies currently in clinical use for hematopoietic stem cell transplantation. To investigate the effects of blocking the ICOS pathway in the canine hematopoietic cell transplantation model, we tested existing murine and human reagents and cloned the full length of the open reading frame of canine ICOS cDNA to allow the development of reagents specific for the canine ICOS. Canine ICOS contains a major open reading frame of 624 nucleotides, encoding a protein of 208 amino acids, and localizes to chromosome 37. Canine ICOS shares 79% sequence identity with human ICOS, 70% with mouse, and 69% with rat. Canine ICOS expression is limited to stimulated PBMC. PMID:15475250

  5. Canine lymphoma: a review.

    PubMed

    Zandvliet, M

    2016-06-01

    Canine lymphoma (cL) is a common type of neoplasia in dogs with an estimated incidence rate of 20-100 cases per 100,000 dogs and is in many respects comparable to non-Hodgkin lymphoma in humans. Although the exact cause is unknown, environmental factors and genetic susceptibility are thought to play an important role. cL is not a single disease, and a wide variation in clinical presentations and histological subtypes is recognized. Despite this potential variation, most dogs present with generalized lymphadenopathy (multicentric form) and intermediate to high-grade lymphoma, more commonly of B-cell origin. The most common paraneoplastic sign is hypercalcemia that is associated with the T-cell immunophenotype. Chemotherapy is the treatment of choice and a doxorubicin-based multidrug protocol is currently the standard of care. A complete remission is obtained for most dogs and lasts for a median period of 7-10 months, resulting in a median survival of 10-14 months. Many prognostic factors have been reported, but stage, immunophenotype, tumor grade, and response to chemotherapy appear of particular importance. Failure to respond to chemotherapy suggests drug resistance, which can be partly attributed to the expression of drug transporters of the ABC-transporter superfamily, including P-gp and BCRP. Ultimately, most lymphomas will become drug resistant and the development of treatments aimed at reversing drug resistance or alternative treatment modalities (e.g. immunotherapy and targeted therapy) are of major importance. This review aims to summarize the relevant data on cL, as well as to provide an update of the recent literature. PMID:26953614

  6. What's eating you? Canine scabies.

    PubMed

    Burroughs, Richard F; Elston, Dirk M

    2003-08-01

    Infestation with Sarcoptes scabiei var canis, the causative strain of canine scabies, can produce a pruritic rash in humans. The rash generally manifests within 24 to 96 hours of contact with the affected pet. Scrapings are generally negative, and the correct diagnosis requires a high index of suspicion. PMID:12953932

  7. 9 CFR 113.305 - Canine Hepatitis and Canine Adenovirus Type 2 Vaccine.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... dilution in a varying serum-constant virus neutralization test using 50 to 300 TCID50 of canine adenovirus... virus neutralization test using 50 to 300 TCID50 of canine adenovirus. (i) A geometric mean titer of...

  8. 9 CFR 113.305 - Canine Hepatitis and Canine Adenovirus Type 2 Vaccine.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... dilution in a varying serum-constant virus neutralization test using 50 to 300 TCID50 of canine adenovirus... virus neutralization test using 50 to 300 TCID50 of canine adenovirus. (i) A geometric mean titer of...

  9. 9 CFR 113.305 - Canine Hepatitis and Canine Adenovirus Type 2 Vaccine.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... dilution in a varying serum-constant virus neutralization test using 50 to 300 TCID50 of canine adenovirus... virus neutralization test using 50 to 300 TCID50 of canine adenovirus. (i) A geometric mean titer of...

  10. Genetics of Human and Canine Dilated Cardiomyopathy

    PubMed Central

    Simpson, Siobhan; Edwards, Jennifer; Ferguson-Mignan, Thomas F. N.; Cobb, Malcolm; Mongan, Nigel P.; Rutland, Catrin S.

    2015-01-01

    Cardiovascular disease is a leading cause of death in both humans and dogs. Dilated cardiomyopathy (DCM) accounts for a large number of these cases, reported to be the third most common form of cardiac disease in humans and the second most common in dogs. In human studies of DCM there are more than 50 genetic loci associated with the disease. Despite canine DCM having similar disease progression to human DCM studies into the genetic basis of canine DCM lag far behind those of human DCM. In this review the aetiology, epidemiology, and clinical characteristics of canine DCM are examined, along with highlighting possible different subtypes of canine DCM and their potential relevance to human DCM. Finally the current position of genetic research into canine and human DCM, including the genetic loci, is identified and the reasons many studies may have failed to find a genetic association with canine DCM are reviewed. PMID:26266250

  11. Canine mammary tumours, an overview.

    PubMed

    Sleeckx, N; de Rooster, H; Veldhuis Kroeze, E J B; Van Ginneken, C; Van Brantegem, L

    2011-12-01

    Canine mammary tumours (CMTs) are the most common neoplasms in intact female dogs. Although the prevalence of these tumours decreases in regions where preventive ovari(ohyster)ectomy is performed, it remains an important disease entity in veterinary medicine. Moreover, treatment options are limited in comparison with human breast cancer. Nevertheless, recent human treatment protocols might have potential in bitches suffering from CMTs. PMID:21645126

  12. Canine adenovirus based rabies vaccines.

    PubMed

    Tordo, N; Foumier, A; Jallet, C; Szelechowski, M; Klonjkowski, B; Eloit, M

    2008-01-01

    Adenovirus based vectors are very attractive candidates for vaccination purposes as they induce in mammalian hosts potent humoral, mucosal and cellular immune responses to antigens encoded by the inserted genes. We have generated E1-deleted and replication-competent recombinant canine type-2 adenoviruses expressing the rabies virus glycoprotein (G). The effectiveness of both vectors to express a native G protein has been characterized in vitro in permissive cell lines. We compared the humoral and cellular immune responses induced in mice by intramuscular injection of the recombinant canine adenovirus vectors with those induced by a human (Ad5) E1-deleted virus expressing the same rabies G protein. Humoral responses specific to the adenoviruses or the rabies glycoprotein antigens were studied. The influence of the mouse strain was observed using replication-competent canine adenovirus. A high level of rabies neutralizing antibody was observed upon i.m. inoculation, and 100% of mice survived lethal challenge. These results are very promising in the perspective of oral vaccine for dog rabies control. PMID:18634509

  13. Genome Sequence of Canine Herpesvirus

    PubMed Central

    Papageorgiou, Konstantinos V.; Suárez, Nicolás M.; Wilkie, Gavin S.; McDonald, Michael; Graham, Elizabeth M.; Davison, Andrew J.

    2016-01-01

    Canine herpesvirus is a widespread alphaherpesvirus that causes a fatal haemorrhagic disease of neonatal puppies. We have used high-throughput methods to determine the genome sequences of three viral strains (0194, V777 and V1154) isolated in the United Kingdom between 1985 and 2000. The sequences are very closely related to each other. The canine herpesvirus genome is estimated to be 125 kbp in size and consists of a unique long sequence (97.5 kbp) and a unique short sequence (7.7 kbp) that are each flanked by terminal and internal inverted repeats (38 bp and 10.0 kbp, respectively). The overall nucleotide composition is 31.6% G+C, which is the lowest among the completely sequenced alphaherpesviruses. The genome contains 76 open reading frames predicted to encode functional proteins, all of which have counterparts in other alphaherpesviruses. The availability of the sequences will facilitate future research on the diagnosis and treatment of canine herpesvirus-associated disease. PMID:27213534

  14. Canine leishmaniosis - an emerging disease.

    PubMed

    Kaszak, Ilona; Planellas, Marta; Dworecka-Kaszak, Bożena

    2015-01-01

    Canine leishmaniosis (CanL) is an invasive disease of dogs, caused by Leishmania spp. parasites transmitted by the bite of an infected phlebotomine sand fly. CanL is declared an important disease by World Organisation for Animal Health (OIE). Due to its zoonotic potential is of a great importance the prevention of this disease in non endemic areas. Canine leishmaniosis is endemic disease in more than 70 countries and is a common disease in Mediterranean region. Recently, many cases have been reported in non endemic areas, like United Kingdom, Germany and Poland as well, where this disease is considered exotic. The aim of this article is to summarize shortly canine leishmaniosis, it's transmission, clinical manifestations, diagnostics procedure, treatment, prognosis and prevention. Increasing knowledge about this disease can be of a great use for veterinary surgeons from countries where CanL is an emerging disease. Multiple clinical presentations of CanL should aware clinicians to include leishmaniosis in the differential diagnosis of most clinical cases. Unfortunately, even if dogs recover clinically after treatment, complete elimination of Leishmania spp. is rarely achieved, and they remain infected and may relapse. PMID:26342500

  15. Coupling of epithelial Na+ and Cl− channels by direct and indirect activation by serine proteases

    PubMed Central

    Gondzik, Veronika; Weber, Wolf Michael

    2012-01-01

    The mammalian collecting duct (CD) is continuously exposed to urinary proteases. The CD expresses an epithelial Na+ channel (ENaC) that is activated after cleavage by serine proteases. ENaC also exists at the plasma membrane in the uncleaved form, rendering activation by extracellular proteases an important mechanism for regulating Na+ transport. Many exogenous and a small number of endogenous extracellular serine proteases have been shown to activate the channel. Recently, kallikrein 1 (KLK1) was shown to increase γENaC cleavage in the native CD indicating a possible direct role of this endogenous protease in Na+ homeostasis. To explore this process, we examined the coordinated effect of this protease on Na+ and Cl− transport in a polarized renal epithelial cell line (Madin-Darby canine kidney). We also examined the role of native urinary proteases in this process. Short-circuit current (Isc) was used to measure transport of these ions. The Isc exhibited an ENaC-dependent Na+ component that was amiloride blockable and a cystic fibrosis transmembrane conductance regulator (CFTR)-dependent Cl− component that was blocked by inhibitor 172. Apical application of trypsin, an exogenous S1 serine protease, activated IENaC but was without effects on ICFTR. Subtilisin an exogenous S8 protease that mimics endogenous furin-type proteases activated both currents. A similar activation was also observed with KLK1 and native rat urinary proteases. Activation with urinary proteases occurred within minutes and at protease concentrations similar to those in the CD indicating physiological significance of this process. ENaC activation was irreversible and mediated by enhanced cleavage of γENaC. The activation of CFTR was indirect and likely dependent on activation of an endogenous apical membrane protease receptor. Collectively, these data demonstrate coordinated stimulation of separate Na+ and Cl− transport pathways in renal epithelia by extracellular luminal proteases. They

  16. Protein 4.1-mediated membrane targeting of human discs large in epithelial cells.

    PubMed

    Hanada, Toshihiko; Takeuchi, Atsuko; Sondarva, Gautam; Chishti, Athar H

    2003-09-01

    Human discs large (hDlg) protein binds to protein 4.1R via a motif encoded by an alternatively spliced exon located between the SH3 and the C-terminal guanylate kinase-like domains. To evaluate the functional significance of protein 4.1R binding for subcellular localization of hDlg in vivo, we expressed full-length recombinant constructs of two naturally occurring isoforms of hDlg termed hDlg-I2 and hDlg-I3. The hDlg-I3 but not the hDlg-I2 isoform binds to the FERM (Four.1-Ezrin-Radixin-Moesin) domain of protein 4.1R in vitro. Upon transient transfection into subconfluent Madine-Darby canine kidney (MDCK) epithelial cells, the hDlg-I3 fused with the green fluorescent protein accumulated predominantly at the plasma membrane of cell-cell contact sites, whereas the hDlg-I2 fusion protein distributed in the cytoplasm. In contrast, in stably transfected confluent MDCK cells, both hDlg-I2 and -I3 isoforms localized efficiently to the lateral membrane, consistent with the previous notion that the N-terminal domain of hDlg mediates its membrane targeting in polarized epithelial cells. We introduced a double mutation (I38A/I40A) into the N-terminal domain of hDlg, which disrupted its interaction with DLG2, a key event in the membrane targeting of hDlg. Interestingly, the hDlg-I2 isoform harboring the I38A/I40A mutation mislocalized from the membrane into cytoplasm. Importantly, the hDlg-I3 isoform with the same mutation localized efficiently to the membrane of confluent MDCK cells. Together, our results demonstrate that in addition to the N-terminal targeting domain, the alternatively spliced I3 insertion plays a critical role in recruiting hDlg to the lateral membrane in epithelial cells via its interaction with protein 4.1R. PMID:12807908

  17. Functional expression and apical localization of the cystic fibrosis transmembrane conductance regulator in MDCK I cells.

    PubMed

    Mohamed, A; Ferguson, D; Seibert, F S; Cai, H M; Kartner, N; Grinstein, S; Riordan, J R; Lukacs, G L

    1997-02-15

    The gene product affected in cystic fibrosis, the cystic fibrosis transmembrane conductance regulator (CFTR), is a chlorideselective ion channel that is regulated by cAMP-dependent protein kinase-mediated phosphorylation, ATP binding and ATP hydrolysis. Mutations in the CFTR gene may result in cystic fibrosis characterized by severe pathology (e.g. recurrent pulmonary infection, male infertility and pancreatic insufficiency) involving organs expressing the CFTR. Interestingly, in the kidney, where expression of the CFTR has been reported, impaired ion transport in patients suffering from cystic fibrosis could not be observed. To understand the role of the CFTR in chloride transport in the kidney, we attempted to identify an epithelial cell line that can serve as a model. We demonstrate that the CFTR is expressed constitutively in Madine-Darby canine kidney (MDCK) type I cells, which are thought to have originated from the distal tubule of the dog nephron. We show expression at the mRNA level, using reverse transcriptase-PCR, and at the protein level, using Western blot analysis with three different monoclonal antibodies. Iodide efflux measurements indicate that CFTR expression confers a plasma membrane anion conductance that is responsive to stimulation by cAMP. The cAMP-stimulated iodide release is sensitive to glybenclamide, diphenylamine carboxylic acid and 5-nitro-2-(3-phenylpropylamino)benzoic acid, but not to 4,4'-di-isothiocyanostilbene-2,2'-disulphonic acid, an inhibitor profile characteristic of the CFTR chloride channel. Finally, the polarized localization of the CFTR to the apical plasma membrane was established by iodide efflux measurements and cell-surface biotinylation on MDCK I monolayers. Interestingly, MDCK type II cells, which are thought to have originated from the proximal tubule of the kidney, lack CFTR protein expression and cAMP-stimulated chloride conductance. In conclusion, we propose that MDCK type I and II cells can serve as convenient

  18. Functional expression and apical localization of the cystic fibrosis transmembrane conductance regulator in MDCK I cells.

    PubMed Central

    Mohamed, A; Ferguson, D; Seibert, F S; Cai, H M; Kartner, N; Grinstein, S; Riordan, J R; Lukacs, G L

    1997-01-01

    The gene product affected in cystic fibrosis, the cystic fibrosis transmembrane conductance regulator (CFTR), is a chlorideselective ion channel that is regulated by cAMP-dependent protein kinase-mediated phosphorylation, ATP binding and ATP hydrolysis. Mutations in the CFTR gene may result in cystic fibrosis characterized by severe pathology (e.g. recurrent pulmonary infection, male infertility and pancreatic insufficiency) involving organs expressing the CFTR. Interestingly, in the kidney, where expression of the CFTR has been reported, impaired ion transport in patients suffering from cystic fibrosis could not be observed. To understand the role of the CFTR in chloride transport in the kidney, we attempted to identify an epithelial cell line that can serve as a model. We demonstrate that the CFTR is expressed constitutively in Madine-Darby canine kidney (MDCK) type I cells, which are thought to have originated from the distal tubule of the dog nephron. We show expression at the mRNA level, using reverse transcriptase-PCR, and at the protein level, using Western blot analysis with three different monoclonal antibodies. Iodide efflux measurements indicate that CFTR expression confers a plasma membrane anion conductance that is responsive to stimulation by cAMP. The cAMP-stimulated iodide release is sensitive to glybenclamide, diphenylamine carboxylic acid and 5-nitro-2-(3-phenylpropylamino)benzoic acid, but not to 4,4'-di-isothiocyanostilbene-2,2'-disulphonic acid, an inhibitor profile characteristic of the CFTR chloride channel. Finally, the polarized localization of the CFTR to the apical plasma membrane was established by iodide efflux measurements and cell-surface biotinylation on MDCK I monolayers. Interestingly, MDCK type II cells, which are thought to have originated from the proximal tubule of the kidney, lack CFTR protein expression and cAMP-stimulated chloride conductance. In conclusion, we propose that MDCK type I and II cells can serve as convenient

  19. Bilateral Mandibular Supernumerary Canines: A Case Report

    PubMed Central

    Abouei Mehrizi, Ehsan; Semyari, Hassan; Eslami Amirabadi, Gholamreza

    2010-01-01

    Supernumerary teeth are defined as the teeth developed in excess of the number found in a normal dentition. Supernumerary canine is an extremely rare finding particularly in the mandible. This case report presents a 25-year-old female patient with the unique feature of bilateral mandibular supplemental supernumerary canines. The patient was non-syndromic without any other supernumerary teeth. PMID:23346342

  20. Canine and feline parasitic zoonoses in China

    PubMed Central

    2012-01-01

    Canine and feline parasitic zoonoses have not been given high priority in China, although the role of companion animals as reservoirs for zoonotic parasitic diseases has been recognized worldwide. With an increasing number of dogs and cats under unregulated conditions in China, the canine and feline parasitic zoonoses are showing a trend towards being gradually uncontrolled. Currently, canine and feline parasitic zoonoses threaten human health, and cause death and serious diseases in China. This article comprehensively reviews the current status of major canine and feline parasitic zoonoses in mainland China, discusses the risks dogs and cats pose with regard to zoonotic transmission of canine and feline parasites, and proposes control strategies and measures. PMID:22839365

  1. Concomitant canine distemper, infectious canine hepatitis, canine parvoviral enteritis, canine infectious tracheobronchitis, and toxoplasmosis in a puppy.

    PubMed

    Headley, Selwyn Arlington; Alfieri, Amauri Alcindo; Fritzen, Juliana Torres Tomazi; Garcia, João Luis; Weissenböck, Herbert; da Silva, Ana Paula; Bodnar, Livia; Okano, Werner; Alfieri, Alice Fernandes

    2013-01-01

    The concomitant infections of Canine distemper virus (CDV), Canine adenovirus A types 1 (CAdV-1) and 2 (CAdV-2), Canine parvovirus type 2 (CPV-2), and Toxoplasma gondii are described in a 43-day-old mixed-breed puppy. Clinically, there were convulsions and blindness with spontaneous death; 14 siblings of this puppy, born to a 10-month-old dam, which was seropositive (titer: 1,024) for T. gondii, also died. Necropsy revealed unilateral corneal edema (blue eye), depletion of intestinal lymphoid tissue, non-collapsible lungs, congestion of meningeal vessels, and a pale area in the myocardium. Histopathology demonstrated necrotizing myocarditis associated with intralesional apicomplexan protozoa; necrotizing and chronic hepatitis associated with rare intranuclear inclusion bodies within hepatocytes; necrotizing bronchitis and bronchiolitis; interstitial pneumonia associated with eosinophilic intracytoplasmic inclusion bodies within epithelial cells; atrophy and fusion of intestinal villi with cryptal necrosis; and white matter demyelination of the cerebrum and cerebellum associated with intranuclear inclusion bodies within astrocytes. Polymerase chain reaction (PCR) amplified the partial fragments (bp) of the CDV N gene (290 bp), CPV-2c VP2 capsid protein gene (583 bp), and CAdV-1 (508 bp) and CAdV-2 (1,030 bp) E gene from urine and tissue samples. The PCR assays demonstrated that the apicomplexan protozoa observed within several organs contained DNA specific for T. gondii; genotyping revealed T. gondii type III. The findings support the characterization of concomitant infections of CDV, CAdV-1, CAdV-2, CPV-2, and T. gondii in this puppy. Further, seroreactivity to T. gondii of the dam in association with the systemic disease observed in the puppy described herein is suggestive of congenital toxoplasmosis. PMID:23293164

  2. Neuroinflammation in advanced canine glaucoma

    PubMed Central

    Jiang, Bing; Harper, Matthew M.; Kecova, Helga; Adamus, Grazyna; Kardon, Randy H.; Grozdanic, Sinisa D.

    2010-01-01

    Purpose The pathophysiological events that occur in advanced glaucoma are not well characterized. The principal purpose of this study is to characterize the gene expression changes that occur in advanced glaucoma. Methods Retinal RNA was obtained from canine eyes with advanced glaucoma as well as from healthy eyes. Global gene expression patterns were determined using oligonucleotide microarrays and confirmed by real-time PCR. The presence of tumor necrosis factor (TNF) and its receptors was evaluated by immunolabeling. Finally, we evaluated the presence of serum autoantibodies directed against retinal epitopes using western blot analyses. Results We identified over 500 genes with statistically significant changes in expression level in the glaucomatous retina. Decreased expression levels were detected for large number of functional groups, including synapse and synaptic transmission, cell adhesion, and calcium metabolism. Many of the molecules with decreased expression levels have been previously shown to be components of retinal ganglion cells. Genes with elevated expression in glaucoma are largely associated with inflammation, such as antigen presentation, protein degradation, and innate immunity. In contrast, expression of many other pro-inflammatory genes, such as interferons or interleukins, was not detected at abnormal levels. Conclusions This study characterizes the molecular events that occur in the canine retina with advanced glaucoma. Our data suggest that in the dog this stage of the disease is accompanied by pronounced retinal neuroinflammation. PMID:21042562

  3. Canine procalcitonin messenger RNA expression.

    PubMed

    Kuzi, Sharon; Aroch, Itamar; Peleg, Keren; Karnieli, Ohad; Klement, Eyal; Dank, Gillian

    2008-09-01

    Procalcitonin is considered an acute phase protein used as both a marker of infection and prognosis in human medicine. Canine procalcitonin has been previously sequenced; however, its use as a diagnostic or prognostic tool in dogs has never been assessed. A quantitative reverse transcription polymerase chain reaction (qRT-PCR) assay for canine procalcitonin messenger RNA (mRNA) was developed. Whole blood samples were collected from ill and healthy dogs. RNA was extracted and the real-time PCR was assessed. The patients' diagnoses, complete blood cell count, and differential leukocyte count results were recorded. Based on the diagnosis, dogs were divided into 5 groups: inflammatory, infectious, neoplastic, other diseases, and healthy controls. Procalcitonin mRNA expression and the hematological measures were compared between groups, and their correlations were assessed. Procalcitonin mRNA expression was assessed in 70 dogs, including infectious (17), noninfectious inflammatory (17), neoplastic (18), other diseases (7), and healthy controls (11), and was significantly (P < 0.001) higher in all ill dogs versus controls. Procalcitonin may therefore be considered an acutephase protein in dogs. However, there were no significant differences in procalcitonin mRNA expression between ill dog groups and no correlations between its expression levels and hematological measures. In 5 dogs of all disease categories, procalcitonin mRNA expression was measured twice during the course of disease. The changes in its levels were in agreement with the clinical evaluation of improvement or deterioration, suggesting a possible prognostic value. PMID:18776098

  4. Evidence for canine rehabilitation and physical therapy.

    PubMed

    Millis, Darryl L; Ciuperca, Ionut Alexandru

    2015-01-01

    This article reviews some important studies regarding canine physical rehabilitation. Bones, cartilage, muscles, ligaments, and tendons undergo atrophy if loading is decreased. Knowledge of the changes that occur with immobilization and the time course of events helps in the development of a rehabilitation program to improve tissue integrity. Outcome assessment instruments are clinically useful indicators of patient progress and the success of rehabilitation programs. A number of physical modalities are used in canine rehabilitation, although there are relatively few canine-specific studies. Rehabilitation has specific benefits in the treatment of various orthopedic and neurologic conditions. PMID:25432679

  5. Comparative functional characterization of canine IgG subclasses.

    PubMed

    Bergeron, Lisa M; McCandless, Erin E; Dunham, Steve; Dunkle, Bill; Zhu, Yaqi; Shelly, John; Lightle, Sandra; Gonzales, Andrea; Bainbridge, Graeme

    2014-01-15

    To date, very little is known about the functional characteristics of the four published canine IgG subclasses. It is not clear how each subclass engages the immune system via complement-dependent cytotoxicity (CDC) or antibody-dependent cell-mediated cytotoxicity (ADCC), or how long each antibody may last in serum. Such information is critical for understanding canine immunology and for the discovery of canine therapeutic monoclonal antibodies. Through both in vitro and ex vivo experiments to evaluate canine Fc's for effector function, complement binding, FcRn binding, and ADCC, we are now able to categorize canine subclasses by function. The subclasses share functional properties with the four human IgG subclasses and are reported herein with their function-based human analog. Canine Fc fusions, canine chimeras, and caninized antibodies were characterized. Canine subclasses A and D appear effector-function negative while subclasses B and C bind canine Fc gamma receptors and are positive for ADCC. All canine subclasses bind the neonatal Fc receptor except subclass C. By understanding canine IgGs in this way, we can apply what is known of human immunology toward translational and veterinary medicine. Thus, this body of work lays the foundation for evaluating canine IgG subclasses for therapeutic antibody development and builds upon the fundamental scholarship of canine immunology. PMID:24268690

  6. Estimating canine tooth crown height in early Australopithecus.

    PubMed

    Plavcan, J Michael; Ward, Carol V; Paulus, Faydre L

    2009-07-01

    Canine tooth size reduction and the associated reduction in canine dimorphism is a basal hominin character that also provides important evidence for models of behavioral evolution. Two specimens of Australopithecus anamensis (KNM-KP 29287 and KNM-KP 29283) that do not preserve the canine crown, but do preserve the root or alveolus, appear to suggest that canine size variation and canine dimorphism in this species may have been greater than in other hominins. We evaluate canine root and crown dimensions in a series of extant hominoids, and estimate canine crown height in Australopithecus afarensis and A. anamensis. Our results demonstrate that it is possible to generate estimates of canine crown height from basal canine crown and root dimensions with a moderate degree of accuracy. Estimates of maxillary canine crown size for A. anamensis are slightly larger than those of A. afarensis, and are approximately the same size as canines of modern female chimpanzees. Estimated mandibular canine crown height is very similar in the two species. Variation within the A. anamensis sample of estimated canine crown heights is similar to that of modern humans, suggesting a low degree of sexual dimorphism. Inclusion of estimates for KNM-KP 29287 and KNM-KP 29283 does not substantially increase either the estimate of overall canine size or variation for A. anamensis. PMID:19482334

  7. The role of pro- and anti-inflammatory cytokines in the pathogenesis of spontaneous canine CNS diseases.

    PubMed

    Spitzbarth, I; Baumgärtner, W; Beineke, A

    2012-06-15

    Dogs are comparatively frequently affected by various spontaneously occurring inflammatory and degenerative central nervous system (CNS) conditions, and immunopathological processes are a hallmark of the associated neuropathology. Due to the low regenerative capacity of the CNS a sophisticated understanding of the underlying molecular basis for disease initiation, progression and remission in canine CNS diseases represents a prerequisite for the development of novel therapeutical approaches. In addition, as many spontaneous canine CNS diseases share striking similarities with their human counterpart, knowledge about the immune pathogenesis may in part be translated for a better understanding of certain human diseases. In addition to cytokine-driven differentiation of peripheral leukocytes including different subsets of T cells recent research suggests a pivotal role of these mediators also in phenotype polarization of resident glial cells. Cytokines thus represent the key mediators of the local and systemic immune response in CNS diseases and their orchestration significantly decides on either lesion progression or remission. The aim of the present review is to summarize the growing number of data focusing on the molecular basis of the immune response during spontaneous canine CNS diseases and to detail the effect of cytokines on the immune pathogenesis of selected idiopathic, infectious, and traumatic canine CNS diseases. Steroid-responsive meningitis arteritis (SRMA) represents a unique idiopathic disease of leptomeningeal blood vessels characterized by excessive IgA secretion into the cerebrospinal fluid. Recent reports have given sophisticated insights into the cytokine-driven, immune-mediated pathogenesis of SRMA that is characterized by a biased T helper 2 cell response. Canine distemper associated leukoencephalitis represents an important spontaneously occurring disease that allows investigations on the basic pathogenesis of immune-mediated myelin loss. It

  8. Etiology of maxillary canine impaction: a review.

    PubMed

    Becker, Adrian; Chaushu, Stella

    2015-10-01

    This article is a review that enumerates the causes of impaction of the maxillary permanent canines, including hard tissue obstructions, soft tissue lesions, and anomalies of neighboring teeth, and discusses the much-argued relationship between environmental and genetic factors. These phenomena have been shown in many investigations to accompany the diagnosis of canine impaction and have been presented as unrelated anomalous features, each of which is etiologically construed as genetic, including the aberrant canine itself. While in general the influence of genetics pervades the wider picture, a guidance theory proposes an alternative etiologic line of reasoning and interpretation of these studies, in which the same genetically determined anomalous features provide an abnormal milieu in which the canine is reared and from which it is guided in its misdirected and often abortive path of eruption. PMID:26432311

  9. Canine adenovirus type 1 in a fennec fox (Vulpes zerda).

    PubMed

    Choi, Jeong-Won; Lee, Hyun-Kyoung; Kim, Seong-Hee; Kim, Yeon-Hee; Lee, Kyoung-Ki; Lee, Myoung-Heon; Oem, Jae-Ku

    2014-12-01

    A 10-mo-old female fennec fox (Vulpes zerda) with drooling suddenly died and was examined postmortem. Histologic examination of different tissue samples was performed. Vacuolar degeneration and diffuse fatty change were observed in the liver. Several diagnostic methods were used to screen for canine parvovirus, canine distemper virus, canine influenza virus, canine coronavirus, canine parainfluenza virus, and canine adenovirus (CAdV). Only CAdV type 1 (CAdV-1) was detected in several organs (liver, lung, brain, kidney, spleen, and heart), and other viruses were not found. CAdV-1 was confirmed by virus isolation and nucleotide sequencing. PMID:25632689

  10. [Nonsurgical endodontic treatment of an invaginated canine].

    PubMed

    Fernández Guerrero, F; Miñana Laliga, R; Bullon Fernandez, P

    1989-01-01

    We present a case of a maxillary canine with a dens invaginatus treated successfully. The patient had pain, swelling and a sinus tract coming from the inmature apex of the canine. The canals were enlarged and cleaned and the main canal was filled with Calcium Hydroxide to allow the root development. Seven months later, the patient was asymptomatic and the tooth was obturated with guttapercha. One year later it was confirm the success in the treatment. PMID:2638021

  11. Rootless eruption of a mandibular permanent canine.

    PubMed

    Shapira, Yehoshua; Kuftinec, Mladen M

    2011-04-01

    The purpose of this article was to describe the rootless eruption of a mandibular permanent canine in a 10-year-old boy; his mandible had been fractured in a car accident. The fracture was at the region of the developing canine, resulting in arrested root formation and causing abnormal, rootless eruption. Current theories on tooth eruption and the important role of the dental follicle in the process of eruption are discussed. PMID:21457868

  12. Canine Cytogenetics - From band to basepair

    PubMed Central

    Breen, Matthew

    2008-01-01

    Humans and dogs have coexisted for thousands of years, during which time we have developed a unique bond, centered on companionship. Along the way, we have developed purebred dog breeds in a manner that has resulted unfortunately in many of them being affected by serious genetic disorders, including cancers. With serendipity and irony the unique genetic architecture of the 21st Century genome of Man's best friend may ultimately provide many of the keys to unlock some of nature's most intriguing biological puzzles. Canine cytogenetics has advanced significantly over the past 10 years, spurred on largely by the surge of interest in the dog as a biomedical model for genetic disease and the availability of advanced genomics resources. As such the role of canine cytogenetics has moved rapidly from one that served initially to define the gross genomic organization of the canine genome and provide a reliable means to determine the chromosomal location of individual genes, to one that enabled the assembled sequence of the canine genome to be anchored to the karyotype. Canine cytogenetics now presents the biomedical research community with a means to assist in our search for a greater understanding of how genome architectures altered during speciation and in our search for genes associated with cancers that affect both dogs and humans. The cytogenetics ‘toolbox’ for the dog is now loaded. This review aims to provide a summary of some of the recent advancements in canine cytogenetics. PMID:18467825

  13. Environmental contamination by canine geohelminths

    PubMed Central

    2014-01-01

    Intestinal nematodes affecting dogs, i.e. roundworms, hookworms and whipworms, have a relevant health-risk impact for animals and, for most of them, for human beings. Both dogs and humans are typically infected by ingesting infective stages, (i.e. larvated eggs or larvae) present in the environment. The existence of a high rate of soil and grass contamination with infective parasitic elements has been demonstrated worldwide in leisure, recreational, public and urban areas, i.e. parks, green areas, bicycle paths, city squares, playgrounds, sandpits, beaches. This review discusses the epidemiological and sanitary importance of faecal pollution with canine intestinal parasites in urban environments and the integrated approaches useful to minimize the risk of infection in different settings. PMID:24524656

  14. Environmental contamination by canine geohelminths.

    PubMed

    Traversa, Donato; Frangipane di Regalbono, Antonio; Di Cesare, Angela; La Torre, Francesco; Drake, Jason; Pietrobelli, Mario

    2014-01-01

    Intestinal nematodes affecting dogs, i.e. roundworms, hookworms and whipworms, have a relevant health-risk impact for animals and, for most of them, for human beings. Both dogs and humans are typically infected by ingesting infective stages, (i.e. larvated eggs or larvae) present in the environment. The existence of a high rate of soil and grass contamination with infective parasitic elements has been demonstrated worldwide in leisure, recreational, public and urban areas, i.e. parks, green areas, bicycle paths, city squares, playgrounds, sandpits, beaches. This review discusses the epidemiological and sanitary importance of faecal pollution with canine intestinal parasites in urban environments and the integrated approaches useful to minimize the risk of infection in different settings. PMID:24524656

  15. Age estimation from canine volumes.

    PubMed

    De Angelis, Danilo; Gaudio, Daniel; Guercini, Nicola; Cipriani, Filippo; Gibelli, Daniele; Caputi, Sergio; Cattaneo, Cristina

    2015-08-01

    Techniques for estimation of biological age are constantly evolving and are finding daily application in the forensic radiology field in cases concerning the estimation of the chronological age of a corpse in order to reconstruct the biological profile, or of a living subject, for example in cases of immigration of people without identity papers from a civil registry. The deposition of teeth secondary dentine and consequent decrease of pulp chamber in size are well known as aging phenomena, and they have been applied to the forensic context by the development of age estimation procedures, such as Kvaal-Solheim and Cameriere methods. The present study takes into consideration canines pulp chamber volume related to the entire teeth volume, with the aim of proposing new regression formulae for age estimation using 91 cone beam computerized scans and a freeware open-source software, in order to permit affordable reproducibility of volumes calculation. PMID:25698302

  16. CANINE: a robotic mine dog

    NASA Astrophysics Data System (ADS)

    Stancil, Brian A.; Hyams, Jeffrey; Shelley, Jordan; Babu, Kartik; Badino, Hernán.; Bansal, Aayush; Huber, Daniel; Batavia, Parag

    2013-01-01

    Neya Systems, LLC competed in the CANINE program sponsored by the U.S. Army Tank Automotive Research Development and Engineering Center (TARDEC) which culminated in a competition held at Fort Benning as part of the 2012 Robotics Rodeo. As part of this program, we developed a robot with the capability to learn and recognize the appearance of target objects, conduct an area search amid distractor objects and obstacles, and relocate the target object in the same way that Mine dogs and Sentry dogs are used within military contexts for exploration and threat detection. Neya teamed with the Robotics Institute at Carnegie Mellon University to develop vision-based solutions for probabilistic target learning and recognition. In addition, we used a Mission Planning and Management System (MPMS) to orchestrate complex search and retrieval tasks using a general set of modular autonomous services relating to robot mobility, perception and grasping.

  17. 9 CFR 113.202 - Canine Hepatitis and Canine Adenovirus Type 2 Vaccine, Killed Virus.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... serum-constant virus neutralization test using 50 to 300 TCID50 of canine adenovirus. (i) The 20 dogs to... negative at a 1:2 final serum dilution in a varying serum-constant virus neutralization test using 50 to.... (2) Potency test for canine hepatitis—serum neutralization test. Bulk or final container samples...

  18. Effects of vaccines on the canine immune system.

    PubMed Central

    Phillips, T R; Jensen, J L; Rubino, M J; Yang, W C; Schultz, R D

    1989-01-01

    The effects of several commercially available polyvalent canine vaccines on the immune system of the dog were examined. The results demonstrated that the polyvalent vaccines used in this study significantly suppressed the absolute lymphocyte count and that most of the polyvalent vaccines significantly suppressed lymphocyte response to mitogen, but had no effect on natural effector cell activity, neutrophil chemiluminescence, nor antibody response to canine distemper virus. The individual vaccine components from the polyvalent vaccines when inoculated alone did not significantly suppress the lymphocyte response to mitogen. However, when canine distemper virus was combined with canine adenovirus type 1 or canine adenovirus type 2, significant suppression in lymphocyte responsiveness to mitogen occurred. The results indicate that interactions between canine distemper virus and canine adenovirus type 1 or canine adenovirus type 2 are responsible for the polyvalent vaccine induced suppression of lymphocyte responsiveness. PMID:2540897

  19. Transmigration of mandibular canine – case report

    PubMed Central

    Gruszka, Katarzyna; Różyło, T. Katarzyna; Różyło-Kalinowska, Ingrid; Denkiewicz, Katarzyna; Masłowska, Klaudia

    2014-01-01

    Summary Background Transmigration is a phenomenon of movement of an unerupted tooth in the bone across the midline. This anomaly is not often found. Transmigration is more prevalent in females than in males, and more often encountered in the mandible than maxilla, it affects mostly canines. Case Report The aim of this study was to present a case report of a mandibular canine transmigration in a patient aged 12. Intraoral examination determined hypodontia of right second premolar and delayed eruption of left second premolar in maxilla, as well as persistent deciduous teeth: right second molar, left canine and second molar. The patient was referred for a Cone-Beam CT examination, which allowed precise visualization of the transmigrating canine as well as ruled out resorption of roots of mandibular incisors. Results The treatment with a maxillary fixed orthodontic appliance was finished after obtaining a satisfactory result. Proper alignment of the incisors in the anterior-posterior plane and correct midline position were accepted by the patient. Transmigrating canine after consultation with the surgeon was designed to further radiological observation. PMID:24520309

  20. Canine kobuvirus infections in Korean dogs.

    PubMed

    Oem, Jae-Ku; Choi, Jeong-Won; Lee, Myoung-Heon; Lee, Kyoung-Ki; Choi, Kyoung-Seong

    2014-10-01

    To investigate canine kobuvirus (CaKoV) infection, fecal samples (n = 59) were collected from dogs with or without diarrhea (n = 21 and 38, respectively) in the Republic of Korea (ROK) in 2012. CaKoV infection was detected in four diarrheic samples (19.0 %) and five non-diarrheic samples (13.2 %). All CaKoV-positive dogs with diarrhea were found to be infected in mixed infections with canine distemper virus and canine parvovirus or canine adenovirus. There was no significant difference in the prevalence of CaKoV in dogs with and without diarrhea. By phylogenetic analysis based on partial 3D genes and complete genome sequences, the Korean isolates were found to be closely related to each other regardless of whether they were associated with diarrhea, and to the canine kobuviruses identified in the USA and UK. This study supports the conclusion that CaKoVs from different countries are not restricted geographically and belong to a single lineage. PMID:24906525

  1. Immunologic Observations in Canine Interstitial Nephritis

    PubMed Central

    Krohn, Kai; Mero, Matti; Oksanen, Aili; Sandholm, Markus

    1971-01-01

    Immunofluorescence studies in cases of chronic interstitial nephritis (CIN) in the dog demonstrated deposition of canine IgC and C'3 in the thickened capillary walls of the glomeruli and in the mesangium. Eluates obtained from the nephritic kidneys contained antibodies of IgG type and reacted with autologous or homologous nephritic kidneys but not with normal kidneys or with any normal canine tissue. The staining pattern of fluorescein-conjugated eluates was similar to that obtained with anti-canine IgG or anti-canine C'3. The eluates did not contain leptospiral antibodies. The findings indicate that complement-fixing immune complexes are deposited in the damaged glomeruli in CIN. The nature of the antigen involved in these complexes is unknown, but it does not seem to be a component of normal canine tissue and could thus be viral or bacterial. ImagesFig 5Fig 6Fig 7Fig 8Fig 13Fig 14Fig 15Fig 16Fig 9Fig 10Fig 11Fig 12Fig 1Fig 2Fig 3Fig 4 PMID:4106382

  2. Furcation lesion in a mandibular canine.

    PubMed

    Fonseca, Dimitri Ribas; Sena, Larryson Goncalves; Santos, Maria Helena; Goncalves, Patricia Furtado

    2011-01-01

    Morphological changes can complicate dental treatment. This report presents a rare case of a furcation lesion in a mandibular canine with two roots. A 39-year-old man in general good health sought dental care for severe pain in his maxillary anterior teeth. The clinical examination showed localized swelling in the vestibular mucosa close to the mandibular left canine. Radiographic examination revealed two distinct roots and vertical bone resorption in the canine's mesial surface. Periodontal evaluation led to a diagnosis of periodontal abscess associated with furcation lesion. Despite the occurrence in an atypical location, the site of periodontal furcation received conventional therapy for initial decontamination, including tissue debridement and a combination of polyvinylpyrrolidone irrigation and antibiotics. To improve access, the decontamination was completed with surgical techniques and scaling and root planing. Early diagnosis of this rare morphological change helped to determine appropriate, timely treatment planning and optimal patient recovery. PMID:21903558

  3. Canine neuroendocrine carcinoma. A tumor resembling histiocytoma.

    PubMed

    Nickoloff, B J; Hill, J; Weiss, L M

    1985-12-01

    The clinical and light- and electron microscopic features of 20 cases of canine neuroendocrine carcinoma, initially classified as atypical histiocytomas, are reported. The locally expansile well-circumscribed dermal tumor nodules were composed of solid masses of cells with high mitotic index and multinucleation, arranged in a trabecular pattern with prominent fibrovascular connective tissue stroma rich in reticulin fibers that outlined compact cell nests. Ultrastructural studies revealed evenly dispersed chromatin, focally indented nuclei and abundant cytoplasm with perinuclear filaments, membrane-bound dense core granules, and prominent interdigitating plasma membrane projections with primitive intercellular junctions. Clinical and pathological comparisons between canine neuroendocrine carcinoma, canine histiocytomas, and human Merkel cell neoplasms are discussed. PMID:4091229

  4. Canine rabies ecology in southern Africa.

    PubMed

    Bingham, John

    2005-09-01

    Rabies is a widespread disease in African domestic dogs and certain wild canine populations. Canine rabies became established in Africa during the 20th century, coinciding with ecologic changes that favored its emergence in canids. I present a conceptual and terminologic framework for understanding rabies ecology in African canids. The framework is underpinned by 2 distinct concepts: maintenance and persistence. Maintenance encompasses the notion of indefinite transmission of infection within a local population and depends on an average transmission ratio > or =1. Maintenance in all local populations is inherently unstable, and the disease frequently becomes extinct. Persistence, the notion of long-term continuity, depends on the presence of rabies in > or =1 local population within the canine metapopulation at any time. The implications for understanding rabies ecology and control are reviewed, as are previous studies on rabies ecology in African canids. PMID:16229759

  5. The orthodontic management of ectopic canine

    PubMed Central

    Thirunavukkarasu, R.; Sriram, G.; Satish, R.

    2015-01-01

    The canines being the cornerstone of the arch and smile is one of the teeth, which has the longest eruption passage that gets influenced by local and general etiological factors easily. The initial calcification of the crowns starts at 4–5 months of age and proceeds toward eruption about 11–13 years of age with mesiobuccal crown angulation that gets corrected toward occlusion. It gets displaced buccally or palatally or may sometimes get impacted. Early intervention is the best suited to manage canine eruption patterns. Once erupted ectopically, they possess a great challenge in repositioning them back into their correct position. This case report discusses an orthodontic treatment planning and execution to correct a buccally placed canine with an anterior crossbite in an adult. PMID:26538959

  6. A Study of Transmigrated Canine in an Indian Population

    PubMed Central

    Sharma, Gaurav; Nagpal, Archna

    2014-01-01

    Aim. The purpose of this study was to investigate the prevalence of transmigrated canines in a north Indian population and association with gender, side, associated pathologies, and dental anomalies. Subjects and methods. The prospective study consisted of panoramic radiographs of 3000 patients from two dental colleges in north India. The panoramic radiographs were screened for radiographically identified position of the transmigrated tooth, retained canine, and other coexisting dental anomalies. Results. The overall prevalence of transmigrated canines (15 mandibular and 5 maxillary) was 0.66%. The prevalence of mandibular transmigrated canine was 0.5% and maxillary transmigrated canine was 0.16%. All the transmigrated canines were unilateral. The age range was 15–53 years (average age 24.1 years) and there were 12 males (60%) and 8 females (40%). Type 1 mandibular canine transmigration was the commonest type found in our study (10 cases), followed by types 2 and 4 (2 cases each) and 1 case of type 5 transmigration. Conclusion. The prevalence of transmigrated canines in the north Indian population was 0.66% and no gender predilection was evident. The transmigrated canines have a low complication rate (10.0%) and no correlation with other dental anomalies was found. Type 3 canine is the rarest form of mandibular canine transmigration.

  7. Oncolytic Virotherapy of Canine and Feline Cancer

    PubMed Central

    Gentschev, Ivaylo; Patil, Sandeep S.; Petrov, Ivan; Cappello, Joseph; Adelfinger, Marion; Szalay, Aladar A.

    2014-01-01

    Cancer is the leading cause of disease-related death in companion animals such as dogs and cats. Despite recent progress in the diagnosis and treatment of advanced canine and feline cancer, overall patient treatment outcome has not been substantially improved. Virotherapy using oncolytic viruses is one promising new strategy for cancer therapy. Oncolytic viruses (OVs) preferentially infect and lyse cancer cells, without causing excessive damage to surrounding healthy tissue, and initiate tumor-specific immunity. The current review describes the use of different oncolytic viruses for cancer therapy and their application to canine and feline cancer. PMID:24841386

  8. Canine Mammary Mixed Tumours: A Review

    PubMed Central

    Dantas Cassali, Geovanni; Cavalheiro Bertagnolli, Angélica; Ferreira, Enio; Araújo Damasceno, Karine; de Oliveira Gamba, Conrado; Bonolo de Campos, Cecília

    2012-01-01

    Mammary mixed tumours are the most frequent neoplasias in female dogs. In humans, mixed tumours are frequently found in the salivary glands and are known as pleomorphic adenomas. In addition to their histomorphologic similarities, mixed tumours and pleomorphic adenomas have the potential to become malignant and give rise to carcinomas in mixed tumours and carcinomas ex-pleomorphic adenoma, respectively. The factors associated with malignant transformation are still poorly known in the case of canine mixed tumours. However, this form of neoplasia tends to be associated with a better prognosis than other malignant histological types. This paper discusses the main features associated with female canine mammary mixed tumours. PMID:23193497

  9. A report of canine tooth syndrome.

    PubMed

    Lee, William B; O'Halloran, Henry S

    2004-03-01

    The authors describe the case of a 5-year-old girl traumatized from a dog bite to the superior aspect of the orbit in the right eye. The dog's canine tooth penetrated deep into the posterior orbit and severed the attachment of the superior oblique muscle from the globe posterior to the trochlea. The management and clinical course of the patient are described and photographs documenting the initial ocular damage and postoperative course are provided. In addition, the entity known as 'canine tooth syndrome' is reviewed. PMID:15513022

  10. Evaluation of the Broad-Range PCR-Electrospray Ionization Mass Spectrometry (PCR/ESI-MS) System and Virus Microarrays for Virus Detection

    PubMed Central

    Taliaferro, Lanyn P.; Galvin, Teresa A.; Ma, Hailun; Shaheduzzaman, Syed; Williams, Dhanya K.; Glasner, Dustin R.; Khan, Arifa S.

    2014-01-01

    Advanced nucleic acid-based technologies are powerful research tools for novel virus discovery but need to be standardized for broader applications such as virus detection in biological products and clinical samples. We have used well-characterized retrovirus stocks to evaluate the limit of detection (LOD) for broad-range PCR with electrospray ionization mass spectrometry (PCR/ESI-MS or PLEX-ID), RT-PCR assays, and virus microarrays. The results indicated that in the absence of background cellular nucleic acids, PLEX-ID and RT-PCR had a similar LOD for xenotropic murine retrovirus-related virus (XMRV; 3.12 particles per µL) whereas sensitivity of virus detection was 10-fold greater using virus microarrays. When virus was spiked into a background of cellular nucleic acids, the LOD using PLEX-ID remained the same, whereas virus detection by RT-PCR was 10-fold less sensitive, and no virus could be detected by microarrays. Expected endogenous retrovirus (ERV) sequences were detected in cell lines tested and known species-specific viral sequences were detected in bovine serum and porcine trypsin. A follow-up strategy was developed using PCR amplification, nucleotide sequencing, and bioinformatics to demonstrate that an RD114-like retrovirus sequence that was detected by PLEX-ID in canine cell lines (Madin-Darby canine kidney (MDCK) and Cf2Th canine thymus) was due to defective, endogenous gammaretrovirus-related sequences. PMID:24777034

  11. Canine adenovirus downstream processing protocol.

    PubMed

    Puig, Meritxell; Piedra, Jose; Miravet, Susana; Segura, María Mercedes

    2014-01-01

    Adenovirus vectors are efficient gene delivery tools. A major caveat with vectors derived from common human adenovirus serotypes is that most adults are likely to have been exposed to the wild-type virus and exhibit active immunity against the vectors. This preexisting immunity limits their clinical success. Strategies to circumvent this problem include the use of nonhuman adenovirus vectors. Vectors derived from canine adenovirus type 2 (CAV-2) are among the best-studied representatives. CAV-2 vectors are particularly attractive for the treatment of neurodegenerative disorders. In addition, CAV-2 vectors have shown great promise as oncolytic agents in virotherapy approaches and as vectors for recombinant vaccines. The rising interest in CAV-2 vectors calls for the development of scalable GMP compliant production and purification strategies. A detailed protocol describing a complete scalable downstream processing strategy for CAV-2 vectors is reported here. Clarification of CAV-2 particles is achieved by microfiltration. CAV-2 particles are subsequently concentrated and partially purified by ultrafiltration-diafiltration. A Benzonase(®) digestion step is carried out between ultrafiltration and diafiltration operations to eliminate contaminating nucleic acids. Chromatography purification is accomplished in two consecutive steps. CAV-2 particles are first captured and concentrated on a propyl hydrophobic interaction chromatography column followed by a polishing step using DEAE anion exchange monoliths. Using this protocol, high-quality CAV-2 vector preparations containing low levels of contamination with empty viral capsids and other inactive vector forms are typically obtained. The complete process yield was estimated to be 38-45 %. PMID:24132487

  12. Polarization developments

    SciTech Connect

    Prescott, C.Y.

    1993-07-01

    Recent developments in laser-driven photoemission sources of polarized electrons have made prospects for highly polarized electron beams in a future linear collider very promising. This talk discusses the experiences with the SLC polarized electron source, the recent progress with research into gallium arsenide and strained gallium arsenide as a photocathode material, and the suitability of these cathode materials for a future linear collider based on the parameters of the several linear collider designs that exist.

  13. Experimental Forelimb Allotransplantation in Canine Model.

    PubMed

    Hong, Sa-Hyeok; Eun, Seok-Chan

    2016-01-01

    As reconstructive transplantation is gaining popularity as a viable alternative for upper limb amputees, it is becoming increasingly important for plastic surgeons to renew surgical skills and knowledge of this area. Forelimb allotransplantation research has been performed previously in rodent and swine models. However, preclinical canine forelimb allotransplantation studies are lacking in the literature. The purpose of this paper is to provide an overview of the surgical skills necessary to successfully perform forelimb transplantation in canines as a means to prepare for clinical application. A total of 18 transplantation operations on canines were performed. The recipient limb was shortened at the one-third proximal forearm level. The operation was performed in the following order: bones (two reconstructive plates), muscles and tendons (separately sutured), nerves (median, ulnar, and radial nerve), arteries (two), and veins (two). The total mean time of transplantation was 5 hours ± 30 minutes. All of the animals that received transplantation were treated with FK-506 (tacrolimus, 2 mg/kg) for 7 days after surgery. Most allografts survived with perfect viability without vascular problems during the early postoperative period. The canine forelimb allotransplantation model is well qualified to be a suitable training model for standard transplantation and future research work. PMID:27597952

  14. [Backshifting of lower canines in occlusion regulation].

    PubMed

    Michałowska-Sarosiek, A; Wedrychowska-Szulc, B; Doniec-Zawidzka, I

    1990-08-01

    In 17 patients the lower canines were shifted back during treatment of prognathic malocclusion and cross-bites. Stable devices with retraction loops, rubber or spring traction were used. The duration of active treatment was about 3 months, on average. During the treatment gaps after the removed premolars were closed completely, although this is nearly impossible when removable devices are applied. PMID:2104380

  15. Prostate histotripsy for BPH: initial canine results

    NASA Astrophysics Data System (ADS)

    Roberts, William W.; Hall, Timothy L.; Hempel, Christopher R.; Cain, Charles A.

    2009-02-01

    Histotripsy is an extracorporeal ablative technology that utilizes microsecond pulses of intense ultrasound (< 1% duty cycle) to produce nonthermal, mechanical fractionation of targeted tissue. We have previously demonstrated the feasibility of histotripsy prostate ablation. In this study we sought to assess the chronic tissue response, tolerability and safety of histotripsy in a chronic in vivo canine model. Five acute and thirteen chronic canine subjects were anesthetized and treated with histotripsy targeting the prostate. Pulses consisted of 3 cycle bursts of 750 kHz ultrasound at a repetition rate of 300 Hz delivered transabdominally from a highly focused 15 cm aperture array. Transrectal ultrasound imaging provided accurate targeting and real-time monitoring of histotripsy treatment. Prostates were harvested at 0, 7, 28, or 56 days after treatment. Consistent mechanical tissue fractionation and debulking of prostate tissue was seen acutely and at delayed time points without collateral injury. Urothelialization of the treatment cavity was apparent 28 days after treatment. Canine subjects tolerated histotripsy with minimal hematuria or discomfort. Only mild transient lab abnormalities were noted. Histotripsy is a promising non-invasive therapy for prostate tissue fractionation and debulking that appears safe and well tolerated without systemic side effects in the canine model.

  16. Experimental Forelimb Allotransplantation in Canine Model

    PubMed Central

    2016-01-01

    As reconstructive transplantation is gaining popularity as a viable alternative for upper limb amputees, it is becoming increasingly important for plastic surgeons to renew surgical skills and knowledge of this area. Forelimb allotransplantation research has been performed previously in rodent and swine models. However, preclinical canine forelimb allotransplantation studies are lacking in the literature. The purpose of this paper is to provide an overview of the surgical skills necessary to successfully perform forelimb transplantation in canines as a means to prepare for clinical application. A total of 18 transplantation operations on canines were performed. The recipient limb was shortened at the one-third proximal forearm level. The operation was performed in the following order: bones (two reconstructive plates), muscles and tendons (separately sutured), nerves (median, ulnar, and radial nerve), arteries (two), and veins (two). The total mean time of transplantation was 5 hours ± 30 minutes. All of the animals that received transplantation were treated with FK-506 (tacrolimus, 2 mg/kg) for 7 days after surgery. Most allografts survived with perfect viability without vascular problems during the early postoperative period. The canine forelimb allotransplantation model is well qualified to be a suitable training model for standard transplantation and future research work. PMID:27597952

  17. Canine brachycephalic airway syndrome: surgical management.

    PubMed

    Trappler, Michelle; Moore, Kenneth

    2011-05-01

    Many surgical options have been described to treat various aspects of canine brachycephalic airway syndrome (BAS). This article describes the surgical management, postoperative care, and prognosis of this condition. The pathophysiology and medical therapy of BAS are described in a companion article. PMID:21870354

  18. Production of monoclonal antibodies against canine leukocytes.

    PubMed

    Aguiar, Paulo Henrique Palis; Borges dos Santos, Roberto Robson; Lima, Carla Andrade; Rios de Sousa Gomes, Hilton; Larangeira, Daniela Farias; Santos, Patrícia Meira; Barrouin-Melo, Stella Maria; Conrado dos-Santos, Washington Luis; Pontes-de-Carvalho, Lain

    2004-04-01

    A panel of anti-canine leukocyte monoclonal antibodies (MAbs) was produced by immunizing BALB/c mice with canine peripheral blood mononuclear cells (PBMC), either resting or stimulated with concanavalin A (ConA). Three out of 28 clones-IH1, AB6, and HG6-screened by ELISA and producing antibody with the highest specificity for canine cell immunostaining, were subjected to three subsequent subcloning steps by limiting dilution, and selected for further characterization. These MAbs belonged to IgG1 (HG6 and IH1) and IgG2a (AB6) isotypes. The distribution of cell populations expressing the antigen recognized by the antibodies was identified by indirect immunoflorescence on canine PBMC and on tissue sections of lymph node, spleen, liver and skin. The possible crossreactivity with human PBMC was also examined in immunocytochemistry. One of the antibodies specifically recognized macrophages. The MAbs presented here can be foreseen as possible valuable diagnostic and research tools to study immune functions in dogs. PMID:15165486

  19. DELINEATING TOXIC AREAS BY CANINE OLFACTION

    EPA Science Inventory

    A research project was undertaken to learn how the highly acute olfactory sensitivity of the canine could be applied with advantage to environmental problems. The objectives were to determine how dogs could be trained to detect hazardous and toxic pollutants in the environment an...

  20. Neuronal polarization.

    PubMed

    Takano, Tetsuya; Xu, Chundi; Funahashi, Yasuhiro; Namba, Takashi; Kaibuchi, Kozo

    2015-06-15

    Neurons are highly polarized cells with structurally and functionally distinct processes called axons and dendrites. This polarization underlies the directional flow of information in the central nervous system, so the establishment and maintenance of neuronal polarization is crucial for correct development and function. Great progress in our understanding of how neurons establish their polarity has been made through the use of cultured hippocampal neurons, while recent technological advances have enabled in vivo analysis of axon specification and elongation. This short review and accompanying poster highlight recent advances in this fascinating field, with an emphasis on the signaling mechanisms underlying axon and dendrite specification in vitro and in vivo. PMID:26081570

  1. Polarization Aberrations

    NASA Technical Reports Server (NTRS)

    Mcguire, James P., Jr.; Chipman, Russell A.

    1990-01-01

    The analysis of the polarization characteristics displayed by optical systems can be divided into two categories: geometrical and physical. Geometrical analysis calculates the change in polarization of a wavefront between pupils in an optical instrument. Physical analysis propagates the polarized fields wherever the geometrical analysis is not valid, i.e., near the edges of stops, near images, in anisotropic media, etc. Polarization aberration theory provides a starting point for geometrical design and facilitates subsequent optimization. The polarization aberrations described arise from differences in the transmitted (or reflected) amplitudes and phases at interfaces. The polarization aberration matrix (PAM) is calculated for isotropic rotationally symmetric systems through fourth order and includes the interface phase, amplitude, linear diattenuation, and linear retardance aberrations. The exponential form of Jones matrices used are discussed. The PAM in Jones matrix is introduced. The exact calculation of polarization aberrations through polarization ray tracing is described. The report is divided into three sections: I. Rotationally Symmetric Optical Systems; II. Tilted and Decentered Optical Systems; and Polarization Analysis of LIDARs.

  2. Mandibular canine dimensions as an aid in gender estimation

    PubMed Central

    Rajarathnam, Basetty Neelakantam; David, Maria Priscilla; Indira, Annamalai Ponnuswamy

    2016-01-01

    Background: All humans have an identity in life; compassionate societies require this identity to be recognized even after death. Objectives: To measure the dimensions of the mandibular canine and assess the usefulness of the mandibular canine as an aid in gender estimation. Materials and Methods: The study population comprised 200 subjects inclusive of 100 males and 100 females with an age range of 18–25 years. Measurements made in mm at the contact point were of mesiodistal width of the right and left canines and intercanine distance both intraorally and on casts, and the mandibular canine index (MCI) was calculated. The obtained data were subjected to t-test/Mann-Whitney test and discriminant function analysis. Results: All parameters of mandibular canines, namely, intercanine distance, canine width, and canine index were greater in males compared to females suggesting significant sexual dimorphism of mandibular canines. On subjecting the data to discriminant function analysis, it classified sex correctly in 73% of the samples. Conclusion: The result of our study establishes the existence of significant sexual dimorphism in mandibular canines. We can therefore, recommend the use of mandibular canine dimensions as an applicable and additional method for gender determination in human identification. PMID:27555724

  3. Canine cytochrome P450 (CYP) pharmacogenetics

    PubMed Central

    Court, Michael H.

    2013-01-01

    Synopsis The cytochrome P450 (CYP) drug metabolizing enzymes are essential for the efficient elimination of many clinically used drugs. These enzymes typically display high interindividual variability in expression and function resulting from enzyme induction, inhibition, and genetic polymorphism thereby predisposing patients to adverse drug reactions or therapeutic failure. There are also substantial species differences in CYP substrate specificity and expression that complicate direct extrapolation of information from humans to veterinary species. This article reviews the available published data regarding the presence and impact of genetic polymorphisms on CYP-dependent drug metabolism in dogs in the context of known human-dog CYP differences. Canine CYP1A2, which metabolizes phenacetin, caffeine, and theophylline, is the most widely studied polymorphic canine CYP. A single nucleotide polymorphism resulting in a CYP1A2 premature stop codon (c.1117C>T; R383X) with a complete lack of enzyme is highly prevalent in certain dog breeds including Beagle and Irish wolfhound. This polymorphism was shown to substantially affect the pharmacokinetics of several experimental compounds in Beagles during preclinical drug development. However, the impact on the pharmacokinetics of phenacetin (a substrate specific for human CYP1A2) was quite modest probably because other canine CYPs are capable of metabolizing phenacetin. Other canine CYPs with known genetic polymorphisms include CYP2C41 (gene deletion), as well as CYP2D15, CYP2E1, and CYP3A12 (coding SNPs). However the impact of these variants on drug metabolism in vitro or on drug pharmacokinetics is unknown. Future systematic investigations are needed to comprehensively identify CYP genetic polymorphisms that are predictive of drug effects in canine patients. PMID:23890236

  4. Canine visceral leishmaniasis in Sicily.

    PubMed

    Orndorff, G R; Cooper, B A; Smith, W; Ryan, J R

    2000-01-01

    The Sicilian province of Catania is an active foci for human visceral leishmaniasis (VL) in the Mediterranean area. Approximately 10 to 15 cases of VL are diagnosed via hospital admissions each year in this community. Recently, an increase in VL case reporting by Sicilian physicians was noted, with 38 and 37 VL cases in 1996 and 1997, respectively. Before 1995, there were no reported VL cases among U.S. military personnel or their family members living in Sicily. However, since 1996, there have been four cases referred to Walter Reed Army Medical Center for diagnosis and treatment, all involving the children of personnel assigned to Naval Air Station Sigonella. Exposure histories for all infected individuals excluded exposure to Leishmania parasites outside of Sicily. All patients lived in areas where vectoring sandflies are present. All had dogs as family pets. To evaluate the level of infection among dogs owned by Navy personnel and their families, U.S. Navy Environmental and Preventive Medicine Unit 7, in a collaborative study with the U.S. Army Veterinary Clinic, Naval Air Station Sigonella, and the Walter Reed Army Institute of Research, performed clinical evaluation and serological testing of 50 dogs residing with U.S. personnel assigned to Naval Air Station Sigonella. The data indicate a high exposure rate to Leishmania (60% of the animals tested had elevated immunoglobulin M antibody levels) in the study population, suggesting that they were infected with Leishmania infantum. Distribution of seropositive dogs by sex was equal. Most of the dogs studied appeared to be in good health. However, inapparent infection of dogs, seen by Italian veterinarians, has been observed throughout all areas of Catania. Sandflies responsible for vectoring L. infantum were trapped in the same locations as the dogs sampled in this study. The level of subclinical infection was 75% among seropositive dogs. The overall level of canine infection observed was higher than expected

  5. Polar Bear

    USGS Publications Warehouse

    Amstrup, S.D.; DeMaster

    1988-01-01

    Polar bears are long-lived, late-maturing carnivores that have relatively low rates of reproduction and natural mortality. Their populations are susceptible to disturbance from human activities, such as the exploration and development of mineral resources or hunting. Polar bear populations have been an important renewable resource available to coastal communities throughout the Arctic for thousands of years.

  6. Effects of canine serum collected from dogs at different estrous cycle stages on in vitro nuclear maturation of canine oocytes.

    PubMed

    Oh, Hyun Ju; Fibrianto, Yuda Heru; Kim, Min Kyu; Jang, Goo; Hossein, M Shamim; Kim, Hye Jin; Kang, Sung Keun; Lee, Byeong Chun; Hwang, Woo Suk

    2005-08-01

    Canine oocytes are ovulated at prophase of the first meiotic division and undergo maturation in the distal part of the oviduct for at least 48-72 h. Because of these differences from other domestic mammals, the efficiency of in vitro maturation (IVM) of canine oocyte is very low. The present study was conducted to evaluate the effects of canine serum on IVM of canine oocytes recovered from ovaries in various reproductive states (follicular, luteal or anestrous stages). Oocytes were recovered by mincing ovaries from bitches presented for ovariohysterectomy at various stages of the estrous cycle. Heat-inactivated canine serum was prepared with blood taken from dogs at the anestrous, estrous or diestrous stage of the estrous cycle as determined by progesterone concentration and vaginal cytology. Oocytes were cultured for 72 h in tissue culture medium (TCM)-199 supplemented with 10% canine anestrous, estrous or diestrous serum or fetal bovine serum (FBS) (experiment 1), or supplemented with 0 (control), 5%, 10% or 20% canine estrous serum (experiment 2). In experiment 1, IVM of oocytes collected at the follicular stage of the estrous cycle to metaphase II (MII) stage was higher (p < 0.05) with canine estrous serum (14.2%) than with canine anestrous (5.2%) or diestrous serum (6.3%), FBS (2.2%) or in the control (2.2%). In experiment 2, oocytes collected at the follicular stage of the estrous cycle cultured in TCM-199 with 10% canine estrous serum showed a higher maturation rate to MII stage (13.5%, p < 0.05) compared with those cultured with 5% (1.3% MII) or 20% canine estrous serum (5.1% MII) or the control (2.7% MII). In conclusion, our results demonstrate that supplementing culture medium with 10% canine estrous serum improves IVM of canine follicular stage oocytes. PMID:16261767

  7. Canine preprorelaxin: nucleic acid sequence and localization within the canine placenta.

    PubMed

    Klonisch, T; Hombach-Klonisch, S; Froehlich, C; Kauffold, J; Steger, K; Steinetz, B G; Fischer, B

    1999-03-01

    Employing uteroplacental tissue at Day 35 of gestation, we determined the nucleic acid sequence of canine preprorelaxin using reverse transcription- and rapid amplification of cDNA ends-polymerase chain reaction. Canine preprorelaxin cDNA consisted of 534 base pairs encoding a protein of 177 amino acids with a signal peptide of 25 amino acids (aa), a B domain of 35 aa, a C domain of 93 aa, and an A domain of 24 aa. The putative receptor binding region in the N'-terminal part of the canine relaxin B domain GRDYVR contained two substitutions from the classical motif (E-->D and L-->Y). Canine preprorelaxin shared highest homology with porcine and equine preprorelaxin. Northern analysis revealed a 1-kilobase transcript present in total RNA of canine uteroplacental tissue but not of kidney tissue. Uteroplacental tissue from two bitches each at Days 30 and 35 of gestation were studied by in situ hybridization to localize relaxin mRNA. Immunohistochemistry for relaxin, cytokeratin, vimentin, and von Willebrand factor was performed on uteroplacental tissue at Day 30 of gestation. The basal cell layer at the core of the chorionic villi was devoid of relaxin mRNA and immunoreactive relaxin or vimentin but was immunopositive for cytokeratin and identified as cytotrophoblast cells. The cell layer surrounding the chorionic villi displayed specific hybridization signals for relaxin mRNA and immunoreactivity for relaxin and cytokeratin but not for vimentin, and was identified as syncytiotrophoblast. Those areas of the chorioallantoic tissue with most intense relaxin immunoreactivity were highly vascularized as demonstrated by immunoreactive von Willebrand factor expressed on vascular endothelium. The uterine glands and nonplacental uterine areas of the canine zonary girdle placenta were devoid of relaxin mRNA and relaxin. We conclude that the syncytiotrophoblast is the source of relaxin in the canine placenta. PMID:10026098

  8. Polarized rainbow.

    PubMed

    Können, G P; de Boer, J H

    1979-06-15

    The Airy theory of the rainbow is extended to polarized light. For both polarization directions a simple analytic expression is obtained for the intensity distribution as a function of the scattering angle in terms of the Airy function and its derivative. This approach is valid at least down to droplet diameters of 0.3 mm in visible light. The degree of polarization of the rainbow is less than expected from geometrical optics; it increases with droplet size. For a droplet diameter >1 mm the locations of the supernumerary rainbows are equal for both polarization directions, but for a diameter <1 mm the supernumerary rainbows of the weaker polarization component are located between those in the strong component. PMID:20212586

  9. Combined orthodontic-surgical management of a transmigrated mandibular canine.

    PubMed

    Cavuoti, Serena; Matarese, Giovanni; Isola, Gaetano; Abdolreza, Jamilian; Femiano, Felice; Perillo, Letizia

    2016-07-01

    The presence of an impacted mandibular canine is one of the most difficult challenges that an orthodontist will meet. Orthodontic treatment is planned on an individual basis after thoroughly considering the patient's overall facial and dentoskeletal characteristics; the duration, risks, and costs of treatment; patient preferences; and the orthodontist's experience. This article reports an orthodontic treatment of a boy, age 12.9 years, with an impacted mandibular canine in the permanent dentition that was successfully managed. Radiographic analysis indicated a transmigration of the mandibular right canine. The orthodontic treatment plan included extraction of the deciduous right canine followed by surgical exposure and ligation of the permanent canine. Eruption was properly guided, and the correct position of the tooth was achieved. This challenging treatment approach is described in detail, including the mechanics used to align the impacted canine. PMID:26502299

  10. Survivin inhibition via EZN-3042 in canine lymphoma and osteosarcoma.

    PubMed

    Shoeneman, J K; Ehrhart, E J; Charles, J B; Thamm, D H

    2016-06-01

    Canine lymphoma (LSA) and osteosarcoma (OS) have high mortality rates and remain in need of more effective therapeutic approaches. Survivin, an inhibitor of apoptosis (IAP) family member protein that inhibits apoptosis and drives cell proliferation, is commonly elevated in human and canine cancer. Survivin expression is a negative prognostic factor in dogs with LSA and OS, and canine LSA and OS cell lines express high levels of survivin. In this study, we demonstrate that survivin downregulation in canine LSA and OS cells using a clinically applicable locked nucleic acid antisense oligonucleotide (EZN-3042, Enzon Pharmaceuticals, Piscataway Township, NJ, USA) inhibits growth, induces apoptosis and enhances chemosensitivity in vitro, and inhibits survivin transcription and protein production in orthotopic canine OS xenografts. Our findings strongly suggest that survivin-directed therapies might be effective in treatment of canine LSA and OS and support evaluation of EZN-3042 in dogs with cancer. PMID:24923332

  11. Unilateral Maxillary Canine Agenesis: A Case Report and Literature Review

    PubMed Central

    Koç, Nagihan; Çağırankaya, L. Berna; Akkaya, Nursel

    2014-01-01

    Congenital absence of maxillary permanent canines is an extremely rare condition, which may appear as part of a syndrome or as a nonsyndromic form. Nonsyndromic canine agenesis combined with other types of tooth agenesis has occasionally been described in the literature but isolated cases are rarely observed. This report presents an isolated case of maxillary permanent canine agenesis in a healthy 18-year-old female patient and a literature review on the prevalence, etiology, and differential diagnosis of the condition. PMID:25177502

  12. Polar Glaciology

    NASA Technical Reports Server (NTRS)

    Robin, G. D.

    1984-01-01

    Two fields of research on polar ice sheets are likely to be of dominant interest during the 1990s. These are: the role of polar ice sheets in the hydrological cycle ocean-atmosphere-ice sheets-oceans, especially in relation to climate change; and the study and interpretation of material in deep ice cores to provide improved knowledge of past climates and of the varying levels of atmospheric constituents such as CO2, NOx, SO2, aerosols, etc., over the past 200,000 years. Both topics require a better knowledge of ice dynamics. Many of the studies that should be undertaken in polar regions by Earth Observing System require similar instruments and techniques to those used elsewhere over oceans and inland surfaces. However to study polar regions two special requirements need to be met: Earth Observing System satellite(s) need to be in a sufficiently high inclination orbit to cover most of the polar regions. Instruments must also be adapted, often by relatively limited changes, to give satisfactory data over polar ice. The observational requirements for polar ice sheets in the 1990s are summarized.

  13. Myoepithelial cells in canine mammary tumours.

    PubMed

    Sánchez-Céspedes, Raquel; Millán, Yolanda; Guil-Luna, Silvia; Reymundo, Carlos; Espinosa de Los Monteros, Antonio; Martín de Las Mulas, Juana

    2016-01-01

    Mammary tumours are the most common neoplasms of female dogs. Compared to mammary tumours of humans and cats, myoepithelial (ME) cell involvement is common in canine mammary tumours (CMT) of any subtype. Since ME cell involvement in CMT influences both histogenetic tumour classification and prognosis, correct identification of ME cells is important. This review describes immunohistochemical methods for identification of canine mammary ME cells used in vivo. In addition, phenotypic and genotypic methods to isolate ME cells for in vitro studies to analyse tumour-suppressor protein production and gene expression are discussed. The contribution of ME cells to both histogenetic classifications and the prognosis of CMT is compared with other species and the potential use of ME cells as a method to identify carcinoma in situ is discussed. PMID:26639832

  14. Rhabdomyolysis as a complication of canine babesiosis.

    PubMed

    Jacobson, L S; Lobetti, R G

    1996-06-01

    Rhabdomyolysis was diagnosed in two dogs with babesiosis. The first animal presented with muscle pain and caramel-coloured urine, and had markedly elevated serum myoglobin and muscle enzymes. Acute renal failure complicated the clinical picture. The second dog exhibited muscle pain and tremors, together with neurological signs and pulmonary oedema, and died soon after admission. Muscle necrosis and haemorrhage were found at necropsy. In human malaria, a disease clinically similar to canine babesiosis, rhabdomyolysis is unusual, but clinically silent muscle damage appears to be common. Likewise, biochemical evidence of muscle damage is readily found in experimental bovine babesiosis. Muscle enzymes were mildly elevated in three dogs with severe babesiosis and pigmenturia but there was no obvious muscle damage, indicating that this might also apply to canine babesiosis. The pathogenesis of infection-associated rhabdomyolysis and acute renal failure remains unclear, but inflammatory cytokines and nitric oxide could play an important role. PMID:8965483

  15. Characterization of pantropic canine coronavirus from Brazil.

    PubMed

    Pinto, Luciane D; Barros, Iracema N; Budaszewski, Renata F; Weber, Matheus N; Mata, Helena; Antunes, Jéssica R; Boabaid, Fabiana M; Wouters, Angélica T B; Driemeier, David; Brandão, Paulo E; Canal, Cláudio W

    2014-12-01

    Characterization of canine coronavirus (CCoV) strains currently in circulation is essential for understanding viral evolution. The aim of this study was to determine the presence of pantropic CCoV type IIa in tissue samples from five puppies that died in Southern Brazil as a result of severe gastroenteritis. Reverse-transcriptase PCR was used to generate amplicons for sequence analysis. Phylogenetic analysis of the CCoV-IIa strains indicated that they were similar to those found in other countries, suggesting a common ancestor of these Brazilian isolates. This is the first report of pantropic CCoV-II in puppies from Latin America and our findings highlight that CCoV should be included as a differential diagnosis when dogs present with clinical signs and lesions typically seen with canine parvovirus infection. PMID:25294661

  16. Polarizing cues.

    PubMed

    Nicholson, Stephen P

    2012-01-01

    People categorize themselves and others, creating ingroup and outgroup distinctions. In American politics, parties constitute the in- and outgroups, and party leaders hold sway in articulating party positions. A party leader's endorsement of a policy can be persuasive, inducing co-partisans to take the same position. In contrast, a party leader's endorsement may polarize opinion, inducing out-party identifiers to take a contrary position. Using survey experiments from the 2008 presidential election, I examine whether in- and out-party candidate cues—John McCain and Barack Obama—affected partisan opinion. The results indicate that in-party leader cues do not persuade but that out-party leader cues polarize. This finding holds in an experiment featuring President Bush in which his endorsement did not persuade Republicans but it polarized Democrats. Lastly, I compare the effect of party leader cues to party label cues. The results suggest that politicians, not parties, function as polarizing cues. PMID:22400143

  17. Polar motion

    NASA Technical Reports Server (NTRS)

    Kolenkiewicz, R.

    1973-01-01

    Tracking of the Beacon Explorer-C satellite by a precision laser system was used to measure the polar motion and solid earth tide. The tidal perturbation of satellite latitude is plotted as variation in maximum latitude in seconds of arc on earth's surface as a function of the date, and polar motion is shown by plotting the variation in latitude of the laser in seconds of arc along the earth's surface as a function of date

  18. Treatment of canine scabies with milbemycin oxime.

    PubMed

    Miller, W H; de Jaham, C; Scott, D W; Cayatte, S M; Bagladi, M S; Buerger, R G

    1996-04-01

    The purpose of this study was to determine the efficacy of orally administered milbemycin oxime in the treatment of canine scabies. Forty dogs were treated. Mean drug dosage for all dogs was approximately 2 mg/kg body weight. Twenty-seven dogs received 3 doses separated by 7 d, and 13 dogs received 2 doses separated by 14 d. All dogs were clinically normal following treatment and no adverse reactions were detected. PMID:8801016

  19. Ultrasonographic characteristics of canine renal lymphoma.

    PubMed

    Taylor, Angela J; Lara-Garcia, Ana; Benigni, Livia

    2014-01-01

    There is little published information on the ultrasonographic appearance of canine renal lymphoma. The purpose of this retrospective study was to provide additional information regarding the ultrasonographic characteristics of canine renal lymphoma, suggest ultrasonographic description criteria, and evaluate the role of fine-needle aspirate cytology in the diagnosis of this disease. The ultrasonographic features of confirmed renal lymphoma were reviewed in ten dogs. Pyelectasia was found in all dogs. Other ultrasonographic findings were loss of corticomedullary distinction (9/10 dogs), renomegaly (8/10 dogs), renal deformity (6/10 dogs), hypoechoic lesion(s) (6/10 dogs), and hyperechoic lesion(s) (2/10 dogs). Hypoechoic lesions were described as masses, nodules, and indistinct areas. In 30% of the cases (3/10 dogs) ultrasound revealed only minor abnormalities, including grade 1 pyelectasia, mild renomegaly, and focal loss of corticomedullary definition. Bilateral lesions were seen in nine dogs (90%). Renal fine-needle aspirates were performed in 9/10 dogs, yielding a diagnosis in seven on first attempt (78%). Two dogs had been given a provisional cytological diagnosis of round cell neoplasia; in one dog lymphoma was confirmed by second aspirate and by tissue core biopsy in the other. In 1/10 dogs, lymphoma was found at necropsy. Findings indicated that ultrasonographic signs of canine renal lymphoma may be subtle, canine renal lymphoma should be included in the differential diagnosis when the above ultrasonographic features are observed, and fine-needle aspirate cytology is a useful method for diagnosing this disease. PMID:24629062

  20. Remote detection of explosives using trained canines

    SciTech Connect

    Smith, J.C.

    1983-03-01

    Use of dogs is a search method which combines high probability of detection, speed of search, and low cost. It was concluded that the canine could be used for explosive screening of personnel, but that it was imperative that the dog be in a position remote from employees and employee traffic. A study was made of the design of booths and air flow for this purpose. Results of tests and conclusions are given and discussed. (DLC)

  1. Increasing Incidence of Canine Leptospirosis in Switzerland

    PubMed Central

    Major, Andrea; Schweighauser, Ariane; Francey, Thierry

    2014-01-01

    A marked increase in canine leptospirosis was observed in Switzerland over 10 years with a peak incidence of 28.1 diagnosed cases/100,000 dogs/year in the most affected canton. With 95% affected dogs living at altitudes <800 m, the disease presented a seasonal pattern associated with temperature (r2 0.73) and rainfall (r2 0.39), >90% cases being diagnosed between May and October. The increasing yearly incidence however was only weakly correlated with climatic data including number of summer (r2 0.25) or rainy days (r2 0.38). Serovars Australis and Bratislava showed the highest seropositivity rates with 70.5% and 69.1%, respectively. Main clinical manifestations included renal (99.6%), pulmonary (76.7%), hepatic (26.0%), and hemorrhagic syndromes (18.2%), leading to a high mortality rate (43.3%). Similar to the human disease, liver involvement had the strongest association with negative outcome (OR 16.3). Based on these data, canine leptospirosis presents similar features and severity as the human infection for which it therefore can be considered a model. Its re-emergence in a temperate country with very high incidence rates in canines should thus be viewed as a warning and emphasize the need for increased awareness in other species. PMID:25032740

  2. Canine kobuviruses in diarrhoeic dogs in Italy.

    PubMed

    Di Martino, Barbara; Di Felice, Elisabetta; Ceci, Chiara; Di Profio, Federica; Marsilio, Fulvio

    2013-09-27

    Canine kobuviruses (CaKVs) are newly recognized picornaviruses recently detected in dogs in the US. By molecular analysis of the whole genome, CaKV that appeared genetically closest to the murine kobuvirus (MuKV) and to the human Aichi virus (AiV), may be classified in the Kobuvirus genus as new genotype (CaKV type 1) within the species Aichivirus A. To date, there are no information on the epidemiology of these novel viruses in other continents. In this study, by screening a collection of 256 dog fecal samples either from diarrhoeic or asymptomatic animals, CaKV was identified in six specimens with an overall prevalence of 2.34% (6/256). All the positive dogs presented diarrhea and were found to be infected by CaKV alone or in mixed infections with canine coronavirus (CCoV) and/or canine parvovirus type 2 (CPV-2). By molecular analysis of the partial 3D gene, all the strains detected displayed a close relatedness with the CaKVs recently identified in the US. This study provides evidence that CaKVs circulate in diarrhoeic dogs in Italy and are not geographically restricted to the North American continent, where they were first signaled. PMID:23806200

  3. Cytodiagnostics of canine lymphomas - possibilities and limitations.

    PubMed

    Sapierzyński, R; Kliczkowska-Klarowicz, K; Jankowska, U; Jagielski, D

    2016-01-01

    Malignant lymphomas are one of the most common malignant tumours occurring in dogs. The basic method of lymphoma diagnosis in human, as well as in canine oncology is histopathology supported by immunohistochemistry. It was suggested that in veterinary medicine excisional biopsy of lymph node and histopathology should be considered only where the cytologic diagnosis is equivocal or needs to be confirmed. There are at least three basic reasons for which cytological examination ought to be accepted as a sufficient and reliable diagnostic method for lymphoma in dogs. Firstly, most dog owners consider the fine-needle biopsy as an acceptable non-harmful method of sample collection. Secondly, an increasing number of studies recommend cytology as an accurate test for diagnosing and subtyping canine lymphoma. Finally, the vast majority of canine lymphoma subtypes belong to 4-5 categories characterized by a typical cytological picture. Immunocytochemical staining of cytological smears gives new diagnostic possibilities, such as detection of markers better characterizing given growth or a potential goal for target therapy in individual cases (for example inhibitors of platelet-derived growth factor). PMID:27487521

  4. Increasing incidence of canine leptospirosis in Switzerland.

    PubMed

    Major, Andrea; Schweighauser, Ariane; Francey, Thierry

    2014-07-01

    A marked increase in canine leptospirosis was observed in Switzerland over 10 years with a peak incidence of 28.1 diagnosed cases/100,000 dogs/year in the most affected canton. With 95% affected dogs living at altitudes <800 m, the disease presented a seasonal pattern associated with temperature (r2 0.73) and rainfall (r2 0.39), >90% cases being diagnosed between May and October. The increasing yearly incidence however was only weakly correlated with climatic data including number of summer (r2 0.25) or rainy days (r2 0.38). Serovars Australis and Bratislava showed the highest seropositivity rates with 70.5% and 69.1%, respectively. Main clinical manifestations included renal (99.6%), pulmonary (76.7%), hepatic (26.0%), and hemorrhagic syndromes (18.2%), leading to a high mortality rate (43.3%). Similar to the human disease, liver involvement had the strongest association with negative outcome (OR 16.3). Based on these data, canine leptospirosis presents similar features and severity as the human infection for which it therefore can be considered a model. Its re-emergence in a temperate country with very high incidence rates in canines should thus be viewed as a warning and emphasize the need for increased awareness in other species. PMID:25032740

  5. Mechanisms of cell death in canine parvovirus-infected cells provide intuitive insights to developing nanotools for medicine.

    PubMed

    Nykky, Jonna; Tuusa, Jenni E; Kirjavainen, Sanna; Vuento, Matti; Gilbert, Leona

    2010-01-01

    Viruses have great potential as nanotools in medicine for gene transfer, targeted gene delivery, and oncolytic cancer virotherapy. Here we have studied cell death mechanisms of canine parvovirus (CPV) to increase the knowledge on the CPV life cycle in order to facilitate the development of better parvovirus vectors. Morphological studies of CPV-infected Norden laboratory feline kidney (NLFK) cells and canine fibroma cells (A72) displayed characteristic apoptotic events. Apoptosis was further confirmed by activation of caspases and cellular DNA damage. However, results from annexin V-propidium iodide (PI) labeling and membrane polarization assays indicated disruption of the plasma membrane uncommon to apoptosis. These results provide evidence that secondary necrosis followed apoptosis. In addition, two human cancer cell lines were found to be infected by CPV. This necrotic event over apoptotic cell death and infection in human cells provide insightful information when developing CPV as a nanotool for cancer treatments. PMID:20957163

  6. Mechanisms of cell death in canine parvovirus-infected cells provide intuitive insights to developing nanotools for medicine

    PubMed Central

    Nykky, Jonna; Tuusa, Jenni E; Kirjavainen, Sanna; Vuento, Matti; Gilbert, Leona

    2010-01-01

    Viruses have great potential as nanotools in medicine for gene transfer, targeted gene delivery, and oncolytic cancer virotherapy. Here we have studied cell death mechanisms of canine parvovirus (CPV) to increase the knowledge on the CPV life cycle in order to facilitate the development of better parvovirus vectors. Morphological studies of CPV-infected Norden laboratory feline kidney (NLFK) cells and canine fibroma cells (A72) displayed characteristic apoptotic events. Apoptosis was further confirmed by activation of caspases and cellular DNA damage. However, results from annexin V-propidium iodide (PI) labeling and membrane polarization assays indicated disruption of the plasma membrane uncommon to apoptosis. These results provide evidence that secondary necrosis followed apoptosis. In addition, two human cancer cell lines were found to be infected by CPV. This necrotic event over apoptotic cell death and infection in human cells provide insightful information when developing CPV as a nanotool for cancer treatments. PMID:20957163

  7. 9 CFR 113.306 - Canine Distemper Vaccine.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... AGRICULTURE VIRUSES, SERUMS, TOXINS, AND ANALOGOUS PRODUCTS; ORGANISMS AND VECTORS STANDARD REQUIREMENTS Live Virus Vaccines § 113.306 Canine Distemper Vaccine. Canine Distemper Vaccine shall be prepared from virus-bearing cell culture fluids or embryonated chicken eggs. Only Master Seed Virus which has been...

  8. 9 CFR 113.306 - Canine Distemper Vaccine.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... AGRICULTURE VIRUSES, SERUMS, TOXINS, AND ANALOGOUS PRODUCTS; ORGANISMS AND VECTORS STANDARD REQUIREMENTS Live Virus Vaccines § 113.306 Canine Distemper Vaccine. Canine Distemper Vaccine shall be prepared from virus-bearing cell culture fluids or embryonated chicken eggs. Only Master Seed Virus which has been...

  9. 9 CFR 113.306 - Canine Distemper Vaccine.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... AGRICULTURE VIRUSES, SERUMS, TOXINS, AND ANALOGOUS PRODUCTS; ORGANISMS AND VECTORS STANDARD REQUIREMENTS Live Virus Vaccines § 113.306 Canine Distemper Vaccine. Canine Distemper Vaccine shall be prepared from virus-bearing cell culture fluids or embryonated chicken eggs. Only Master Seed Virus which has been...

  10. 9 CFR 113.306 - Canine Distemper Vaccine.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... AGRICULTURE VIRUSES, SERUMS, TOXINS, AND ANALOGOUS PRODUCTS; ORGANISMS AND VECTORS STANDARD REQUIREMENTS Live Virus Vaccines § 113.306 Canine Distemper Vaccine. Canine Distemper Vaccine shall be prepared from virus-bearing cell culture fluids or embryonated chicken eggs. Only Master Seed Virus which has been...

  11. Sex differences in anthropoid mandibular canine lateral enamel formation.

    PubMed

    Guatelli-Steinberg, Debbie; Ferrell, Rebecca J; Spence, Jennifer; Talabere, Tiffany; Hubbard, Amelia; Schmidt, Stacey

    2009-10-01

    Previous research has demonstrated that great ape and macaque males achieve large canine crown sizes primarily through extended canine growth periods. Recent work has suggested, however, that platyrrhine males may achieve larger canine sizes by accelerating rather than prolonging growth. This study tested the hypothesis that the ontogenetic pathway leading to canine sexual dimorphism in catarrhines differs from that of platyrrhines. To test this hypothesis, males and females of several catarrhine genera (Hylobates, Papio, Macaca, Cercopithecus, and Cercocebus) and three platyrrhine genera (Cebus, Ateles, and Callicebus) were compared in the number and spacing of perikymata (enamel growth increments) on their canine crowns. In addition, perikymata periodicities (the number of days of growth perikymata represent) were determined for five genera (Hylobates, Papio, Macaca, Cebus, and Ateles) using previously published as well as original data gathered for this study. The central findings are as follows: 1) males have more perikymata than females for seven of eight genera (in five of the seven, the differences are statistically significant); 2) in general, the greater the degree of sexual dimorphism, the greater the sex difference in male and female perikymata numbers; 3) there is no evidence of a systematic sex difference in primate periodicities; and 4) there is some evidence that sex differences in enamel formation rates may make a minor contribution to canine sexual dimorphism in Papio and Cercopithecus. These findings strongly suggest that in both catarrhines and platyrrhines prolongation of male canine growth is the primary mechanism by which canine crown sexual dimorphism is achieved. PMID:19350641

  12. First detection of canine parvovirus type 2c in Brazil

    PubMed Central

    Streck, André Felipe; de Souza, Carine Kunzler; Gonçalves, Karla Rathje; Zang, Luciana; Pinto, Luciane Dubina; Canal, Cláudio Wageck

    2009-01-01

    The presence of canine parvovirus type 2 (CPV-2), 2a and 2b has been described in Brazil, however, the type 2c had not been reported until now. In the current study, seven out of nine samples from dogs with diarrhea were characterized as CPV-2c, indicating that this virus is already circulating in the Brazilian canine population. PMID:24031389

  13. 9 CFR 113.306 - Canine Distemper Vaccine.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 9 Animals and Animal Products 1 2010-01-01 2010-01-01 false Canine Distemper Vaccine. 113.306 Section 113.306 Animals and Animal Products ANIMAL AND PLANT HEALTH INSPECTION SERVICE, DEPARTMENT OF AGRICULTURE VIRUSES, SERUMS, TOXINS, AND ANALOGOUS PRODUCTS; ORGANISMS AND VECTORS STANDARD REQUIREMENTS Live Virus Vaccines § 113.306 Canine...

  14. Canine evolution in sabretoothed carnivores: natural selection or sexual selection?

    PubMed

    Randau, Marcela; Carbone, Chris; Turvey, Samuel T

    2013-01-01

    The remarkable elongated upper canines of extinct sabretoothed carnivorous mammals have been the subject of considerable speculation on their adaptive function, but the absence of living analogues prevents any direct inference about their evolution. We analysed scaling relationships of the upper canines of 20 sabretoothed feliform carnivores (Nimravidae, Barbourofelidae, Machairodontinae), representing both dirk-toothed and scimitar-toothed sabretooth ecomorphs, and 33 non-sabretoothed felids in relation to body size in order to characterize and identify the evolutionary processes driving their development, using the scaling relationships of carnassial teeth in both groups as a control. Carnassials display isometric allometry in both sabretooths and non-sabretooths, supporting their close relationship with meat-slicing, whereas the upper canines of both groups display positive allometry with body size. Whereas there is no statistical difference in allometry of upper canine height between dirk-toothed and scimitar-toothed sabretooth ecomorphs, the significantly stronger positive allometry of upper canine height shown by sabretooths as a whole compared to non-sabretooths reveals that different processes drove canine evolution in these groups. Although sabretoothed canines must still have been effective for prey capture and processing by hypercarnivorous predators, canine morphology in these extinct carnivores was likely to have been driven to a greater extent by sexual selection than in non-sabretooths. Scaling relationships therefore indicate the probable importance of sexual selection in the evolution of the hypertrophied sabretooth anterior dentition. PMID:23951334

  15. Elastase Deficiency Phenotype of Pseudomonas aeruginosa Canine Otitis Externa Isolates

    PubMed Central

    Petermann, Shana R.; Doetkott, Curt; Rust, Lynn

    2001-01-01

    Pseudomonas aeruginosa veterinary isolates were assayed for elastase and total matrix protease activity. The elastase activity of canine ear isolates was much less than that of strain PAO1 and that of all other veterinary isolates (P < 0.0001). The results indicate that canine ear isolates have a distinct elastase phenotype. PMID:11329471

  16. Cutaneous immune mechanisms in canine leishmaniosis due to Leishmania infantum.

    PubMed

    Papadogiannakis, E I; Koutinas, A F

    2015-02-15

    Canine leishmaniosis (CanL) caused by the parasite Leishmania infantum is a systemic disease with variable clinical signs. The disease is endemic in the Mediterranean countries and dogs are the main domestic reservoir of the parasite. The quite complicated immune response against the parasite is crucial for the evolution of CanL infection with the skin playing a major role in its immunopathogenesis. After the inoculation of Leishmania promastigotes into the dermis by sand fly bites, complement factors, Langerhan's cells, neutrophils, fibroblasts and keratinocytes are involved in the activation of the innate arm of the skin immune system, with the macrophages and dendritic cells to play a major key role. The effective activation of cellular immunity is the cornerstone of dog's resistance against the parasite. Promastigotes reaching the dermis are engulfed, processed and transferred by APCs to draining lymph nodes to stimulate naïve T-cells for proliferation and differentiation into armed effector T-cells. Th1 cells activate the infected macrophages to kill Leishmania, whereas Th2 cells divert the immune response to humoral immunity and down regulation of cellular immunity with Th1 cell anergy. Inhibition of co-stimulatory molecules expression by infected macrophages contributes to T-cell anergy. In canine subclinical infections cutaneous lymphocytic infiltrate and parasites are absent, as opposed to dogs with clinical leishmaniosis. CD8+ cells constitute a significant population of cellular immunity in CanL since they outnumber CD4+ cells in the dermis, producing IFN-γ in sub clinically infected dogs and high levels of IL-4 in dogs with clinical leishmaniosis. Numerous B-lymphocytes have been shown to heavily infiltrate the dermis at least in exfoliative dermatitis in CanL. A mixed Th1/Th2 cytokine profile has been found in the dermis of naturally infected with L. infantum dogs. In the skin of dogs with clinical leishmaniosis, where plasma cells outnumber T

  17. Cyclooxygenase-2 (COX-2) expression in canine intracranial meningiomas.

    PubMed

    Rossmeisl, J H; Robertson, J L; Zimmerman, K L; Higgins, M A; Geiger, D A

    2009-09-01

    Meningiomas are the most common canine intracranial tumour. Neurologic disability and death from treatment failure remain problematic despite current surgical and radiotherapeutic treatments for canine intracranial meningiomas. Cyclooxygenase-2 (COX-2) over-expression has been demonstrated in multiple canine malignancies, and COX-2 inhibitory treatment strategies have been shown to have both preventative and therapeutic effects in spontaneous and experimental models of cancer. The purpose of this study was to evaluate COX-2 expression in canine intracranial meningiomas. Immunohistochemical and Western blot (WB) analyses showed COX-2 expression in multiple tissues of the normal canine brain, and 87% (21/24) of intracranial meningiomas studied were immunoreactive to COX-2. No significant associations between COX-2 immunoreactivity and tumour grade were identified. Further studies are required to elucidate the physiologic roles of constitutive COX-2 expression in the central nervous system as well as its participation in meningioma tumourigenesis. PMID:19691646

  18. Apicotomy: surgical management of maxillary dilacerated or ankylosed canines.

    PubMed

    Araújo, Eustáquio A; Araújo, Cristiana V; Tanaka, Orlando M

    2013-12-01

    This clinical article reports a technique, apicotomy, for managing dilacerated or ankylosed canines. The records of 3 patients successfully treated with apicotomy are presented. Orthodontists observe clinically significant incidences of impacted maxillary canines in their daily practices. Several procedures have been described to bring an ankylosed, impacted tooth into occlusion. Luxation is the most widely used solution, but there are risks involved with that approach, and the success rate is low. Surgical repositioning has also been used, but morbidity is high, and the aggressiveness of the procedure might also contraindicate it. Ankylosis might be related to the anatomic position of the canine's root apex and its adjacent anatomic structures. Apicotomy is a guided fracture of a canine root apex, followed by its orthodontic traction. It is a conservative surgical alternative for treating impacted canines with dilacerations or apical root ankylosis. PMID:24286914

  19. The effects of oncolytic reovirus in canine lymphoma cell lines.

    PubMed

    Hwang, C C; Umeki, S; Igase, M; Coffey, M; Noguchi, S; Okuda, M; Mizuno, T

    2016-08-01

    Reovirus is a potent oncolytic virus in many human neoplasms that has reached phase II and III clinical trials. Our laboratory has previously reported the oncolytic effects of reovirus in canine mast cell tumour (MCT). In order to further explore the potential of reovirus in veterinary oncology, we tested the susceptibility of reovirus in 10 canine lymphoma cell lines. Reovirus-induced cell death, virus replication and infectivity were confirmed in four cell lines with variable levels of susceptibility. The level of Ras activation varied among the cell lines with no correlation with reovirus susceptibility. Reovirus-susceptible cell lines underwent apoptosis as proven by propidium iodide (PI) staining, Annexin V-FITC/PI assay, cleavage of PARP and inhibition of cell death by caspase inhibitor. A single intratumoral injection of reovirus suppressed the growth of canine lymphoma subcutaneous tumour in NOD/SCID mice. Unlike canine MCT, canine lymphoma is less susceptible to reovirus. PMID:25319493

  20. Recombinant canine distemper virus serves as bivalent live vaccine against rabies and canine distemper.

    PubMed

    Wang, Xijun; Feng, Na; Ge, Jinying; Shuai, Lei; Peng, Liyan; Gao, Yuwei; Yang, Songtao; Xia, Xianzhu; Bu, Zhigao

    2012-07-20

    Effective, safe, and affordable rabies vaccines are still being sought. Attenuated live vaccine has been widely used to protect carnivores from canine distemper. In this study, we generated a recombinant canine distemper virus (CDV) vaccine strain, rCDV-RVG, expressing the rabies virus glycoprotein (RVG) by using reverse genetics. The recombinant virus rCDV-RVG retained growth properties similar to those of vector CDV in Vero cell culture. Animal studies demonstrated that rCDV-RVG was safe in mice and dogs. Mice inoculated intracerebrally or intramuscularly with rCDV-RVG showed no apparent signs of disease and developed a strong rabies virus (RABV) neutralizing antibody response, which completely protected mice from challenge with a lethal dose of street virus. Canine studies showed that vaccination with rCDV-RVG induced strong and long-lasting virus neutralizing antibody responses to RABV and CDV. This is the first study demonstrating that recombinant CDV has the potential to serve as bivalent live vaccine against rabies and canine distemper in animals. PMID:22698451

  1. Breed predisposition to canine gastric carcinoma - a study based on the Norwegian canine cancer register

    PubMed Central

    2013-01-01

    Background Previous research has indicated a breed predisposition to gastric carcinoma in dogs. However, results to date are inconsistent since several studies have failed to prove such a predisposition. Better knowledge of breeds at risk could facilitate early detection of gastric carcinoma in dogs. The aim of the study was to retrospectively investigate the proportion and possible breed predisposition to canine gastric carcinoma using the Norwegian Canine Cancer Register for calculations of proportional morbidity ratios (PMRs) for the period 1998–2009. Results Histologically verified tumours recorded in the Norwegian Canine Cancer Register were studied (n = 19,715). A total of 31 (0.16%) cases of canine gastric carcinomas were identified. The median age of affected dogs was 10 years. The most commonly reported clinical signs were vomiting, anorexia, and weight loss. Males had significantly higher odds of gastric carcinoma than females (P = 0.02). The PMR with 95% confidence interval (CI) was calculated for each breed, and a breed predisposition was identified. Individuals of the breeds Tervuren (PMR 56.1), Bouvier des Flandres (PMR 36.5), Groenendael (PMR 34.5), Collie (PMR 26.1), Standard poodle (PMR 7.6), and Norwegian elkhound (PMR 6.1) had a significantly increased risk of developing gastric carcinoma. Discussion and conclusion The proportion of cases of gastric carcinoma recorded in the Norwegian Canine Cancer Register was found to be 0.16%, and a breed predisposition was identified. The breed predisposition observed in the current study indicates a genetic susceptibility to gastric carcinoma. PMID:23514604

  2. Polar Stratigraphy

    NASA Technical Reports Server (NTRS)

    1999-01-01

    These three images were taken on three different orbits over the north polar cap in April 1999. Each shows a different part of the same ice-free trough. The left and right images are separated by a distance of more than 100 kilometers (62 miles). Note the similar layers in each image.

  3. Political polarization

    PubMed Central

    Dixit, Avinash K.; Weibull, Jörgen W.

    2007-01-01

    Failures of government policies often provoke opposite reactions from citizens; some call for a reversal of the policy, whereas others favor its continuation in stronger form. We offer an explanation of such polarization, based on a natural bimodality of preferences in political and economic contexts and consistent with Bayesian rationality. PMID:17452633

  4. Canine hip extension range during gait.

    PubMed

    van der Walt, A M; Stewart, A V; Joubert, K E; Bekker, P

    2008-12-01

    Assessment of canine gait is frequently used by veterinary clinicians to establish the presence of orthopaedic pain. As up to 30% of canine orthopaedic conditions affect the pelvic limb, knowledge of pelvic limb biomechanics during gait is very important. Previous studies have investigated the biomechanics at the tarsus and stifle, but little information is available regarding hip motion during gait. The aim of this study was to determine the maximum hip extension range achieved during the stance phase of gait in normal canines. In addition, this study aimed to determine the difference between maximum passive hip extension and maximum hip extension during gait. Using a sample of 30 morphologically similar normal dogs, mean maximum passive hip extension was measured using a goniometer and mean maximum hip extension range during gait was determined videographically. Inter- and intra-assessor reliability studies performed at the start of the study showed that the measurement tools and techniques used in this study were valid and reliable. The goniometric data showed that mean maximum passive hip extension range was 162.44 degrees (+/-3.94) with no significant difference between the left and the right hind limbs. The videographic data showed that mean maximum hip extension range during gait was 119.9 degrees (+/-9.26) with no significant difference between the left and right hind limbs. The results of this study provided reference values for active and passive hip extension range and showed that the degree of hip extension range required for normal gait is significantly less than maximum passive hip extension range. PMID:19496317

  5. Extraglandular and intraglandular vascularization of canine prostate.

    PubMed

    Stefanov, Miroslav

    2004-03-01

    The literature on the vascularization of the canine prostate is reviewed and the clinical significance of prostate morphology is described. Scanning Electron Microscopy (SEM), combined with improved corrosion casting methods, reveal new morphological details that promise better diagnostics and treatment but also require expansion of clinical nomenclature. A proposal is made for including two previously unnamed veins in Nomina Anatomica Veterinaria (NAV). The canine prostate has two lobes with independent vascularization. Each lobe is supplied through the left and right a. prostatica, respectively. The a. prostatica sprouts three small vessels (cranial, middle, and caudal) towards the prostate gland. A. prostatica is a small-size artery whose wall structure is similar to the arteries of the muscular type. V. prostatica is a small-size valved vein. The canine prostate has capsular, parenchymal, and urethral vascular zones. The surface vessels of the capsule are predominantly veins and the diameter of arterial vessels is larger than that of the veins. The trabecular vessels are of two types: direct and branched. The prostate parenchyma is supplied by branches of the trabecular vessels. The periacinary capillaries are fenestrated and form a net in a circular pattern. The processes of the myoepithelial cells embrace both the acins and the periacinar capillaries. In the prostate ductal system. there are spermatozoa. The prostatic part of the urethra is supplied by an independent branch of a. prostatica. The prostatic urethral part is drained by v. prostatica, the vein of the urethral bulb and the ventral prostate veins. M. urethralis begins as early as the urethral prostatic part. The greater part of the white muscle fibers in m. urethralis suggest an enhanced anaerobic metabolism. PMID:14988915

  6. Canine blood groups: description of 20 specificities.

    PubMed

    Symons, M; Bell, K

    1992-01-01

    Twenty blood typing reagents, four agglutinins and 16 operable in the antiglobulin test, were prepared from 54 antisera which were produced in 24 dogs. Two of the reagents were identified as anti-B and Nf6. Two of the antigens were shown by absorption and family studies to be linear subtypes. In most cases, detailed family studies demonstrated a Mendelian dominant inheritance for the genes controlling the canine red cell antigens. Gene frequencies were determined in various breeds of dogs and in the dingo. PMID:1492701

  7. Functional characterization of canine interferon-lambda.

    PubMed

    Fan, Wenhui; Xu, Lei; Ren, Liqian; Qu, Hongren; Li, Jing; Liang, Jingjing; Liu, Wenjun; Yang, Limin; Luo, Tingrong

    2014-11-01

    In this study, we provide the first comprehensive annotation of canine interferon-λ (CaIFN-λ, type III IFN). Phylogenetic analysis based on genomic sequences indicated that CaIFN-λ is located in the same branch with Swine IFN-λ1 (SwIFN-λ), Bat IFN-λ1 (BaIFN-λ), and human IFN-λ1 (HuIFN-λ1). CaIFN-λ was cloned, expressed in Escherichia coli, and purified to further investigate the biological activity in vitro. The recombinant CaIFN-λ (rCaIFN-λ) displayed potent antiviral activity on both homologous and heterologous animal cells in terms of inhibiting the replication of the New Jersey serotype of vesicular stomatitis virus (VSV), canine parvovirus, and influenza virus A/WSN/33 (H1N1), respectively. In addition, we also found that rCaIFN-λ exhibits a significant antiproliferative response against A72 canine tumor cells and MDCK cells in a dose-dependent manner. Furthermore, CaIFN-λ activated the JAK-STAT signaling pathway. To evaluate the expression of CaIFN-λ induced by virus and the expression of IFN-stimulated genes (ISGs) induced by rCaIFN-λ in the MDCK cells, we measured the relative mRNA level of CaIFN-λ and ISGs (ISG15, Mx1, and 2'5'-OAS) by quantitative real-time PCR and found that the mRNA level of CaIFN-λ and the ISGs significantly increased after treating the MDCK cells with viruses and rCaIFN-λ protein, respectively. Finally, to evaluate the binding activity of rCaIFN-λ to its receptor, we expressed the extracellular domain of the canine IFN-λ receptor 1 (CaIFN-λR1-EC) and determined the binding activity via ELISA. Our results demonstrated that rCaIFN-λ bound tightly to recombinant CaIFN-λR1-EC (rCaIFN-λR1-EC). PMID:24950142

  8. Diagnosis and management of canine claw diseases.

    PubMed

    Mueller, R S

    1999-11-01

    The diagnostic workup for canine claw disease consists of a good history and complete clinical examination which may provide clues for a possible underlying disorder. In dogs with claw disease but no other clinical or historical signs, further recommended diagnostic procedures include cytological evaluation of impression smears or discharge from the claw fold, bacterial culture and sensitivity testing, biopsy of the claw matrix, and an elimination diet for 6 to 8 weeks. If no underlying disease can be identified, trial treatment with essential fatty acid supplementation, vitamin E, or a combination of doxycycline hydrochloride and niacinamide may be useful. In some patients, onychectomy of all claws may be considered. PMID:10563005

  9. Definition, Classification, and Pathophysiology of Canine Glaucoma.

    PubMed

    Pizzirani, Stefano

    2015-11-01

    Glaucoma is a common ocular condition in humans and dogs leading to optic nerve degeneration and irreversible blindness. Primary glaucoma is a group of spontaneous heterogeneous diseases. Multiple factors are involved in its pathogenesis and these factors vary across human ethnic groups and canine breeds, so the clinical phenotypes are numerous and their classification can be challenging and remain superficial. Aging and oxidative stress are major triggers for the manifestation of disease. Multiple, intertwined inflammatory and biochemical cascades eventually alter cellular and extracellular physiology in the optic nerve and trabecular meshwork and lead to vision loss. PMID:26456751

  10. The Genetics of Canine Skull Shape Variation

    PubMed Central

    Schoenebeck, Jeffrey J.; Ostrander, Elaine A.

    2013-01-01

    A dog’s craniofacial diversity is the result of continual human intervention in natural selection, a process that began tens of thousands of years ago. To date, we know little of the genetic underpinnings and developmental mechanisms that make dog skulls so morphologically plastic. In this Perspectives, we discuss the origins of dog skull shapes in terms of history and biology and highlight recent advances in understanding the genetics of canine skull shapes. Of particular interest are those molecular genetic changes that are associated with the development of distinct breeds. PMID:23396475

  11. Canine congenital portosystemic shunts: Disconnections dissected.

    PubMed

    Van den Bossche, L; van Steenbeek, F G

    2016-05-01

    Canine congenital portosystemic shunts (CPSS) are vascular anomalies that connect the portal vein with the systemic circulation, therefore bypassing the hepatic parenchyma. Portosystemic shunts exist in two different subtypes: extrahepatic and intrahepatic. This congenital disorder is also described in mice, cat, sheep and man. Research has been focused on pathophysiology, diagnostics and treatment of CPSS and this has resulted in increased knowledge, although the aetiology of the disease remains unclear. This review focuses on the aetiology and genetic basis of both intra- and extrahepatic shunts. PMID:27061656

  12. Cone-beam computed tomography findings of impacted upper canines

    PubMed Central

    Bastos, Luana Costa; Oliveira-Santos, Christiano; da Silva, Silvio José Albergaria; Neves, Frederico Sampaio; Campos, Paulo Sérgio Flores

    2014-01-01

    Purpose To describe the features of impacted upper canines and their relationship with adjacent structures through three-dimensional cone-beam computed tomography (CBCT) images. Materials and Methods Using the CBCT scans of 79 upper impacted canines, we evaluated the following parameters: gender, unilateral/bilateral occurrence, location, presence and degree of root resorption of adjacent teeth (mild, moderate, or severe), root dilaceration, dental follicle width, and presence of other associated local conditions. Results Most of the impacted canines were observed in females (56 cases), unilaterally (51 cases), and at a palatine location (53 cases). Root resorption in adjacent teeth and root dilaceration were observed in 55 and 47 impacted canines, respectively. In most of the cases, the width of the dental follicle of the canine was normal; it was abnormally wide in 20 cases. A statistically significant association was observed for all variables, except for root dilaceration (p=0.115) and the side of impaction (p=0.260). Conclusion Root resorption of adjacent teeth was present in most cases of canine impaction, mostly affecting adjacent lateral incisors to a mild degree. A wide dental follicle of impacted canines was not associated with a higher incidence of external root resorption of adjacent teeth. PMID:25473636

  13. Stem Cell-Associated Marker Expression in Canine Hair Follicles.

    PubMed

    Gerhards, Nora M; Sayar, Beyza S; Origgi, Francesco C; Galichet, Arnaud; Müller, Eliane J; Welle, Monika M; Wiener, Dominique J

    2016-03-01

    Functional hair follicle (HF) stem cells (SCs) are crucial to maintain the constant recurring growth of hair. In mice and humans, SC subpopulations with different biomarker expression profiles have been identified in discrete anatomic compartments of the HF. The rare studies investigating canine HF SCs have shown similarities in biomarker expression profiles to that of mouse and human SCs. The aim of our study was to broaden the current repertoire of SC-associated markers and their expression patterns in the dog. We combined analyses on the expression levels of CD34, K15, Sox9, CD200, Nestin, LGR5 and LGR6 in canine skin using RT-qPCR, the corresponding proteins in dog skin lysates, and their expression patterns in canine HFs using immunohistochemistry. Using validated antibodies, we were able to define the location of CD34, Sox9, Keratin15, LGR5 and Nestin in canine HFs and confirm that all tested biomarkers are expressed in canine skin. Our results show similarities between the expression profile of canine, human and mouse HF SC markers. This repertoire of biomarkers will allow us to conduct functional studies and investigate alterations in the canine SC compartment of different diseases, like alopecia or skin cancer with the possibility to extend relevant findings to human patients. PMID:26739040

  14. Rapid maxillary canine retraction by dental distraction: A clinical study

    PubMed Central

    Koteswara Prasad, N. K.; Chitharanjan, Arun; Kailasam, Vignesh

    2014-01-01

    Aim: The aim of this clinical study was to perform rapid maxillary canine retraction through distraction of the periodontal ligament and investigate the rate and amount of canine retraction, amount of anchor loss, the nature of tooth movement achieved, and radiographic changes in the periodontal ligament region during and after canine distraction. Materials and Methods: This study was conducted on 10 distractions ranging in age from 14 years to 25 years who needed canine retraction and first premolar extraction in the maxillary arch. Ten canine distractions were carried out with custom-made, tooth-borne intra-oral distraction device. Results: The results indicate that the periodontal ligament can be distracted just like the mid-palatal suture in rapid palatal expansion and the maxillary canines are retracted rapidly into the first premolar extraction space at the rate of about 2.53 mm/week. Conclusion: Though this study indicates that the periodontal ligament can be distracted to elicit rapid tooth movement, the long-term effects of canine distraction are not well known and need close monitoring. PMID:25298710

  15. Cell polarity

    PubMed Central

    Romereim, Sarah M

    2011-01-01

    Despite extensive genetic analysis of the dynamic multi-phase process that transforms a small population of lateral plate mesoderm into the mature limb skeleton, the mechanisms by which signaling pathways regulate cellular behaviors to generate morphogenetic forces are not known. Recently, a series of papers have offered the intriguing possibility that regulated cell polarity fine-tunes the morphogenetic process via orienting cell axes, division planes and cell movements. Wnt5a-mediated non-canonical signaling, which may include planar cell polarity, has emerged as a common thread in the otherwise distinct signaling networks that regulate morphogenesis in each phase of limb development. These findings position the limb as a key model to elucidate how global tissue patterning pathways direct local differences in cell behavior that, in turn, generate growth and form. PMID:22064549

  16. Osteocalcin and Osteonectin Expression in Canine Osteosarcoma.

    PubMed

    Wehrle-Martinez, A S; Dittmer, K E; Aberdein, D; Thompson, K G

    2016-07-01

    Osteosarcoma (OSA) is a malignant heterogeneous primary bone tumor responsible for up to 90% of all primary bone tumors in dogs. In this study, osteocalcin (OC) and osteonectin (ON) immunoreactivity was evaluated in 23 canine OSAs, 4 chondrosarcomas, 4 fibrosarcomas, 2 hemangiosarcomas, and 4 histiocytic sarcomas. The effects of three different decalcification agents (ethylenediaminetetraetic acid [EDTA], formic acid and hydrochloric acid [HCl]) on the immunoreactivity for OC and ON was also assessed. Immunoreactivity to OC was present in 19/23 (83%) cases of OSA and all cases of chondrosarcoma. In three OSAs the extracellular matrix showed immunoreactivity to OC. None of the fibrosarcomas, histiocytic sarcomas or hemangiosarcomas showed immunoreactivity to OC. The sensitivity and specificity for OC in canine OSA in this study was 83% and 71% respectively. For ON, 100% of both OSAs (23/23) and non-OSAs (14/14) showed cytoplasmic immunoreactivity to this antibody, giving a sensitivity of 100% but a complete lack of specificity. There were no significant differences in immunoreactivity for OC and ON between the different decalcification agents used. In conclusion, OC showed high sensitivity for identifying OSA but it failed to distinguish between OSA and chondrosarcoma, and the osteoid produced by neoplastic cells in most cases did not show immunoreactivity to OC. These factors may limit the practical utility of OC in the diagnosis of OSA in dogs when chondrosarcoma is a differential diagnosis. ON showed no specificity in detecting OSA and has little practical application for the diagnosis of OSA in dogs. PMID:26926085

  17. The Evolutionary Processes of Canine Coronaviruses

    PubMed Central

    Pratelli, Annamaria

    2011-01-01

    Since the first identification of the virus in 1971, the disease caused by canine coronavirus (CCoV) has not been adequately investigated, and the role that the virus plays in canine enteric illness has not been well established. Only after the emergence in 2002 of SARS in human has new attention been focused on coronaviruses. As a consequence of the relatively high mutation frequency of RNA-positive stranded viruses, CCoV has evolved and, with the biomolecular techniques developed over the last two decades, new virus strains, serotypes, and subtypes have been identified in infected dogs. Considering the widespread nature of CCoV infections among dog populations, several studies have been carried out, focusing upon the epidemiological relevance of these viruses and underlining the need for further investigation into the biology of CCoVs and into the pathogenetic role of the infections. This paper reports the evolutionary processes of CCoVs with a note onto recent diagnostic methods. PMID:22315601

  18. Immunology and pathogenesis of canine demodicosis.

    PubMed

    Ferrer, Lluis; Ravera, Ivan; Silbermayr, Katja

    2014-10-01

    Demodex mites colonized the hair follicles and sebaceous glands of mammals millions of years ago and have remained relatively unchanged in this protected ecologic niche since then. The host immune system detects and tolerates their presence. Toll-like receptor-2 of keratinocytes has been demonstrated to recognize mite chitin and to elicit an innate immune response. The subsequent acquired immune response is poorly understood at present, but there is experimental and clinical evidence that this is the main mechanism in the control of mite proliferation. A transgenic mouse model (STAT(-/-) /CD28(-/-) ) has demonstrated that the immune response is complex, probably involving both cellular and humoral mechanisms and requiring the role of co-stimulatory molecules (CD28). It is known that a genetic predisposition for developing canine juvenile generalized demodicosis exists; however, the primary defect leading to the disease remains unknown. Once the mite proliferation is advanced, dogs show a phenotype that is similar to the T-cell exhaustion characterized by low interleukin-2 production and high interleukin-10 and transforming growth factor-β production by lymphocytes, as described in other viral and parasitic diseases. Acaricidal treatment (macrocyclic lactones) decreases the antigenic load and reverses T-cell exhaustion, leading to a clinical cure. Although in recent years there have been significant advances in the management and understanding of this important and complex canine disease, more research in areas such as the aetiology of the genetic predisposition and the immune control of the mite populations is clearly needed. PMID:24910252

  19. Diagnostic immunohistochemistry of canine round cell tumors.

    PubMed

    Sandusky, G E; Carlton, W W; Wightman, K A

    1987-11-01

    Sixty-five canine skin neoplasms studied using immunocytochemistry, included 22 histiocytomas, 18 amelanotic melanomas, 14 cutaneous lymphosarcomas, six mast cell tumors, and five transmissible venereal tumors. Formalin-fixed, paraffin-embedded sections were stained using the avidin-biotin-peroxidase complex (ABC) immunoperoxidase technique for reactivity with S-100 protein, kappa and lambda immunoglobulin light chains, alpha-1-antitrypsin, alpha-1-antichymotrypsin, leukocyte common antigen (LCA), neuron-specific enolase, keratin, cytokeratin, muramidase, and vimentin. Detection of S-100, kappa and lambda light chains, neuron-specific enolase, and vimentin were most useful for screening these neoplasms. None of the markers examined was consistent in staining histiocytomas. While reactivity of S-100 (ten cases) and neuron-specific enolase (ten cases) was detected in some amelanotic melanomas, lambda light chain immunoglobulin (eight cases) was relatively consistent in cutaneous lymphomas. Mast cell neoplasms reacted with avidin and, therefore, were positive, even on negative control sections. Vimentin reacted strongly on all amelanotic melanomas and transmissible venereal tumors examined. These antibodies are helpful adjuncts in the differential diagnosis of canine skin tumors. PMID:3137715

  20. Intracellular Route of Canine Parvovirus Entry

    PubMed Central

    Vihinen-Ranta, Maija; Kalela, Anne; Mäkinen, Päivi; Kakkola, Laura; Marjomäki, Varpu; Vuento, Matti

    1998-01-01

    The present study was designed to investigate the endocytic pathway involved in canine parvovirus (CPV) infection. Reduced temperature (18°C) or the microtubule-depolymerizing drug nocodazole was found to inhibit productive infection of canine A72 cells by CPV and caused CPV to be retained in cytoplasmic vesicles as indicated by immunofluorescence microscopy. Consistent with previously published results, these data indicate that CPV enters a host cell via an endocytic route and further suggest that microtubule-dependent delivery of CPV to late endosomes is required for productive infection. Cytoplasmic microinjection of CPV particles was used to circumvent the endocytosis and membrane fusion steps in the entry process. Microinjection experiments showed that CPV particles which were injected directly into the cytoplasm, thus avoiding the endocytic pathway, were unable to initiate progeny virus production. CPV treated at pH 5.0 prior to microinjection was unable to initiate virus production, showing that factors of the endocytic route other than low pH are necessary for the initiation of infection by CPV. PMID:9420290

  1. Canine Models for Copper Homeostasis Disorders

    PubMed Central

    Wu, Xiaoyan; Leegwater, Peter A. J.; Fieten, Hille

    2016-01-01

    Copper is an essential trace nutrient metal involved in a multitude of cellular processes. Hereditary defects in copper metabolism result in disorders with a severe clinical course such as Wilson disease and Menkes disease. In Wilson disease, copper accumulation leads to liver cirrhosis and neurological impairments. A lack in genotype-phenotype correlation in Wilson disease points toward the influence of environmental factors or modifying genes. In a number of Non-Wilsonian forms of copper metabolism, the underlying genetic defects remain elusive. Several pure bred dog populations are affected with copper-associated hepatitis showing similarities to human copper metabolism disorders. Gene-mapping studies in these populations offer the opportunity to discover new genes involved in copper metabolism. Furthermore, due to the relatively large body size and long life-span of dogs they are excellent models for development of new treatment strategies. One example is the recent use of canine organoids for disease modeling and gene therapy of copper storage disease. This review addresses the opportunities offered by canine genetics for discovery of genes involved in copper metabolism disorders. Further, possibilities for the use of dogs in development of new treatment modalities for copper storage disorders, including gene repair in patient-derived hepatic organoids, are highlighted. PMID:26861285

  2. Peptide neurons in the canine small intestine.

    PubMed

    Daniel, E E; Costa, M; Furness, J B; Keast, J R

    1985-07-01

    The distributions of peptide-containing nerve fibers and cell bodies in the canine small intestine were determined with antibodies raised against seven peptides: enkephalin, gastrin-releasing peptide (GRP), neuropeptide Y, neurotensin, somatostatin, substance P, and vasoactive intestinal peptide (VIP). Immunoreactive nerve cell bodies and fibers were found for each peptide except neurotensin. In the muscle layers there were numerous substance P, VIP, and enkephalin fibers, fewer neuropeptide Y fibers, and very few GRP or somatostatin fibers. The mucosa contained many VIP and substance P fibers, moderate numbers of neuropeptide Y, somatostatin, and GRP fibers and rare enkephalin fibers. Nerve cell bodies reactive for each of the six neural peptides were located in both the myenteric and submucous plexuses. The distributions of nerve cell bodies and processes in the canine small intestine show many similarities with other mammals, for example, in the distributions of VIP, substance P, neuropeptide Y, and somatostatin nerves. There are some major differences, such as the presence in dogs of numerous submucosal nerve cell bodies with enkephalinlike immunoreactivity and of GRP-like immunoreactivity in submucous nerve cell bodies and mucosal fibers. PMID:2411766

  3. Masitinib monotherapy in canine epitheliotropic lymphoma.

    PubMed

    Holtermann, N; Kiupel, M; Kessler, M; Teske, E; Betz, D; Hirschberger, J

    2016-08-01

    This study evaluated efficacy and side effects of masitinib in canine epitheliotropic lymphoma. Complete remission occurred in 2 of 10 dogs and lasted for median 85 days. Five dogs went into partial remission for median 60.5 days. Three pretreated dogs did not respond to therapy. Side effects occurred in six dogs and were mostly mild to moderate. Immunohistochemistry was available for eight dogs. KIT receptor was negative in all of them, six of eight lymphomas stained strongly positive for stem cell factor (SCF). platelet-derived growth factor (PDGF)-AA was weakly positive in two and negative in six. PDGF-BB was negative in four tumours, weakly positive in one and strongly positive in three. One was strongly positive for PDGF receptor (PDGFR)-β, seven were negative for that receptor. Five showed strong expression of PDGFR-α, two showed weak expression, one was negative. In conclusion, masitinib is effective in treating canine epitheliotropic lymphoma. But its effects are most likely not generated through the KIT receptor. PMID:26364581

  4. Polar Textures

    NASA Technical Reports Server (NTRS)

    2005-01-01

    [figure removed for brevity, see original site] Context image for PIA03646 Polar Textures

    This VIS image shows part of the south polar region. The ejecta from the relatively young crater covers the rougher textured polar surface.

    Image information: VIS instrument. Latitude 81S, Longitude 54.5E. 17 meter/pixel resolution.

    Note: this THEMIS visual image has not been radiometrically nor geometrically calibrated for this preliminary release. An empirical correction has been performed to remove instrumental effects. A linear shift has been applied in the cross-track and down-track direction to approximate spacecraft and planetary motion. Fully calibrated and geometrically projected images will be released through the Planetary Data System in accordance with Project policies at a later time.

    NASA's Jet Propulsion Laboratory manages the 2001 Mars Odyssey mission for NASA's Office of Space Science, Washington, D.C. The Thermal Emission Imaging System (THEMIS) was developed by Arizona State University, Tempe, in collaboration with Raytheon Santa Barbara Remote Sensing. The THEMIS investigation is led by Dr. Philip Christensen at Arizona State University. Lockheed Martin Astronautics, Denver, is the prime contractor for the Odyssey project, and developed and built the orbiter. Mission operations are conducted jointly from Lockheed Martin and from JPL, a division of the California Institute of Technology in Pasadena.

  5. Canine Pluripotent Stem Cells: Are They Ready for Clinical Applications?

    PubMed

    Betts, Dean H; Tobias, Ian C

    2015-01-01

    The derivation of canine embryonic stem cells and generation of canine-induced pluripotent stem cells are significant achievements that have unlocked the potential for developing novel cell-based disease models, drug discovery platforms, and transplantation therapies in the dog. A progression from concept to cure in this clinically relevant companion animal will not only help our canine patients but also help advance human regenerative medicine. Nevertheless, many issues remain to be resolved before pluripotent cells can be used clinically in a safe and reproducible manner. PMID:26664969

  6. Canine hematopoiesis in a model of combined injury

    SciTech Connect

    MacVittie, T.J.; Monroy, R.L.; Fink, M.; Gruber, D.F.; Patchen, M.L.

    1983-04-29

    The development of a large animal model for CI within the context of a nuclear disaster required that we describe, experimentally, the essential features of the radiobiology of acute effects in the canine. The large-animal model is also appropriate for assessing the immunologic, pharmacologic, and surgical modes of intervention of following CI. The canine model of CI at the AFRRI has stressed three developmental aspects: (a) establishing the radiobiology of the canine hemopoietic system, (b) choosing a relevant model for peritoneal sepsis, and (c) identifying several choices for physical trauma. This paper stresses the relevance of the first aspect, the radiation-induced suppression and recovery of the hemopoietic system.

  7. Kinetics of canine dental calculus crystallization: an in vitro study on the influence of inorganic components of canine saliva.

    PubMed

    Borah, Ballav M; Halter, Timothy J; Xie, Baoquan; Henneman, Zachary J; Siudzinski, Thomas R; Harris, Stephen; Elliott, Matthew; Nancollas, George H

    2014-07-01

    This work identifies carbonated hydroxyapatite (CAP) as the primary component of canine dental calculus, and corrects the long held belief that canine dental calculus is primarily CaCO3 (calcite). CAP is known to be the principal crystalline component of human dental calculus, suggesting that there are previously unknown similarities in the calcification that occurs in these two unique oral environments. In vitro kinetic experiments mimicking the inorganic components of canine saliva have examined the mechanisms of dental calculus formation. The solutions were prepared so as to mimic the inorganic components of canine saliva; phosphate, carbonate, and magnesium ion concentrations were varied individually to investigate the roll of these ions in controlling the nature of the phases that is nucleated. To date, the inorganic components of the canine oral systems have not been investigated at concentrations that mimic those in vivo. The mineral composition of the synthetic calculi grown under these conditions closely resembled samples excised from canines. This finding adds new information about calculus formation in humans and canines, and their sensitivity to chemicals used to treat these conditions. PMID:24776659

  8. Collective Cell Motion in an Epithelial Sheet Can Be Quantitatively Described by a Stochastic Interacting Particle Model

    PubMed Central

    Cochet, Olivier; Grasland-Mongrain, Erwan; Silberzan, Pascal; Hakim, Vincent

    2013-01-01

    Modelling the displacement of thousands of cells that move in a collective way is required for the simulation and the theoretical analysis of various biological processes. Here, we tackle this question in the controlled setting where the motion of Madin-Darby Canine Kidney (MDCK) cells in a confluent epithelium is triggered by the unmasking of free surface. We develop a simple model in which cells are described as point particles with a dynamic based on the two premises that, first, cells move in a stochastic manner and, second, tend to adapt their motion to that of their neighbors. Detailed comparison to experimental data show that the model provides a quantitatively accurate description of cell motion in the epithelium bulk at early times. In addition, inclusion of model “leader” cells with modified characteristics, accounts for the digitated shape of the interface which develops over the subsequent hours, providing that leader cells invade free surface more easily than other cells and coordinate their motion with their followers. The previously-described progression of the epithelium border is reproduced by the model and quantitatively explained. PMID:23505356

  9. UDP-N-acetylglucosamine Transporter (SLC35A3) Regulates Biosynthesis of Highly Branched N-Glycans and Keratan Sulfate*

    PubMed Central

    Maszczak-Seneczko, Dorota; Sosicka, Paulina; Olczak, Teresa; Jakimowicz, Piotr; Majkowski, Michał; Olczak, Mariusz

    2013-01-01

    SLC35A3 is considered the main UDP-N-acetylglucosamine transporter (NGT) in mammals. Detailed analysis of NGT is restricted because mammalian mutant cells defective in this activity have not been isolated. Therefore, using the siRNA approach, we developed and characterized several NGT-deficient mammalian cell lines. CHO, CHO-Lec8, and HeLa cells deficient in NGT activity displayed a decrease in the amount of highly branched tri- and tetraantennary N-glycans, whereas monoantennary and diantennary ones remained unchanged or even were accumulated. Silencing the expression of NGT in Madin-Darby canine kidney II cells resulted in a dramatic decrease in the keratan sulfate content, whereas no changes in biosynthesis of heparan sulfate were observed. We also demonstrated for the first time close proximity between NGT and mannosyl (α-1,6-)-glycoprotein β-1,6-N-acetylglucosaminyltransferase (Mgat5) in the Golgi membrane. We conclude that NGT may be important for the biosynthesis of highly branched, multiantennary complex N-glycans and keratan sulfate. We hypothesize that NGT may specifically supply β-1,3-N-acetylglucosaminyl-transferase 7 (β3GnT7), Mgat5, and possibly mannosyl (α-1,3-)-glycoprotein β-1,4-N-acetylglucosaminyltransferase (Mgat4) with UDP-GlcNAc. PMID:23766508

  10. Cyclooxygenase-2-dependent phosphorylation of the pro-apoptotic protein Bad inhibits tonicity-induced apoptosis in renal medullary cells.

    PubMed

    Küper, Christoph; Bartels, Helmut; Beck, Franz-X; Neuhofer, Wolfgang

    2011-11-01

    During antidiuresis, cell survival in the renal medulla requires cyclooxygenase-2 (COX-2) activity. We have recently found that prostaglandin E2 (PGE2) promotes cell survival by phosphorylation and, hence, inactivation of the pro-apoptotic protein Bad during hypertonic stress in Madin-Darby canine kidney (MDCK) cells in vitro. Here we determine the role of COX-2-derived PGE(2) on phosphorylation of Bad and medullary apoptosis in vivo using COX-2-deficient mice. Both wild-type and COX-2-knockout mice constitutively expressed Bad in tubular epithelial cells of the renal medulla. Dehydration caused a robust increase in papillary COX-2 expression, PGE2 excretion, and Bad phosphorylation in wild-type, but not in the knockout mice. The abundance of cleaved caspase-3, a marker of apoptosis, was significantly higher in papillary homogenates, especially in tubular epithelial cells of the knockout mice. Knockdown of Bad in MDCK cells decreased tonicity-induced caspase-3 activation. Furthermore, the addition of PGE2 to cells with knockdown of Bad had no effect on caspase-3 activation; however, PGE2 caused phosphorylation of Bad and substantially improved cell survival in mock-transfected cells. Thus, tonicity-induced COX-2 expression and PGE2 synthesis in the renal medulla entails phosphorylation and inactivation of the pro-apoptotic protein Bad, thereby counteracting apoptosis in renal medullary epithelial cells. PMID:21716255

  11. Lipids that determine detergent resistance of MDCK cell membrane fractions.

    PubMed

    Manni, Marco M; Cano, Ainara; Alonso, Cristina; Goñi, Félix M

    2015-10-01

    A comparative lipidomic study has been performed of whole Madin-Darby canine kidney epithelial cells and of the detergent-resistant membrane fraction (DRM) obtained after treating the cells with the non-ionic detergent Triton X-100. The DRM were isolated following a standard procedure that is extensively used in cell biology studies. Significant differences were found in the lipid composition of the whole cells and of DRM. The latter were enriched in all the analyzed sphingolipid classes: sphingomyelins, ceramides and hexosylceramides. Diacylglycerols were also preferentially found in DRM. The detergent-resistant fraction was also enriched in saturated over unsaturated fatty acyl chains, and in sn-1 acyl chains containing 16 carbon atoms, over the longer and shorter ones. The glycerophospholipid species phosphatidylethanolamines and phosphatidylinositols, that were mainly unsaturated, did not show a preference for DRM. Phosphatidylcholines were an intermediate case: the saturated, but not the unsaturated species were found preferentially in DRM. The question remains on whether these DRM, recovered from detergent-membrane mixtures by floatation over a sucrose gradient, really correspond to membrane domains existing in the cell membrane prior to detergent treatment. PMID:26320877

  12. A toxin-based probe reveals cytoplasmic exposure of Golgi sphingomyelin.

    PubMed

    Bakrac, Biserka; Kladnik, Ales; Macek, Peter; McHaffie, Gavin; Werner, Andreas; Lakey, Jeremy H; Anderluh, Gregor

    2010-07-16

    Although sphingomyelin is an important cellular lipid, its subcellular distribution is not precisely known. Here we use a sea anemone cytolysin, equinatoxin II (EqtII), which specifically binds sphingomyelin, as a new marker to detect cellular sphingomyelin. A purified fusion protein composed of EqtII and green fluorescent protein (EqtII-GFP) binds to the SM rich apical membrane of Madin-Darby canine kidney (MDCK) II cells when added exogenously, but not to the SM-free basolateral membrane. When expressed intracellularly within MDCK II cells, EqtII-GFP colocalizes with markers for Golgi apparatus and not with those for nucleus, mitochondria, endoplasmic reticulum or plasma membrane. Colocalization with the Golgi apparatus was confirmed by also using NIH 3T3 fibroblasts. Moreover, EqtII-GFP was enriched in cis-Golgi compartments isolated by gradient ultracentrifugation. The data reveal that EqtII-GFP is a sensitive probe for membrane sphingomyelin, which provides new information on cytosolic exposure, essential to understand its diverse physiological roles. PMID:20463009

  13. Three-dimensional morphogenesis of MDCK cells induced by cellular contractile forces on a viscous substrate.

    PubMed

    Imai, Misako; Furusawa, Kazuya; Mizutani, Takeomi; Kawabata, Kazushige; Haga, Hisashi

    2015-01-01

    Substrate physical properties are essential for many physiological events such as embryonic development and 3D tissue formation. Physical properties of the extracellular matrix such as viscoelasticity and geometrical constraints are understood as factors that affect cell behaviour. In this study, we focused on the relationship between epithelial cell 3D morphogenesis and the substrate viscosity. We observed that Madin-Darby Canine Kidney (MDCK) cells formed 3D structures on a viscous substrate (Matrigel). The structures appear as a tulip hat. We then changed the substrate viscosity by genipin (GP) treatment. GP is a cross-linker of amino groups. Cells cultured on GP-treated-matrigel changed their 3D morphology in a substrate viscosity-dependent manner. Furthermore, to elucidate the spatial distribution of the cellular contractile force, localization of mono-phosphorylated and di-phosphorylated myosin regulatory light chain (P-MRLCs) was visualized by immunofluorescence. P-MRLCs localized along the periphery of epithelial sheets. Treatment with Y-27632, a Rho-kinase inhibitor, blocked the P-MRLCs localization at the edge of epithelial sheets and halted 3D morphogenesis. Our results indicate that the substrate viscosity, the substrate deformation, and the cellular contractile forces induced by P-MRLCs play crucial roles in 3D morphogenesis. PMID:26374384

  14. Synthesis-Dependent Surface Defects and Morphology of Hematite Nanoparticles and Their Effect on Cytotoxicity in Vitro.

    PubMed

    Cardillo, Dean; Tehei, Moeava; Hossain, Md Shahriar; Islam, Md Monirul; Bogusz, Kathrin; Shi, Dongqi; Mitchell, David; Lerch, Michael; Rosenfeld, Anatoly; Corde, Stéphanie; Konstantinov, Konstantin

    2016-03-01

    In this study, we investigate the toxicity of hematite (α-Fe2O3) nanoparticles on the Madin-Darby Canine Kidney (MDCK) cell line. The oxide particles have been synthesized through two different methods and annealing conditions. These two methods, spray precipitation and precipitation, resulted in particles with rod-like and spherical morphology and feature different particle sizes, surface features, and magnetic properties. Through flow cytometry it was found that particle morphology heavily influences the degree to which the nanomaterials are internalized into the cells. It was also found that the ability of the nanoparticles to generate free radicals species is hindered by the formation of tetrahedrally coordinated maghemite-like (γ-Fe2O3) spinel defects on the surfaces of the particles. The combination of these two factors resulted in variable cytotoxic effects of the hematite nanoparticles synthesized with different conditions. This article highlights the importance on the fabrication method, materials properties, and surface characteristics on the cytotoxicity of hematite nanomaterials. PMID:26881459

  15. Development of high-yield influenza A virus vaccine viruses.

    PubMed

    Ping, Jihui; Lopes, Tiago J S; Nidom, Chairul A; Ghedin, Elodie; Macken, Catherine A; Fitch, Adam; Imai, Masaki; Maher, Eileen A; Neumann, Gabriele; Kawaoka, Yoshihiro

    2015-01-01

    Vaccination is one of the most cost-effective ways to prevent infection. Influenza vaccines propagated in cultured cells are approved for use in humans, but their yields are often suboptimal. Here, we screened A/Puerto Rico/8/34 (PR8) virus mutant libraries to develop vaccine backbones (defined here as the six viral RNA segments not encoding haemagglutinin and neuraminidase) that support high yield in cell culture. We also tested mutations in the coding and regulatory regions of the virus, and chimeric haemagglutinin and neuraminidase genes. A combination of high-yield mutations from these screens led to a PR8 backbone that improved the titres of H1N1, H3N2, H5N1 and H7N9 vaccine viruses in African green monkey kidney and Madin-Darby canine kidney cells. This PR8 backbone also improves titres in embryonated chicken eggs, a common propagation system for influenza viruses. This PR8 vaccine backbone thus represents an advance in seasonal and pandemic influenza vaccine development. PMID:26334134

  16. Synthetic generation of influenza vaccine viruses for rapid response to pandemics.

    PubMed

    Dormitzer, Philip R; Suphaphiphat, Pirada; Gibson, Daniel G; Wentworth, David E; Stockwell, Timothy B; Algire, Mikkel A; Alperovich, Nina; Barro, Mario; Brown, David M; Craig, Stewart; Dattilo, Brian M; Denisova, Evgeniya A; De Souza, Ivna; Eickmann, Markus; Dugan, Vivien G; Ferrari, Annette; Gomila, Raul C; Han, Liqun; Judge, Casey; Mane, Sarthak; Matrosovich, Mikhail; Merryman, Chuck; Palladino, Giuseppe; Palmer, Gene A; Spencer, Terika; Strecker, Thomas; Trusheim, Heidi; Uhlendorff, Jennifer; Wen, Yingxia; Yee, Anthony C; Zaveri, Jayshree; Zhou, Bin; Becker, Stephan; Donabedian, Armen; Mason, Peter W; Glass, John I; Rappuoli, Rino; Venter, J Craig

    2013-05-15

    During the 2009 H1N1 influenza pandemic, vaccines for the virus became available in large quantities only after human infections peaked. To accelerate vaccine availability for future pandemics, we developed a synthetic approach that very rapidly generated vaccine viruses from sequence data. Beginning with hemagglutinin (HA) and neuraminidase (NA) gene sequences, we combined an enzymatic, cell-free gene assembly technique with enzymatic error correction to allow rapid, accurate gene synthesis. We then used these synthetic HA and NA genes to transfect Madin-Darby canine kidney (MDCK) cells that were qualified for vaccine manufacture with viral RNA expression constructs encoding HA and NA and plasmid DNAs encoding viral backbone genes. Viruses for use in vaccines were rescued from these MDCK cells. We performed this rescue with improved vaccine virus backbones, increasing the yield of the essential vaccine antigen, HA. Generation of synthetic vaccine seeds, together with more efficient vaccine release assays, would accelerate responses to influenza pandemics through a system of instantaneous electronic data exchange followed by real-time, geographically dispersed vaccine production. PMID:23677594

  17. Inhibition of influenza A virus infection in vitro by saliphenylhalamide-loaded porous silicon nanoparticles.

    PubMed

    Bimbo, Luis M; Denisova, Oxana V; Mäkilä, Ermei; Kaasalainen, Martti; De Brabander, Jef K; Hirvonen, Jouni; Salonen, Jarno; Kakkola, Laura; Kainov, Denis; Santos, Hélder A

    2013-08-27

    Influenza A viruses (IAVs) cause recurrent epidemics in humans, with serious threat of lethal worldwide pandemics. The occurrence of antiviral-resistant virus strains and the emergence of highly pathogenic influenza viruses have triggered an urgent need to develop new anti-IAV treatments. One compound found to inhibit IAV, and other virus infections, is saliphenylhalamide (SaliPhe). SaliPhe targets host vacuolar-ATPase and inhibits acidification of endosomes, a process needed for productive virus infection. The major obstacle for the further development of SaliPhe as antiviral drug has been its poor solubility. Here, we investigated the possibility to increase SaliPhe solubility by loading the compound in thermally hydrocarbonized porous silicon (THCPSi) nanoparticles. SaliPhe-loaded nanoparticles were further investigated for the ability to inhibit influenza A infection in human retinal pigment epithelium and Madin-Darby canine kidney cells, and we show that upon release from THCPSi, SaliPhe inhibited IAV infection in vitro and reduced the amount of progeny virus in IAV-infected cells. Overall, the PSi-based nanosystem exhibited increased dissolution of the investigated anti-IAV drug SaliPhe and displayed excellent in vitro stability, low cytotoxicity, and remarkable reduction of viral load in the absence of organic solvents. This proof-of-principle study indicates that PSi nanoparticles could be used for efficient delivery of antivirals to infected cells. PMID:23889734

  18. Lindane-induced generation of reactive oxygen species and depletion of glutathione do not result in necrosis in renal distal tubule cells.

    PubMed

    Piskac-Collier, Amanda L; Smith, Mary Ann

    2009-01-01

    Lindane is a chlorinated hydrocarbon pesticide, currently used in prescription shampoos and lotions to treat scabies and lice infestations. Lindane is known to be nephrotoxic; however, the mechanism of action is not well understood. In other organ systems, lindane produces cellular damage by generation of free radicals and oxidative stress. Morphological changes were observed in lindane-treated Madin-Darby canine kidney (MDCK) cells indicative of apoptosis. Lindane treatment induced time-dependent reactive oxygen species (ROS) generation. Onset of ROS generation correlated with an initial increase in total glutathione (GSH) levels above control values, with a subsequent decline in a time-dependent manner. This decline may be attributed to quenching of free radicals by GSH, thereby decreasing the cellular stores of this antioxidant. Necrotic injury was assessed by measuring lactate dehydrogenase (LDH) leakage from the cell after lindane exposure. No significant LDH leakage was noted for all concentrations tested over time. Generation of ROS and alterations in cellular protective mechanisms did not result in necrotic injury in MDCK cells, which corresponds with our morphological findings of lindane-induced apoptotic changes as opposed to necrosis in MDCK cells. Thus, lindane exposure results in oxidative damage and alterations in antioxidant response in renal distal tubule cells, followed by cell death not attributed to necrotic injury. PMID:20077184

  19. Effect of salt intake on bioavailability of mizoribine in healthy Japanese males.

    PubMed

    Ishida, Kazuya; Fukao, Miki; Watanabe, Hitomi; Taguchi, Masato; Miyawaki, Toshio; Matsukura, Hiroyoshi; Uemura, Osamu; Zhang, Zufei; Unadkat, Jashvant D; Hashimoto, Yukiya

    2013-01-01

    Bioavailability of mizoribine in subjects with the concentrative nucleoside transporter 1 (CNT1, SLC28A1) 565-A/A allele is significantly lower than that in subjects with the SLC28A1 565-G/G allele. The aims of the present study were to investigate the cellular uptake of mizoribine in CNT1- and CNT2-expressing Madin-Darby canine kidney type II (MDCKII) cells, and to evaluate the effect of salt intake on bioavailability of mizoribine in healthy Japanese volunteers with SLC28A1 565-A/A and -G/A alleles. Eight healthy males participated in the present study, and took 150 mg mizoribine concomitantly with/without 300 mg salt. Bioavailability of mizoribine was estimated by total cumulative urinary excretion of the drug. Mizoribine was taken up Na(+)-dependently into not only CNT1-expressing but also CNT2-expressing MDCKII cells, indicating that mizoribine is a substrate for both CNT1 and CNT2. Mean bioavailability of mizoribine taken with salt (83.8%) was significantly higher than that taken without salt (73.0%). These findings suggest that the salt intake is expected to improve the bioavailability of mizoribine in patients with insufficient intestinal absorption. PMID:22813716

  20. Endocellular polyamine availability modulates epithelial-to-mesenchymal transition and unfolded protein response in MDCK cells.

    PubMed

    Prunotto, Marco; Compagnone, Alessandra; Bruschi, Maurizio; Candiano, Giovanni; Colombatto, Sebastiano; Bandino, Andrea; Petretto, Andrea; Moll, Solange; Bochaton-Piallat, Marie Luce; Gabbiani, Giulio; Dimuccio, Veronica; Parola, Maurizio; Citti, Lorenzo; Ghiggeri, Gianmarco

    2010-06-01

    Epithelial-to-mesenchymal transition (EMT) is involved in embryonic development as well as in several pathological conditions. Literature indicates that polyamine availability may affect transcription of c-myc, matrix metalloproteinase (MMP)1, MMP2, TGFbeta(1), and collagen type I mRNA. The aim of this study was to elucidate polyamines role in EMT in vitro. Madin-Darby canine kidney (MDCK) cells were subjected to experimental manipulation of intracellular levels of polyamines. Acquisition of mesenchymal phenotype was evaluated by means of immunofluorescence, western blots, and zymograms. MDCK cells were then subjected to 2D gel proteomic study and incorporation of a biotinilated polyamine (BPA). Polyamine endocellular availability modulated EMT process. Polyamine-depleted cells treated with TGFbeta(1) showed enhanced EMT with a marked decrease of E-cadherin expression at plasma membrane level and an increased expression of mesenchymal markers such as fibronectin and alpha-smooth muscle actin. Polyamine-depleted cells showed a twofold increased expression of the rough endoplasmic reticulum (ER)-stress proteins GRP78, GRP94, and HSP90 alpha/beta in 2D gels. The latter data were confirmed by western blot analysis. Administration of BPA showed that polyamines are covalently linked, within the cell, to ER-stress proteins. Intracellular polyamine availability affects EMT in MDCK cells possibly through the modulation of ER-stress protein homeostasis. PMID:20212449

  1. Feasibility Study of the Permeability and Uptake of Mesoporous Silica Nanoparticles across the Blood-Brain Barrier

    PubMed Central

    Baghirov, Habib; Karaman, Didem; Viitala, Tapani; Duchanoy, Alain; Lou, Yan-Ru; Mamaeva, Veronika; Pryazhnikov, Evgeny; Khiroug, Leonard; de Lange Davies, Catharina; Sahlgren, Cecilia; Rosenholm, Jessica M.

    2016-01-01

    Drug delivery into the brain is impeded by the blood-brain-barrier (BBB) that filters out the vast majority of drugs after systemic administration. In this work, we assessed the transport, uptake and cytotoxicity of promising drug nanocarriers, mesoporous silica nanoparticles (MSNs), in in vitro models of the BBB. RBE4 rat brain endothelial cells and Madin-Darby canine kidney epithelial cells, strain II, were used as BBB models. We studied spherical and rod-shaped MSNs with the following modifications: bare MSNs and MSNs coated with a poly(ethylene glycol)-poly(ethylene imine) (PEG-PEI) block copolymer. In transport studies, MSNs showed low permeability, whereas the results of the cellular uptake studies suggest robust uptake of PEG-PEI-coated MSNs. None of the MSNs showed significant toxic effects in the cell viability studies. While the shape effect was detectable but small, especially in the real-time surface plasmon resonance measurements, coating with PEG-PEI copolymers clearly facilitated the uptake of MSNs. Finally, we evaluated the in vivo detectability of one of the best candidates, i.e. the copolymer-coated rod-shaped MSNs, by two-photon in vivo imaging in the brain vasculature. The particles were clearly detectable after intravenous injection and caused no damage to the BBB. Thus, when properly designed, the uptake of MSNs could potentially be utilized for the delivery of drugs into the brain via transcellular transport. PMID:27547955

  2. Functional rescue of a kidney anion exchanger 1 trafficking mutant in renal epithelial cells.

    PubMed

    Chu, Carmen Y S; King, Jennifer C; Berrini, Mattia; Alexander, R Todd; Cordat, Emmanuelle

    2013-01-01

    Mutations in the SLC4A1 gene encoding the anion exchanger 1 (AE1) can cause distal renal tubular acidosis (dRTA), a disease often due to mis-trafficking of the mutant protein. In this study, we investigated whether trafficking of a Golgi-retained dRTA mutant, G701D kAE1, or two dRTA mutants retained in the endoplasmic reticulum, C479W and R589H kAE1, could be functionally rescued to the plasma membrane of Madin-Darby Canine Kidney (MDCK) cells. Treatments with DMSO, glycerol, the corrector VX-809, or low temperature incubations restored the basolateral trafficking of G701D kAE1 mutant. These treatments had no significant rescuing effect on trafficking of the mis-folded C479W or R589H kAE1 mutants. DMSO was the only treatment that partially restored G701D kAE1 function in the plasma membrane of MDCK cells. Our experiments show that trafficking of intracellularly retained dRTA kAE1 mutants can be partially restored, and that one chemical treatment rescued both trafficking and function of a dRTA mutant. These studies provide an opportunity to develop alternative therapeutic solutions for dRTA patients. PMID:23460825

  3. ATP-binding cassette transporters as pitfalls in selection of transgenic cells.

    PubMed

    Theile, Dirk; Staffen, Bianca; Weiss, Johanna

    2010-04-15

    Puromycin, hygromycin, and geneticin (G418) are antibiotics frequently used to select genetically engineered eukaryotic cells after transfection or transduction. Because intrinsic or acquired high expression of ATP-binding cassette (ABC) transporters, such as P-glycoprotein (Pgp/ABCB1) and multidrug resistance-associated proteins (MRP/ABCC1), can hamper efficient selection, it is important to know whether these antibiotics are substrates and/or inducers of efflux transporters. Therefore, we investigated the influence of these antibiotics on drug transporter expression by quantitative real-time polymerase chain reaction in the induction model cell line LS180. Moreover, we assessed whether ABC transporters influence the growth inhibitory effects of these antibiotics by proliferation assays using Madin-Darby canine kidney II (MDCKII) cells overexpressing the particular transporter. The results obtained indicate that puromycin and G418 are substrates of several ABC transporters, mainly Pgp/ABCB1. In contrast, hygromycin seems to be no good substrate for any of the ABC transporters investigated. Puromycin induced ABCC1/MRP1, whereas G418 suppressed ABCB1/Pgp, at the messenger RNA (mRNA) level. In contrast, hygromycin had no effect on ABC transporter mRNA expressions. In conclusion, this study emphasizes the significance of ABC transporters for the efficacy of selection processes. Consciousness of the results is supposed to guide the molecular biologist to the right choice of adequate experimental conditions for successful selection of genetically engineered eukaryotic cells. PMID:20018165

  4. Shear-induced Volume Decrease in MDCK Cells

    PubMed Central

    Heo, Jinseok; Sachs, Frederick; Wang, Jianbin; Hua, Susan Z.

    2013-01-01

    Using a microfluidic cell volume sensor we measured the change in the cell volume of Madin-Darby Canine Kidney (MDCK) cells induced by shear stress. An increase in shear stress from 0.2 to 2.0 dyn/cm2 resulted in a volume decrease to a steady state volume ~ 20 – 30 % smaller than the initial resting cell volume. Independent experiments based on fluorescence quenching confirmed the volume reduction. This shear-induced cell shrinkage was irreversible on the time scale of the experiment (~ 30 min). Treatment of 0.1 μM Hg2+ significantly inhibited the volume decrease, suggesting that the shear-induced cell shrinkage is associated with water efflux through aquaporins. The volume decrease cannot be inhibited by 75 mM TEA, 100 μM DIDS, or 100 μM Gd3+ suggesting that volume reduction is not directly mediated by K+ and Cl− channels that typically function during regulatory volume decrease (RVD), nor is it through cationic stretch-activated ion channels (SACs). The process also appears to be Ca2+ independent because it was insensitive to intracellular Ca2+ level. Since cell volume is determined by the intracellular water content, we postulate that the shear induced reductions in cell volume may arise from increased intracellular hydrostatic pressure as the cell is deformed under flow, which promotes the efflux of water. The increase in internal pressure in a deformable object under the flow is supported by the finite element mechanical model. PMID:22759987

  5. Method for recovering ATP content and mitochondrial function after chemical anoxia in renal cell cultures.

    PubMed

    Doctor, R B; Bacallao, R; Mandel, L J

    1994-06-01

    Cultured renal cells provide a highly reproducible and malleable model to study cellular responses to metabolic perturbations. Nevertheless, there is currently no good method to achieve metabolic inhibition and complete recovery in cultured cells. This study describes a specific method for reversibly inhibiting both glycolytic and oxidative metabolism. Glycolysis was inhibited by removing all glycolytic substrates, and mitochondrial respiration was inhibited with rotenone, a site I inhibitor of the electron transport chain. Within 30 min, ATP values were decreased by 98%. Glycolysis was restored through the reintroduction of glucose. Oxidative metabolism was restored by the addition of heptanoate, a short odd-chain fatty acid, which supplies reducing equivalents to site II of the electron transport chain. Employing Madin-Darby canine kidney and LLC-PK1 cell lines, this protocol caused the immediate and complete recovery of mitochondrial respiration and, by 60 min, the complete recovery of cellular ATP levels. Application of this protocol should allow the investigation of the cellular effects and alterations that occur within cells recovering from sublethal energy depletion. PMID:8023909

  6. Dynamic-SERS Optophysiology: A Nanosensor for Monitoring Cell Secretion Events.

    PubMed

    Lussier, Félix; Brulé, Thibault; Vishwakarma, Medhavi; Das, Tamal; Spatz, Joachim P; Masson, Jean-François

    2016-06-01

    We monitored metabolite secretion near living cells using a plasmonic nanosensor. The nanosensor created from borosilicate nanopipettes analogous to the patch clamp was decorated with Au nanoparticles and served as a surface-enhanced Raman scattering (SERS) substrate with addressable location. With this nanosensor, we acquired SERS locally near Madin-Darby canine kidney (MDCKII) epithelial cells, and we detected multiple metabolites, such as pyruvate, lactate, ATP, and urea simultaneously. These plasmonic nanosensors were capable of monitoring metabolites in the extracellular medium with enough sensitivity to detect an increase in metabolite concentration following the lyses of MDCKII cells with a nonionic surfactant. The plasmonic nanosensors also allowed a relative quantification of a chemical gradient for a metabolite near cells, as demonstrated with a decrease in relative lactate to pyruvate concentration further away from the MDCKII cells. This SERS optophysiology technique for the sensitive and nondestructive monitoring of extracellular metabolites near living cells is broadly applicable to different cellular and tissue models and should therefore provide a powerful tool for cellular studies. PMID:27172291

  7. Functional Rescue of a Kidney Anion Exchanger 1 Trafficking Mutant in Renal Epithelial Cells

    PubMed Central

    Chu, Carmen Y. S.; King, Jennifer C.; Berrini, Mattia; Alexander, R. Todd; Cordat, Emmanuelle

    2013-01-01

    Mutations in the SLC4A1 gene encoding the anion exchanger 1 (AE1) can cause distal renal tubular acidosis (dRTA), a disease often due to mis-trafficking of the mutant protein. In this study, we investigated whether trafficking of a Golgi-retained dRTA mutant, G701D kAE1, or two dRTA mutants retained in the endoplasmic reticulum, C479W and R589H kAE1, could be functionally rescued to the plasma membrane of Madin-Darby Canine Kidney (MDCK) cells. Treatments with DMSO, glycerol, the corrector VX-809, or low temperature incubations restored the basolateral trafficking of G701D kAE1 mutant. These treatments had no significant rescuing effect on trafficking of the mis-folded C479W or R589H kAE1 mutants. DMSO was the only treatment that partially restored G701D kAE1 function in the plasma membrane of MDCK cells. Our experiments show that trafficking of intracellularly retained dRTA kAE1 mutants can be partially restored, and that one chemical treatment rescued both trafficking and function of a dRTA mutant. These studies provide an opportunity to develop alternative therapeutic solutions for dRTA patients. PMID:23460825

  8. Direct sequencing of the HA gene of influenza (H3N2) virus in original clinical samples reveals sequence identity with mammalian cell-grown virus.

    PubMed Central

    Katz, J M; Wang, M; Webster, R G

    1990-01-01

    When influenza (H3N2) viruses from infected individuals are grown in embryonated chicken eggs, viruses are isolated which differ antigenically and structurally from viruses grown in mammalian Madin-Darby canine kidney (MDCK) cell culture [G.C. Schild, J.S. Oxford, J.C. de Jong, and R.G. Webster, Nature (London) 303:706-709, 1983]. To determine which of these viruses is most representative of virus replicating in the infected individual, a region of the HA gene of virus present in original clinical samples was amplified by using the polymerase chain reaction and sequenced directly. Comparison of 170 amino acid residues of HA1 flanking and containing the receptor-binding site and antigenic sites indicated that over this region, the HA of virus replicating in the infected individual was identical to that of virus after growth in MDCK cells and was distinct from the HA of viruses grown in eggs. Therefore, cultivation of human influenza H3N2 virus in mammalian MDCK cells results in a virus similar to the predominant population of virus found in the infected individual. PMID:2319652

  9. Mycophenolic acid glucuronide is transported by multidrug resistance-associated protein 2 and this transport is not inhibited by cyclosporine, tacrolimus or sirolimus.

    PubMed

    Patel, Chirag G; Ogasawara, Ken; Akhlaghi, Fatemeh

    2013-03-01

    1. The purpose of this study was to investigate the contribution of MRP2 to the efflux of mycophenolic acid (MPA), and its phenyl glucuronide (MPAG) and acyl glucuronide (AcMPAG) metabolites, using Madin-Darby canine kidney II cells stably transfected with human MRP2 gene (MDCKII/MRP2 cells). 2. Compared to parental MDCKII cells, MPAG was significantly translocated from basolateral (BL) to apical (AP) side in MDCKII/MRP2 cells, indicating MPAG is a substrate for MRP2. AcMPAG is highly translocated from BL to AP side in both cells, suggesting that AcMPAG is actively secreted possibly through an efflux transporter other than MRP2. Appreciable translocation of MPA was not observed in MDCKII/MRP2 cells. 3. Furthermore, using MRP2-expressing Sf9 membrane vesicles, the Michaelis-Menten constant (Km) value for MRP2-mediated MPAG transport was calculated at 224.2 ± 42.7 µM. In the vesicle system, cyclosporine, tacrolimus and sirolimus did not inhibit the uptake of MPAG via MRP2. 4. These findings indicate that only MPAG not MPA and AcMPAG is a substrate for MRP2 and that the interaction between MPAG and concomitantly administered immunosuppressive agents does not occur at MRP2 level. PMID:22934787

  10. ZO-1 Knockout by TALEN-Mediated Gene Targeting in MDCK Cells: Involvement of ZO-1 in the Regulation of Cytoskeleton and Cell Shape

    PubMed Central

    Tokuda, Shinsaku; Higashi, Tomohito; Furuse, Mikio

    2014-01-01

    ZO-1, ZO-2 and ZO-3 are tight junction-associated scaffold proteins that bind to transmembrane proteins of tight junctions and the underlying cytoskeleton. ZO-1 is involved in the regulation of cytoskeletal organization, but its detailed molecular mechanism is less well understood. Gene knockout is an ideal method to investigate the functions of proteins that might have redundant functions such as ZO proteins, when compared with methods such as RNA interference-mediated suppression of gene expression. In this study we applied transcription activator-like effector nucleases (TALENs), a recently developed genome editing method for gene knockout, and established ZO-1 knockout clones in Madin-Darby canine kidney (MDCK) cells. ZO-1 knockout induced striking changes in myosin organization at cell–cell contacts and disrupted the localization of tight junction proteins; these findings were previously unseen in studies of ZO-1 knockdown by RNA interference. Rescue experiments revealed that trace ZO-1 expression reversed these changes while excessive ZO-1 expression induced an intensive zigzag shape of cell–cell junctions. These results suggest a role for ZO-1 in the regulation of cytoskeleton and shape of cell–cell junctions in MDCK cells and indicate the advantage of knockout analysis in cultured cells. PMID:25157572

  11. Effects of Hydrostatic Pressure on Carcinogenic Properties of Epithelia

    PubMed Central

    Tokuda, Shinsaku; Kim, Young Hak; Matsumoto, Hisako; Muro, Shigeo; Hirai, Toyohiro; Mishima, Michiaki; Furuse, Mikio

    2015-01-01

    The relationship between chronic inflammation and cancer is well known. The inflammation increases the permeability of blood vessels and consequently elevates pressure in the interstitial tissues. However, there have been only a few reports on the effects of hydrostatic pressure on cultured cells, and the relationship between elevated hydrostatic pressure and cell properties related to malignant tumors is less well understood. Therefore, we investigated the effects of hydrostatic pressure on the cultured epithelial cells seeded on permeable filters. Surprisingly, hydrostatic pressure from basal to apical side induced epithelial stratification in Madin-Darby canine kidney (MDCK) I and Caco-2 cells, and cavities with microvilli and tight junctions around their surfaces were formed within the multi-layered epithelia. The hydrostatic pressure gradient also promoted cell proliferation, suppressed cell apoptosis, and increased transepithelial ion permeability. The inhibition of protein kinase A (PKA) promoted epithelial stratification by the hydrostatic pressure whereas the activation of PKA led to suppressed epithelial stratification. These results indicate the role of the hydrostatic pressure gradient in the regulation of various epithelial cell functions. The findings in this study may provide clues for the development of a novel strategy for the treatment of the carcinoma. PMID:26716691

  12. Blockade of swelling-induced chloride channels by phenol derivatives.

    PubMed Central

    Gschwentner, M.; Jungwirth, A.; Hofer, S.; Wöll, E.; Ritter, M.; Susanna, A.; Schmarda, A.; Reibnegger, G.; Pinggera, G. M.; Leitinger, M.; Frick, J.; Deetjen, P.; Paulmichl, M.

    1996-01-01

    1. In NIH3T3 fibroblasts, the chloride channel involved in regulatory volume decrease (RVD) was identified as ICln, a protein isolated from a cDNA library derived from Madin Darby canine Kidney (MDCK) cells. ICln expressed in Xenopus laevis oocytes gives rise to an outwardly rectifying chloride current, sensitive to the extracellular addition of nucleotides and the known chloride channel blockers, DIDS (4,4'-diisothiocyanatostilbene-2,2'-disulphonic acid) and NPPB (5-nitro-2-(3-phenylpropylamino)-benzoic acid). We set out to study whether substances structurally similar to NPPB are able to interfere with RVD. 2. RVD in NIH3T3 fibroblasts and MDCK cells is temperature-dependent. 3. RVD, the swelling-dependent chloride current and the depolarization seen after reducing extracellular osmolarity can be blocked by gossypol and NDGA (nordihydroguaiaretic acid), both structurally related to NPPB. 4. The cyclic AMP-dependent chloride current elicited in CaCo cells is less sensitive to the two substances tested while the calcium-activated chloride current in fibroblasts is insensitive. 5. The binding site for the two phenol derivatives onto ICln seems to be distinct but closely related to the nucleotide binding site identified as G x G x G, a glycine repeat located at the predicted outer mouth of the ICln channel protein. PMID:8733574

  13. Intracellular and extracellular forces drive primary cilia movement.

    PubMed

    Battle, Christopher; Ott, Carolyn M; Burnette, Dylan T; Lippincott-Schwartz, Jennifer; Schmidt, Christoph F

    2015-02-01

    Primary cilia are ubiquitous, microtubule-based organelles that play diverse roles in sensory transduction in many eukaryotic cells. They interrogate the cellular environment through chemosensing, osmosensing, and mechanosensing using receptors and ion channels in the ciliary membrane. Little is known about the mechanical and structural properties of the cilium and how these properties contribute to ciliary perception. We probed the mechanical responses of primary cilia from kidney epithelial cells [Madin-Darby canine kidney-II (MDCK-II)], which sense fluid flow in renal ducts. We found that, on manipulation with an optical trap, cilia deflect by bending along their length and pivoting around an effective hinge located below the basal body. The calculated bending rigidity indicates weak microtubule doublet coupling. Primary cilia of MDCK cells lack interdoublet dynein motors. Nevertheless, we found that the organelles display active motility. 3D tracking showed correlated fluctuations of the cilium and basal body. These angular movements seemed random but were dependent on ATP and cytoplasmic myosin-II in the cell cortex. We conclude that force generation by the actin cytoskeleton surrounding the basal body results in active ciliary movement. We speculate that actin-driven ciliary movement might tune and calibrate ciliary sensory functions. PMID:25605896

  14. A comprehensive DNA methylation profile of epithelial-to-mesenchymal transition.

    PubMed

    Carmona, F Javier; Davalos, Veronica; Vidal, Enrique; Gomez, Antonio; Heyn, Holger; Hashimoto, Yutaka; Vizoso, Miguel; Martinez-Cardus, Anna; Sayols, Sergi; Ferreira, Humberto J; Sánchez-Mut, Jose V; Morán, Sebastián; Margelí, Mireia; Castella, Eva; Berdasco, Maria; Stefansson, Olafur A; Eyfjord, Jorunn E; Gonzalez-Suarez, Eva; Dopazo, Joaquín; Orozco, Modesto; Gut, Ivo G; Esteller, Manel

    2014-10-01

    Epithelial-to-mesenchymal transition (EMT) is a plastic process in which fully differentiated epithelial cells are converted into poorly differentiated, migratory and invasive mesenchymal cells, and it has been related to the metastasis potential of tumors. This is a reversible process and cells can also eventually undergo mesenchymal-to-epithelial transition. The existence of a dynamic EMT process suggests the involvement of epigenetic shifts in the phenotype. Herein, we obtained the DNA methylomes at single-base resolution of Madin-Darby canine kidney cells undergoing EMT and translated the identified differentially methylated regions to human breast cancer cells undergoing a gain of migratory and invasive capabilities associated with the EMT phenotype. We noticed dynamic and reversible changes of DNA methylation, both on promoter sequences and gene-bodies in association with transcription regulation of EMT-related genes. Most importantly, the identified DNA methylation markers of EMT were present in primary mammary tumors in association with the epithelial or the mesenchymal phenotype of the studied breast cancer samples. PMID:25106427

  15. Podocyte-specific expression of organic cation transporter PMAT: implication in puromycin aminonucleoside nephrotoxicity

    PubMed Central

    Xia, Li; Zhou, Mingyan; Kalhorn, Thomas F.; Ho, Horace T. B.; Wang, Joanne

    2009-01-01

    Plasma membrane monoamine transporter (PMAT) is a novel polyspecific organic cation transporter that transports organic cations and the purine nucleoside, adenosine. PMAT is expressed in the kidney, but the specific localization and function of this transporter in renal cells are unclear. In this study, we developed a polyclonal antibody toward a 14-amino acid sequence in the last intracellular loop of PMAT and determined the precise cellular localization of PMAT in human and rat kidneys. Surprisingly, we found that the PMAT protein was predominantly expressed in the glomerulus with minimal expression in tubular cells. Within the glomerulus, dual-color immunofluorescence labeling showed that the PMAT protein was specifically localized to the visceral glomerular epithelial cells, i.e., podocytes. There was no significant PMAT immunoreactivity in mesangial or glomerular endothelial cells. We further showed that puromycin aminonucleoside (PAN), a classic podocyte toxin that induces massive proteinuria and severe glomerulopathy, is transported by PMAT. Expression of PMAT in Madin-Darby canine kidney cells significantly increased cell sensitivity to PAN. Decynium 22, a potent PMAT inhibitor, abolished PAN toxicity in PMAT-expressing cells. Together, our data suggest that PMAT is specifically expressed in podocytes and may play an important role in PAN-induced kidney injury. PMID:19357181

  16. Characterization of the interaction between protein 4.1R and ZO-2. A possible link between the tight junction and the actin cytoskeleton.

    PubMed

    Mattagajasingh, S N; Huang, S C; Hartenstein, J S; Benz, E J

    2000-09-29

    Multiple isoforms of the red cell protein 4.1R are expressed in nonerythroid cells, including novel 135-kDa isoforms. Using a yeast two-hybrid system, immunocolocalization, immunoprecipitation, and in vitro binding studies, we found that two 4.1R isoforms of 135 and 150 kDa specifically interact with the protein ZO-2 (zonula occludens-2). 4.1R is colocalized with ZO-2 and occludin at Madin-Darby canine kidney (MDCK) cell tight junctions. Both isoforms of 4.1R coprecipitated with proteins that organize tight junctions such as ZO-2, ZO-1, and occludin. Western blot analysis also revealed the presence of actin and alpha-spectrin in these immunoprecipitates. Association of 4.1R isoforms with these tight junction and cytoskeletal proteins was found to be specific for the tight junction and was not seen in nonconfluent MDCK cells. The amino acid residues that sustain the interaction between 4.1R and ZO-2 reside within the amino acids encoded by exons 19-21 of 4.1R and residues 1054-1118 of ZO-2. Exogenously expressed 4.1R containing the spectrin/actin- and ZO-2-binding domains was recruited to tight junctions in confluent MDCK cells. Taken together, our results suggest that 4.1R might play an important role in organization and function of the tight junction by establishing a link between the tight junction and the actin cytoskeleton. PMID:10874042

  17. Photocatalytic inactivation efficiency of anatase Nano-TiO(2) Sol on the H(9) N(2) avian influenza virus.

    PubMed

    Cui, Haixin; Jiang, Jianfang; Gu, Wei; Sun, Changjiao; Wu, Donglai; Yang, Tao; Yang, Guochen

    2010-01-01

    This study was conducted to investigate efficiency of TiO(2) nanomaterial as a novel environment-friendly disinfectant to control avian influenza (AI) by its photochemical sterilization ability. Anatase nano-TiO(2) sol, a neutral, viscous aqueous colloid of 1.6% TiO(2) , was synthesized from peroxotitanic acid solution according to the Ichinose method. Transmission electron microscope images showed that the TiO(2) particles were spindle-shaped with an average size of 50 nm. X-ray diffraction patterns revealed that the crystal phase of TiO(2) particles was anatase type with photocatalytic effect. A photocatalytic film of nano-TiO(2) sol was tested as a means of inactivating H(9) N(2) avian influenza virus (AIV). Inactivation capabilities were examined with 365nm ultraviolet (UV) radiation under black light by adjusting the UV intensity, the UV irradiation time and the quantity of AIV. The titer change of AIV was determined by hemagglutination tests. Cytopathic effect of Madin Darby canine kidney (MDCK) cells was monitored by inverted fluorescence microscope. The results showed that anatase nano-TiO(2) sol significantly inactivated AIV under UV irradiation of 365nm. The inactivation of AIV viruses reached up to 100%. Therefore, anatase nano-TiO(2) sol is a potentially environment-friendly antivirus agent to prevent AI. PMID:20553405

  18. Preparation of single cells for imaging mass spectrometry.

    PubMed

    Berman, Elena S F; Fortson, Susan L; Kulp, Kristen S

    2010-01-01

    Characterizing the molecular contents of individual cells is critical for understanding fundamental mechanisms of biological processes. Imaging mass spectrometry (IMS) of biological systems has been steadily gaining popularity for its ability to create precise chemical images of biological samples, thereby revealing new biological insights and improving understanding of disease. In order to acquire mass spectral images from single cells that contain relevant molecular information, samples must be prepared such that cell-culture components, especially salts, are eliminated from the cell surface and that the cell contents are accessible to the mass spectrometer. We have demonstrated a cellular preparation technique for IMS that preserves the basic morphology of cultured cells, allows mass spectrometric chemical profiling of cytosol, and removes the majority of the interfering species derived from the cellular growth medium. Using this protocol, we achieve high-quality, reproducible IMS images from three diverse cell types: MCF7 human breast cancer cells, Madin-Darby canine kidney (MDCK) cells, and NIH/3T3 mouse fibroblasts. This preparation method allows rapid and routine IMS analysis of cultured cells, making possible a wide variety of experiments to further scientific understanding of molecular processes within individual cells. PMID:20680596

  19. A continuous peptide epitope reacting with pandemic influenza AH1N1 predicted by bioinformatic approaches.

    PubMed

    Carrillo-Vazquez, Jonathan P; Correa-Basurto, José; García-Machorro, Jazmin; Campos-Rodríguez, Rafael; Moreau, Violaine; Rosas-Trigueros, Jorge L; Reyes-López, Cesar A; Rojas-López, Marlon; Zamorano-Carrillo, Absalom

    2015-09-01

    Computational identification of potential epitopes with an immunogenic capacity challenges immunological research. Several methods show considerable success, and together with experimental studies, the efficiency of the algorithms to identify potential peptides with biological activity has improved. Herein, an epitope was designed by combining bioinformatics, docking, and molecular dynamics simulations. The hemagglutinin protein of the H1N1 influenza pandemic strain served as a template, owing to the interest of obtaining a scheme of immunization. Afterward, we performed enzyme-linked immunosorbent assay (ELISA) using the epitope to analyze if any antibodies in human sera before and after the influenza outbreak in 2009 recognize this peptide. Also, a plaque reduction neutralization test induced by virus-neutralizing antibodies and the IgG determination showed the biological activity of this computationally designed peptide. The results of the ELISAs demonstrated that the serum of both prepandemic and pandemic recognized the epitope. Moreover, the plaque reduction neutralization test evidenced the capacity of the designed peptide to neutralize influenza virus in Madin-Darby canine cells. PMID:25788327

  20. Abcb1 in Pigs: Molecular cloning, tissues distribution, functional analysis, and its effect on pharmacokinetics of enrofloxacin

    PubMed Central

    Guo, Tingting; Huang, Jinhu; Zhang, Hongyu; Dong, Lingling; Guo, Dawei; Guo, Li; He, Fang; Bhutto, Zohaib Ahmed; Wang, Liping

    2016-01-01

    P-glycoprotein (P-gp) is one of the best-known ATP-dependent efflux transporters, contributing to differences in pharmacokinetics and drug-drug interactions. Until now, studies on pig P-gp have been scarce. In our studies, the full-length porcine P-gp cDNA was cloned and expressed in a Madin-Darby Canine Kidney (MDCK) cell line. P-gp expression was then determined in tissues and its role in the pharmacokinetics of oral enrofloxacin in pigs was studied. The coding region of pig Abcb1 gene was 3,861 bp, encoding 1,286 amino acid residues (Mw = 141,966). Phylogenetic analysis indicated a close evolutionary relationship between porcine P-gp and those of cow and sheep. Pig P-gp was successfully stably overexpressed in MDCK cells and had efflux activity for rhodamine 123, a substrate of P-gp. Tissue distribution analysis indicated that P-gp was highly expressed in brain capillaries, small intestine, and liver. In MDCK-pAbcb1 cells, enrofloxacin was transported by P-gp with net efflux ratio of 2.48 and the efflux function was blocked by P-gp inhibitor verapamil. High expression of P-gp in the small intestine could modify the pharmacokinetics of orally administrated enrofloxacin by increasing the Cmax, AUC and Ka, which was demonstrated using verapamil, an inhibitor of P-gp. PMID:27572343

  1. Ankyrin-G palmitoylation and βII-spectrin binding to phosphoinositide lipids drive lateral membrane assembly

    PubMed Central

    He, Meng; Abdi, Khadar M.

    2014-01-01

    Ankyrin-G and βII-spectrin colocalize at sites of cell–cell contact in columnar epithelial cells and promote lateral membrane assembly. This study identifies two critical inputs from lipids that together provide a rationale for how ankyrin-G and βII-spectrin selectively localize to Madin-Darby canine kidney (MDCK) cell lateral membranes. We identify aspartate-histidine-histidine-cysteine 5/8 (DHHC5/8) as ankyrin-G palmitoyltransferases required for ankyrin-G lateral membrane localization and for assembly of lateral membranes. We also find that βII-spectrin functions as a coincidence detector that requires recognition of both ankyrin-G and phosphoinositide lipids for its lateral membrane localization. DHHC5/8 and βII-spectrin colocalize with ankyrin-G in micrometer-scale subdomains within the lateral membrane that are likely sites for palmitoylation of ankyrin-G. Loss of either DHHC5/8 or ankyrin-G–βII-spectrin interaction or βII-spectrin–phosphoinositide recognition through its pleckstrin homology domain all result in failure to build the lateral membrane. In summary, we identify a functional network connecting palmitoyltransferases DHHC5/8 with ankyrin-G, ankyrin-G with βII-spectrin, and βII-spectrin with phosphoinositides that is required for the columnar morphology of MDCK epithelial cells. PMID:25049274

  2. The Rab5 Effector Rabankyrin-5 Regulates and Coordinates Different Endocytic Mechanisms

    PubMed Central

    2004-01-01

    The small GTPase Rab5 is a key regulator of clathrin-mediated endocytosis. On early endosomes, within a spatially restricted domain enriched in phosphatydilinositol-3-phosphate [PI(3)P], Rab5 coordinates a complex network of effectors that functionally cooperate in membrane tethering, fusion, and organelle motility. Here we discovered a novel PI(3)P-binding Rab5 effector, Rabankyrin-5, which localises to early endosomes and stimulates their fusion activity. In addition to early endosomes, however, Rabankyrin-5 localises to large vacuolar structures that correspond to macropinosomes in epithelial cells and fibroblasts. Overexpression of Rabankyrin-5 increases the number of macropinosomes and stimulates fluid-phase uptake, whereas its downregulation inhibits these processes. In polarised epithelial cells, this function is primarily restricted to the apical membrane. Rabankyrin-5 localises to large pinocytic structures underneath the apical surface of kidney proximal tubule cells, and its overexpression in polarised Madin-Darby canine kidney cells stimulates apical but not basolateral, non-clathrin-mediated pinocytosis. In demonstrating a regulatory role in endosome fusion and (macro)pinocytosis, our studies suggest that Rab5 regulates and coordinates different endocytic mechanisms through its effector Rabankyrin-5. Furthermore, its active role in apical pinocytosis in epithelial cells suggests an important function of Rabankyrin-5 in the physiology of polarised cells. PMID:15328530

  3. Effectiveness of Periodic Treatment of Quercetin against Influenza A Virus H1N1 through Modulation of Protein Expression.

    PubMed

    Vaidya, Bipin; Cho, Se-Young; Oh, Kyung-Seo; Kim, Song Hak; Kim, Yeong O; Jeong, Eun-Hye; Nguyen, Thoa Thi; Kim, Sung Hyun; Kim, In Seon; Kwon, Joseph; Kim, Duwoon

    2016-06-01

    Kimchi, a traditional fermented food regularly consumed in Korea, contains various types of antimicrobial compounds. Among the tested compounds present in common spices used in Kimchi, quercetin showed the highest selectivity index against influenza A virus (IAV) H1N1. In this study, the effect of pretreatment and periodic treatment with quercetin against IAV in Madin-Darby canine kidney cells was observed. Compared to pretreatment, periodic treatment resulted in significantly higher cell viability but lower relative expression of the IAV PA gene and total apoptosis and cell death. To explain the mechanisms underlying the antiviral effects of quercetin treatment, a comparative proteomic analysis was performed in four samples (mock, quercetin-treated, IAV-infected, and quercetin-treated IAV-infected). Among the 220 proteins, 56 proteins were classified nonhierarchically into three clusters and were differentially modulated by quercetin treatment in IAV-infected cells. Post-translational modifications were identified in 68 proteins. In conclusion, periodic treatment with quercetin is effective in reducing IAV infection, and differentially regulates the expression of key proteins, including heat shock proteins, fibronectin 1, and prohibitin to reduce IAV replication. PMID:27157719

  4. Antiviral Protein of Momordica charantia L. Inhibits Different Subtypes of Influenza A

    PubMed Central

    Pongthanapisith, Viroj; Ikuta, Kazuyoshi; Puthavathana, Pilaipan; Leelamanit, Wichet

    2013-01-01

    The new antiviral activity of the protein extracted from Momordica charantia was determined with different subtypes of influenza A. The protein was purified from the seed of M. charantia using an anion exchanger and a Fast Protein Liquid Chromatography (FPLC) system. At the concentration of 1.401 mg/mL, the protein did not exhibit cytotoxicity in Madin-Darby canine kidney cells (MDCK) but inhibited 1 × 105 FFU influenza A/PR/8/34 H1N1 virus at 56.50%, 65.72%, and 100% inhibition by the protein treated before the virus (pretreated), the protein treated alongside with the virus (simultaneously treated), and the protein treated after the virus (posttreated) during incubation, respectively. Using 5, 25, and 100 TCID50 of influenza A/New Caledonia/20/99 H1N1, A/Fujian/411/01 H3N2 and A/Thailand/1(KAN-1)/2004 H5N1, the IC50 was calculated to be 100, 150, and 200; 75, 175, and 300; and 40, 75, and 200 μg/mL, respectively. Our present finding indicated that the plant protein inhibited not only H1N1 and H3N2 but also H5N1 subtype. As a result of the broad spectrum of its antiviral activity, this edible plant can be developed as an effective therapeutic agent against various and even new emerging subtypes of influenza A. PMID:23935676

  5. Development of a sensitive in vitro assay to quantify the biological activity of pro-inflammatory phorbol esters in Jatropha oil.

    PubMed

    Pelletier, Guillaume; Padhi, Bhaja K; Hawari, Jalal; Sunahara, Geoffrey I; Poon, Raymond

    2015-06-01

    New health safety concerns may arise from the increasing production and use of Jatropha oil, a biodiesel feedstock that also contains toxic, pro-inflammatory, and co-carcinogenic phorbol esters. Based on the exceptional sensitivity of Madin-Darby canine kidney (MDCK) cells to the model phorbol ester 12-O-tetradecanoylphorbol-13-acetate (TPA), a robust bioassay was developed to quantify the biological activity of Jatropha phorbol esters directly in oil, without sample extraction. We first verified that the characteristic response of MDCK cells to TPA was also observed following direct exposure to phorbol esters in Jatropha oil. We further confirmed that similarly to TPA, Jatropha oil's phorbol esters can activate protein kinase C (PKC). We then assessed the transcriptional response of MDCK cells to Jatropha oil exposure by measuring the expression of cyclooxygenase-2 (COX-2), a gene involved in inflammatory processes which is strongly upregulated following PKC activation. Based on the parameterization of a TPA dose-response curve, the transcriptional response of MDCK cells to Jatropha oil exposure was expressed in term of TPA toxic equivalent (TEQ), a convenient metric to report the inflammatory potential of complex mixtures. The sensitive bioassay described in this manuscript may prove useful for risk assessment, as it provides a quantitative method and a convenient metric to report the inflammatory potential of phorbol esters in Jatropha oil. This bioassay may also be adapted for the detection of bioactive phorbol esters in other matrices. PMID:25588777

  6. Inhibition of endosomal fusion activity of influenza virus by Rheum tanguticum (da-huang).

    PubMed

    Lin, Ta-Jen; Lin, Chwan-Fwu; Chiu, Cheng-Hsun; Lee, Ming-Chung; Horng, Jim-Tong

    2016-01-01

    Rhubarb (Rheum tanguticum; da-huang in Chinese medicine) is a herbal medicine that has been used widely for managing fever and removing toxicity. In this study, we investigated how rhubarb inhibits influenza virus during the early stage of the infectious cycle using different functional assays. A non-toxic ethanolic extract of rhubarb (Rex) inhibited several H1N1 subtypes of influenza A viruses in Madin-Darby canine kidney cells, including strains that are clinically resistant to oseltamivir. Time course analysis of Rex addition showed that viral entry was one of the steps that was inhibited by Rex. We also confirmed that Rex effectively inhibited viral attachment and penetration into the host cells. The inhibition of red blood cell haemolysis and cell-cell fusion by Rex suggests that Rex may block haemagglutinin-mediated fusion (virus-endosome fusion) during the fusion/uncoating step. Rex has the capacity to inhibit influenza viruses by blocking viral endocytosis. Thus, rhubarb might provide an alternative therapeutic approach when resistant viruses become more prevalent. PMID:27302738

  7. Numb directs the subcellular localization of EAAT3 through binding the YxNxxF motif.

    PubMed

    Su, Jin-Feng; Wei, Jian; Li, Pei-Shan; Miao, Hong-Hua; Ma, Yong-Chao; Qu, Yu-Xiu; Xu, Jie; Qin, Jie; Li, Bo-Liang; Song, Bao-Liang; Xu, Zheng-Ping; Luo, Jie

    2016-08-15

    Excitatory amino acid transporter type 3 (EAAT3, also known as SLC1A1) is a high-affinity, Na(+)-dependent glutamate carrier that localizes primarily within the cell and at the apical plasma membrane. Although previous studies have reported proteins and sequence regions involved in EAAT3 trafficking, the detailed molecular mechanism by which EAAT3 is distributed to the correct location still remains elusive. Here, we identify that the YVNGGF sequence in the C-terminus of EAAT3 is responsible for its intracellular localization and apical sorting in rat hepatoma cells CRL1601 and Madin-Darby canine kidney (MDCK) cells, respectively. We further demonstrate that Numb, a clathrin adaptor protein, directly binds the YVNGGF motif and regulates the localization of EAAT3. Mutation of Y503, N505 and F508 within the YVNGGF motif to alanine residues or silencing Numb by use of small interfering RNA (siRNA) results in the aberrant localization of EAAT3. Moreover, both Numb and the YVNGGF motif mediate EAAT3 endocytosis in CRL1601 cells. In summary, our study suggests that Numb is a pivotal adaptor protein that mediates the subcellular localization of EAAT3 through binding the YxNxxF (where x stands for any amino acid) motif. PMID:27358480

  8. Poor oral bioavailability of a promising anticancer agent andrographolide is due to extensive metabolism and efflux by P-glycoprotein.

    PubMed

    Ye, Ling; Wang, Tao; Tang, Lan; Liu, Wei; Yang, Zhen; Zhou, Juan; Zheng, Zhijie; Cai, Zheng; Hu, Ming; Liu, Zhongqiu

    2011-11-01

    Andrographolide (AP), isolated from Andrographis paniculata (Burm. F.) Nees, is an anticancer agent with significant clinical potential. This study determined its oral bioavailability and how intestinal disposition affects its bioavailability. Pharmacokinetics was evaluated in rats. Intestinal disposition was determined using a single-pass rat intestinal perfusion model and the cultured Caco-2 cells and Madin-Darby canine kidney II cells over expressing human P-gp (MDR1-MDCKII). Absolute bioavailability of AP was 2.67%. In the duodenum and jejunum, AP was rapidly metabolized to a sulfonate, identified as 14-deoxy-12-sulfo- andrographolide. AP was also rapidly metabolized by liver S9 fraction and in blank perfusates collected from duodenum and jejunum. The apparent permeability (P(app) ) of AP from basolateral (B) to apical (A) (4.94 × 10 cm/s) in the Caco-2 model was four times higher than the P(app) from A to B (1.14 × 10(-5) cm/s). Moreover, AP was significantly more permeable in the B to A direction than the opposite direction in MDR1-MDCKII cells. In the perfusion model, the effective permeability (P*(eff) ) for AP was highest in the duodenum, followed by jejunum, and then ileum and colon. In the ileum and colon, the P*(eff) for AP was significantly increased by verapamil, a P-glycoprotein (P-gp) inhibitor. AP has poor oral bioavailability because of its rapid biotransformation and efflux by P-gp. PMID:21721007

  9. Preparation of Single Cells for Imaging Mass Spectrometry

    SciTech Connect

    Berman, E S; Fortson, S L; Kulp, K S; Checchi, K D; Wu, L; Felton, J S; Wu, K J

    2007-10-24

    Characterizing chemical changes within single cells is important for determining fundamental mechanisms of biological processes that will lead to new biological insights and improved disease understanding. Imaging biological systems with mass spectrometry (MS) has gained popularity in recent years as a method for creating precise chemical maps of biological samples. In order to obtain high-quality mass spectral images that provide relevant molecular information about individual cells, samples must be prepared so that salts and other cell-culture components are removed from the cell surface and the cell contents are rendered accessible to the desorption beam. We have designed a cellular preparation protocol for imaging MS that preserves the cellular contents for investigation and removes the majority of the interfering species from the extracellular matrix. Using this method, we obtain excellent imaging results and reproducibility in three diverse cell types: MCF7 human breast cancer cells, Madin-Darby canine kidney (MDCK) cells, and NIH/3T3 mouse fibroblasts. This preparation technique allows routine imaging MS analysis of cultured cells, allowing for any number of experiments aimed at furthering scientific understanding of molecular processes within individual cells.

  10. Herpesvirus infections in rock hyraxes (Procavia capensis).

    PubMed

    Galeota, Judith A; Napier, Julia E; Armstrong, Douglas L; Riethoven, Jean-Jack; Rogers, Douglas G

    2009-07-01

    Seven juveniles and 3 adults from a closed group of 19 rock hyraxes (Procavia capensis) housed in a zoo's indoor rock exhibit died or were euthanized after developing blepharoconjunctivitis and orofacial ulcers over a 2-week period. Histopathologic examination of dermal ulcers and ulcerated tongues revealed amphophilic to basophilic intranuclear inclusion bodies in epithelial cells bordering ulcers. Epithelial cells with inclusion bodies were often characterized by cytomegaly and karyomegaly, and many cells had formed syncytia. Examination of inclusion bodies in tongue epithelium by transmission electron microscopy revealed icosahedral nucleocapsids, approximately 80-95 nm in diameter, with morphologic features consistent with herpesvirus. Cytopathic effect (CPE) typical of alphaherpesvirus infection was seen in bovine turbinate, equine dermal, and Vero cell monolayers after inoculation with homogenates of the skin lesions, but CPE was not seen after inoculation onto Madin-Darby canine kidney or swine testicle cell monolayers. Polymerase chain reaction analysis using degenerate primers that targeted a portion of the herpesvirus polymerase gene generated a product of approximately 227 base pairs. The product was cloned, sequenced, and then analyzed using BLAST. At the nucleotide level, there was 86%, 77%, and 76% shared identity with Eidolon herpesvirus 1, Human herpesviruses 1 and 2, and Cercopithecine herpesvirus 2, respectively. Herpesvirus infections in rock hyraxes have not been characterized. The data presented in the current study suggest that a novel alphaherpesvirus caused the lesions seen in these rock hyraxes. The molecular characteristics of this virus would tentatively support its inclusion in the genus Simplexvirus. PMID:19564505

  11. Disruption of Adherens Junctions Liberates Nectin-1 To Serve as Receptor for Herpes Simplex Virus and Pseudorabies Virus Entry

    PubMed Central

    Yoon, Miri; Spear, Patricia G.

    2002-01-01

    Nectin-1, a cell adhesion molecule belonging to the immunoglobulin superfamily, can bind to virion glycoprotein D (gD) to mediate entry of herpes simplex viruses (HSV) and pseudorabies virus (PRV). Nectin-1 colocalizes with E-cadherin at adherens junctions in epithelial cells. The disruption of cell junctions can result in the redistribution of nectin-1. To determine whether disruption of junctions by calcium depletion influenced the susceptibility of epithelial cells to viral entry, Madin-Darby canine kidney cells expressing endogenous nectin-1 or transfected human nectin-1 were tested for the ability to bind soluble forms of viral gD and to be infected by HSV and PRV, before and after calcium depletion. Confocal microscopy revealed that binding of HSV and PRV gD was localized to adherens junctions in cells maintained in normal medium but was distributed, along with nectin-1, over the entire cell surface after calcium depletion. Both the binding of gD and the fraction of cells that could be infected by HSV-1 and PRV were enhanced by calcium depletion. Taken together, these results provide evidence that nectin-1 confined to adherens junctions in epithelial cells is not very accessible to virus, whereas dissociation of cell junctions releases nectin-1 to serve more efficiently as an entry receptor. PMID:12072519

  12. Constitutively expressing cell lines that secrete a truncated bovine herpes virus-1 glycoprotein (gpI) stimulate T-lymphocyte responsiveness.

    PubMed Central

    Leary, T P; Gao, Y; Splitter, G A

    1992-01-01

    The desire to obtain authentically glycosylated viral protein products in sufficient quantity for immunological study has led to the use of eucaryotic expression vectors for protein production. An additional advantage is that these protein products can be studied individually in the absence of their native viral environment. We have cloned a complementary DNA (cDNA) encoding bovine herpes virus-1 (BHV-1) glycoprotein 1 (gpI) into the eucaryotic expression vector, pZipNeo SVX1. Since this protein is normally embedded within the membrane of BHV-1 infected cells, we removed sequences encoding the transmembrane domain of the native protein. After transfection of the plasmid construct into the canine osteosarcoma cell line, D17, or Madin-Darby bovine kidney (MDBK) cells, a truncated BHV-1 (gpI) was secreted into the culture medium as demonstrated by radioimmunoprecipitation and SDS-PAGE. Both a CD4+ T-lymphocyte line specific for BHV-1 and freshly isolated T lymphocytes could recognize and respond to the secreted recombinant gpI. Further, recombinant gpI could elicit both antibody and cellular responses in cattle when used as an immunogen. Having established constitutively glycoprotein producing cell lines, future studies in vaccine evaluation of gpI will be facilitated. Images Figure 1 Figure 2 PMID:1526647

  13. Safety, tolerability and immunogenicity of a mammalian cell-culture-derived influenza vaccine: a sequential Phase I and Phase II clinical trial.

    PubMed

    Groth, N; Montomoli, E; Gentile, C; Manini, I; Bugarini, R; Podda, A

    2009-01-29

    This sequential, observer-blind, randomised, single-centre, combined Phase I and Phase II clinical trial compared the tolerability and immunogenicity of a single intramuscular dose of a novel cell-culture-derived influenza vaccine (CCIV), produced in Madin-Darby canine kidney cells, with a conventional egg-based vaccine. The immunogenicity of both vaccines was assessed by SRH assay, a well-recognized test by EMEA, in compliance with the requirements of the EU Committee for Medicinal Products for Human Use (CHMP). The Phase I part of the trial comprised 40 healthy adults (18-40 years of age); the subsequent Phase II part involved 200 healthy adult (n=80, 18-60 years of age) and elderly (n=120, > or =61 years of age) subjects. Both vaccines showed similar reactogenicity and any solicited local or systemic reactions were mostly mild or moderate. Regarding immunogenicity, both the CCIV and the control vaccine met all of the EU Committee for Medicinal Products for Human Use criteria for influenza vaccines for each strain and in both age groups. In conclusion, the CCIV produced in mammalian cell-culture is as well tolerated and as immunogenic as the control egg-based vaccine in non-elderly and elderly adults. PMID:19027046

  14. Human cell lines used in a micro neutralization test for measuring influenza-neutralizing antibodies.

    PubMed

    Mittelholzer, C M; Brokstad, K A; Pauksens, K; Jonsson, R; Brytting, M; Linde, A

    2006-04-01

    An in situ neutralization test (NT) including ELISA for the measurement of influenza antigen was developed and evaluated. Two human cell lines, fibroblasts (HS27) cells and salivary gland epithelial duct (HSG) cells, were compared with Madin-Darby Canine Kidney (MDCK) cells. The viral production in the human cell lines was lower than that for MDCK cells, which influenced the results of the assay in the HSG and HS27 cells. However, when lowering the infectious dose, the NT using HS27 cells gave a sensitive and stable assay with low background in the ELISA. The NT titres were very low when using HSG cells compared to MDCK cells. The HS27 NT was used to analyze the humoral response after an influenza A infection in patients from a placebo-controlled zanamivir study. We found no differences in NT titres between patients treated with zanamivir or placebo. The MDCK and HS27 NT gave higher titres and more pronounced titre differences than the gold standard haemagglutinin inhibition (HAI) assay. Compared to the HAI assay, the sensitive NT using HS27 cells also revealed heterologous NT-titre rises after influenza infection in the patients. PMID:16623925

  15. Abcb1 in Pigs: Molecular cloning, tissues distribution, functional analysis, and its effect on pharmacokinetics of enrofloxacin.

    PubMed

    Guo, Tingting; Huang, Jinhu; Zhang, Hongyu; Dong, Lingling; Guo, Dawei; Guo, Li; He, Fang; Bhutto, Zohaib Ahmed; Wang, Liping

    2016-01-01

    P-glycoprotein (P-gp) is one of the best-known ATP-dependent efflux transporters, contributing to differences in pharmacokinetics and drug-drug interactions. Until now, studies on pig P-gp have been scarce. In our studies, the full-length porcine P-gp cDNA was cloned and expressed in a Madin-Darby Canine Kidney (MDCK) cell line. P-gp expression was then determined in tissues and its role in the pharmacokinetics of oral enrofloxacin in pigs was studied. The coding region of pig Abcb1 gene was 3,861 bp, encoding 1,286 amino acid residues (Mw = 141,966). Phylogenetic analysis indicated a close evolutionary relationship between porcine P-gp and those of cow and sheep. Pig P-gp was successfully stably overexpressed in MDCK cells and had efflux activity for rhodamine 123, a substrate of P-gp. Tissue distribution analysis indicated that P-gp was highly expressed in brain capillaries, small intestine, and liver. In MDCK-pAbcb1 cells, enrofloxacin was transported by P-gp with net efflux ratio of 2.48 and the efflux function was blocked by P-gp inhibitor verapamil. High expression of P-gp in the small intestine could modify the pharmacokinetics of orally administrated enrofloxacin by increasing the Cmax, AUC and Ka, which was demonstrated using verapamil, an inhibitor of P-gp. PMID:27572343

  16. IQGAP1 controls tight junction formation through differential regulation of claudin recruitment

    PubMed Central

    Tanos, Barbara E.; Perez Bay, Andres E.; Salvarezza, Susana; Vivanco, Igor; Mellinghoff, Ingo; Osman, Mahasin; Sacks, David B.; Rodriguez-Boulan, Enrique

    2015-01-01

    ABSTRACT IQGAP1 is a scaffolding protein previously implicated in adherens junction formation. However, its role in the establishment or maintenance of tight junctions (TJs) has not been explored. We hypothesized that IQGAP1 could regulate TJ formation by modulating the expression and/or localization of junctional proteins, and we systematically tested this hypothesis in the model Madin-Darby canine kidney (MDCK) cell line. We find that IQGAP1 silencing enhances a transient increase in transepithelial electrical resistance (TER) observed during the early stages of TJ formation (Cereijido et al., 1978). Quantitative microscopy and biochemical experiments suggest that this effect of IQGAP1 on TJ assembly is accounted for by reduced expression and TJ recruitment of claudin 2, and increased TJ recruitment of claudin 4. Furthermore, we show that IQGAP1 also regulates TJ formation through its interactor CDC42, because IQGAP1 knockdown increases the activity of the CDC42 effector JNK and dominant-negative CDC42 prevents the increase in TER caused by IQGAP1 silencing. Hence, we provide evidence that IQGAP1 modulates TJ formation by a twofold mechanism: (1) controlling the expression and recruitment of claudin 2 and recruitment of claudin 4 to the TJ, and (2) transient inhibition of the CDC42–JNK pathway. PMID:25588839

  17. A Series of New Ligustrazine-Triterpenes Derivatives as Anti-Tumor Agents: Design, Synthesis, and Biological Evaluation

    PubMed Central

    Xu, Bing; Chu, Fuhao; Zhang, Yuzhong; Wang, Xiaobo; Li, Qiang; Liu, Wei; Xu, Xin; Xing, Yanyi; Chen, Jing; Wang, Penglong; Lei, Haimin

    2015-01-01

    A series of novel ligustrazine-triterpenes derivatives was designed, synthesized and screened for their cytotoxicity against five cancer cell lines (Bel-7402, HepG2, HT-29, Hela, and MCF-7) and Madin-Darby canine kidney (MDCK). Current study suggested that most of the ligustrazine-triterpenes conjunctions showed better cytotoxicity than the starting materials. In particular, compound 4a exhibited better cytotoxic activity (IC50 < 5.23 μM) against Bel-7402, HT-29, MCF-7, Hela, and HepG2 than the standard anticancer drug cisplatin (DDP). The cytotoxicity selectivity detection revealed that 4a exhibited low cytotoxicity (IC50 > 20 μM) towards MDCK cells. A combination of fluorescence staining observation and flow cytometric analysis indicated that 4a could induce HepG2 cell apoptosis. Further studies suggested that 4a-induced apoptosis is mediated through depolarization of the mitochondrial membrane potential and increase of intracellular free Ca2+ concentration. In addition, the structure-activity relationships of these derivatives were briefly discussed. PMID:26404253

  18. Chrysin-benzothiazole conjugates as antioxidant and anticancer agents.

    PubMed

    Mistry, Bhupendra M; Patel, Rahul V; Keum, Young-Soo; Kim, Doo Hwan

    2015-12-01

    7-(4-Bromobutoxy)-5-hydroxy-2-phenyl-4H-chromen-4-one, obtained from chrysin with 1,4-dibromobutane, was combined with a wide range of 6-substituted 2-aminobenzthiazoles, which had been prepared from the corresponding anilines with potassium thiocyanate. Free radical scavenging efficacies of newer analogues were measured using DPPH and ABTS assays, in addition to the assessment of their anticancer activity against cervical cancer cell lines (HeLa and CaSki) and ovarian cancer cell line (SK-OV-3) implementing the SRB assay. Cytotoxicity of titled compounds was checked using Madin-Darby canine kidney (MDCK) non-cancer cell line. Overall, 6a-r indicated remarkable antioxidant power as DPPH and ABTS(+) scavengers; particularly the presence of halogen(s) (6g, 6h, 6j-6l) was favourable with IC50 values comparable to the control ascorbic acid. Unsubstituted benzothiazole ring favored the activity of resultant compounds (6a and 6r) against HeLa cell line, whereas presence of chlorine (6g) or a di-fluoro group (6k) was a key to exert strong action against CaSki. Moreover, a mono-fluoro (6j) and a ketonic functionality (6o) were beneficial to display anticipated anticancer effects against ovarian cancer cell line SK-OV-3. The structural assignments of the new products were done on the basis of IR, (1)H NMR, (13)C NMR spectroscopy and elemental analysis. PMID:26514745

  19. Electrical Measurement of Biological Oscillations in Unicellular Systems.

    NASA Astrophysics Data System (ADS)

    Giaever, Ivar; Linton, Michael; Halvorsrud, Ragnhild; Male, Tone

    1997-03-01

    Many different rhythms or oscillations exists in biological systems; the circadian rhythm is probably the best known, but other oscillations are also common. The slimemold Physarum is a classical example of ultradian oscillations occurring in a single multinucleated cell, and the period is a few minutes long. It is also reasonable well known that oscillations occur in yeast with a periods of a few minutes These oscillations result from the action of a single allosteric enzyme in the glycolytic pathway. We have recently discovered a new oscillating system in the Madin-Darby Canine Kidney (MDCK) epithelial cells that have a period of 2-3 hours, and at present the origin of these oscillations is unknown. All these systems have been studied for the first time using a biosensor that measures impedance changes caused by mammalian cells and is referred to as Electrical Cell-surface Impedance Sensing or ECIS for short. The characteristic behavior of these three systems will be contrasted and discussed in detail. In particular we will focus on how the oscillations can be induced, how long they persists, and finally the stability of the observed frequencies.

  20. Clostridium perfringens epsilon toxin is cytotoxic for human renal tubular epithelial cells.

    PubMed

    Fernandez Miyakawa, Mariano E; Zabal, Osvaldo; Silberstein, Claudia

    2011-04-01

    Clostridium perfringens epsilon toxin (ETX) is responsible for a fatal enterotoxemia in different animal species, producing extensive renal damage, neurological disturbance and edema of lungs, heart and kidneys. However, there is no information about the susceptibility of humans to ETX. Here, we report that primary cultures of human renal tubular epithelial cells (HRTEC) exposed to ETX showed a marked swelling with subsequent large blebs surrounding most cells. The incubation of HRTEC with ETX produced a reduction of cell viability in a dose- and time-dependent manner. The CD(50) after 1-hour and 24-hour incubation were 3 µg/mL and 0.5 µg/mL, respectively. The pulse with ETX for 3 min was enough to produce a significant cytotoxic effect on HRTEC after 1-hour incubation. ETX binds to HRTEC forming a large complex of about 160 kDa similar to what was found in the Madin-Darby canine kidney (MDCK) cell line. The HRTEC could be a useful cell model to improve the understanding of the mechanisms involved on the cell damage mediated by ETX. PMID:20488848

  1. Interaction of Clostridium perfringens epsilon-toxin with biological and model membranes: A putative protein receptor in cells.

    PubMed

    Manni, Marco M; Sot, Jesús; Goñi, Félix M

    2015-03-01

    Epsilon-toxin (ETX) is a powerful toxin produced by some strains of Clostridium perfringens (classified as types B and D) that is responsible for enterotoxemia in animals. ETX forms pores through the plasma membrane of eukaryotic cells, consisting of a β-barrel of 14 amphipathic β-strands. ETX shows a high specificity for certain cell lines, of which Madin-Darby canine kidney (MDCK) is the first sensitive cell line identified and the most studied one. The aim of this study was to establish the role of lipids in the toxicity caused by ETX and the correlation of its activity in model and biological membranes. In MDCK cells, using cell counting and confocal microscopy, we have observed that the toxin causes cell death mediated by toxin binding to plasma membrane. Moreover, ETX binds and permeabilizes the membranes of giant plasma membrane vesicles (GPMV). However, little effect is observed on protein-free vesicles. The data suggest the essential role of a protein receptor for the toxin in cell membranes. PMID:25485476

  2. Is Trichomonas tenax a Parasite or a Commensal?

    PubMed

    Ribeiro, Luiz Carlos; Santos, Carlos; Benchimol, Marlene

    2015-05-01

    Trichomonas tenax is considered a commensal organism found under poor oral hygiene conditions. T. tenax presents morphological similarities with T. vaginalis, and there are doubts concerning whether this protist is a parasite and whether it is a genetic variant of T. vaginalis. This study aimed to investigate the capacity of T. tenax to cause mammalian cell damage and compare its cytotoxicity with that of T. vaginalis. Protozoan-host cell interaction assays were performed with Madin-Darby canine kidney, HeLa, and gum cells and 3D spheroids, which were examined by scanning electron and transmission electron microscopy. Cellular viability experiments were also performed. T. tenax attached and had different forms when interacting with mammalian cells and caused damage with time-dependent host-cell viability. We observed that T. tenax produced plasma membrane projections and phagocytosed portions of the mammalian cells. In addition, T. tenax caused membrane blebbing and apoptotic bodies in HeLa cells, thus inducing cell death. Spheroids were also used in interaction assays with T. tenax and they were damaged by these cells. This study shows that T. tenax fulfills the requisites of a parasite, causing damage to different mammalian cells and behaving similarly to T. vaginalis when in contact with target cells in vitro. PMID:25835639

  3. Crawling Cells Can Close Wounds without Purse Strings or Signaling

    PubMed Central

    Lee, Pilhwa; Wolgemuth, Charles W.

    2011-01-01

    When a gash or gouge is made in a confluent layer of epithelial cells, the cells move to fill in the “wound.” In some cases, such as in wounded embryonic chick wing buds, the movement of the cells is driven by cortical actin contraction (i.e., a purse string mechanism). In adult tissue, though, cells apparently crawl to close wounds. At the single cell level, this crawling is driven by the dynamics of the cell's actin cytoskeleton, which is regulated by a complex biochemical network, and cell signaling has been proposed to play a significant role in directing cells to move into the denuded area. However, wounds made in monolayers of Madin-Darby canine kidney (MDCK) cells still close even when a row of cells is deactivated at the periphery of the wound, and recent experiments show complex, highly-correlated cellular motions that extend tens of cell lengths away from the boundary. These experiments suggest a dominant role for mechanics in wound healing. Here we present a biophysical description of the collective migration of epithelial cells during wound healing based on the basic motility of single cells and cell-cell interactions. This model quantitatively captures the dynamics of wound closure and reproduces the complex cellular flows that are observed. These results suggest that wound healing is predominantly a mechanical process that is modified, but not produced, by cell-cell signaling. PMID:21423710

  4. Designing cell lines for viral vaccine production: Where do we stand?

    PubMed

    Genzel, Yvonne

    2015-05-01

    Established animal cells, such as Vero, Madin Darby canine kidney (MDCK) or chicken embryo fibroblasts (CEFs), are still the main cell lines used for viral vaccine production, although new "designer cells" have been available for some years. These designer cell lines were specifically developed as a cell substrate for one application and are well characterized. Later screening for other possible applications widened the product range. These cells grow in suspension in chemically defined media under controlled conditions and can be used for up to 100 passages. Scale-up is easier and current process options allow cultivation in disposable bioreactors at cell concentrations higher than 1 × 10(7) cells/mL. This review covers the limitations of established cell lines and discusses the requirements and screening options for new host cells. Currently available designer cells for viral vaccine production (PER.C6, CAP, AGE1.CR, EB66 cells), together with other new cell lines (PBS-1, QOR/2E11, SogE, MFF-8C1 cells) that were recently described as possible cell substrates are presented. Using current process knowledge and cell line development tools, future upstream processing could resemble today's Chinese hamster ovary (CHO) cell processes for monoclonal antibody production: small scale bioreactors (disposable) in perfusion or fed-batch mode with cell concentrations above 1 × 10(8) cells/mL. PMID:25903999

  5. Characterization of Neuraminidase Inhibitors in Korean Papaver rhoeas Bee Pollen Contributing to Anti-Influenza Activities In Vitro.

    PubMed

    Lee, In-Kyoung; Hwang, Byung Soon; Kim, Dae-Won; Kim, Ji-Yul; Woo, E-Eum; Lee, Yoon-Ju; Choi, Hwa Jung; Yun, Bong-Sik

    2016-04-01

    The active constituents of Korean Papaver rhoeas bee pollen conferring neuraminidase inhibitory activities (H1N1, H3N2, and H5N1) were investigated. Six flavonoids and one alkaloid were isolated and characterized by nuclear magnetic resonance and mass spectrometry data. These included kaempferol-3-sophoroside (1), kaempferol-3-neohesperidoside (2), kaempferol-3-sambubioside (3), kaempferol-3-glucoside (4), quercetin-3-sophoroside (5), luteolin (6), and chelianthifoline (7). All compounds showed neuraminidase inhibitory activities with IC50 values ranging from 10.7 to 151.1 µM. The most potent neuraminidase inhibitor was luteolin, which was the dominant content in the ethyl acetate fraction. All tested compounds displayed noncompetitive inhibition of H3N2 neuraminidase. Furthermore, compounds 1-7 all reduced the severity of virally induced cytopathic effects as determined by the Madin-Darby canine kidney cell-based assay showing antiviral activity with IC50 values ranging from 10.7 to 33.4 µM (zanamivir: 58.3 µM). The active compounds were quantified by high-performance liquid chromatography, and the total amount of compounds 1-7 made up about 0.592 g/100 g bee pollen, contributing a rich resource of a natural antiviral product. PMID:26848705

  6. Transformed MDCK cells secrete elevated MMP1 that generates LAMA5 fragments promoting endothelial cell angiogenesis.

    PubMed

    Gopal, Shashi K; Greening, David W; Zhu, Hong-Jian; Simpson, Richard J; Mathias, Rommel A

    2016-01-01

    Epithelial-mesenchymal transition (EMT) enhances the migration and invasion of cancer cells, and is regulated by various molecular mechanisms including extracellular matrix metalloproteinase (MMP) activity. Previously, we reported transformation of epithelial Madin-Darby canine kidney (MDCK) cells with oncogenic H-Ras (21D1 cells) induces EMT, and significantly elevates MMP1 expression. To explore the biological significance, in this study we characterized 21D1 cells with knocked-down MMP1 expression (21D1(-MMP1)). MMP1 silencing diminished 21D1 cell migration, invasion and anchorage-independent growth in vitro. Additionally, 21D1(-MMP1) cells displayed reduced tumour volume when grown as in vivo subcutaneous xenografts in mice. Depletion of MMP1 lowered the ability of the cellular secretome (extracellular culture medium) to influence recipient cell behaviour. For example, supplementation with 21D1 secretome elevated cell migration of recipient fibroblasts, and enhanced endothelial cell angiogenesis (vessel length and branching). By contrast, 21D1(-MMP1) secretome was less potent in both functional assays. We reveal laminin subunit alpha-5 (LAMA5) as a novel biological substrate of MMP1, that generates internal and C-terminal proteolytic fragments in 21D1 secretome. Furthermore, antibody-based inhibition of integrin αvβ3 on endothelial cells nullified the angiogenic capability of 21D1 secretome. Therefore, we report this as a new VEGF-independent mechanism that oncogenic cells may employ to promote tumour angiogenesis. PMID:27324842

  7. Heterogeneous transport of digitalis-like compounds by P-glycoprotein in vesicular and cellular assays.

    PubMed

    Gozalpour, Elnaz; Wilmer, Martijn J; Bilos, Albert; Masereeuw, Rosalinde; Russel, Frans G M; Koenderink, Jan B

    2016-04-01

    Digitalis-like compounds (DLCs), the ancient medication of heart failure and Na,K-ATPase inhibitors, are characterized by their toxicity. Drug-drug interactions (DDIs) at absorption and excretion levels play a key role in their toxicity, hence, knowledge about the transporters involved might prevent these unwanted interactions. In the present study, the transport of fourteen DLCs with human P-glycoprotein (P-gp; ABCB1) was studied using a liquid chromatography-mass spectrometry (LC-MS) quantification method. DLC transport by P-gp overexpressing Madin-Darby canine kidney (MDCK) and immortalized human renal cells (ciPTEC) was compared to vesicular DLC transport. Previously, we identified convallatoxin as a substrate using membrane vesicles overexpressing P-gp; however, we could not measure transport of other DLCs in this assay (Gozalpour et al., 2014a). Here, we showed that lipophilic digitoxin, digoxigenin, strophanthidin and proscillaridin A are P-gp substrates in cellular accumulation assays, whereas the less lipophilic convallatoxin was not. P-gp function in the cellular accumulation assays depends on the entrance of lipophilic compounds by passive diffusion, whereas the vesicular transport assay is more appropriate for hydrophilic substrates. In conclusion, we identified digitoxin, digoxigenin, strophanthidin and proscillaridin A as P-gp substrates using cellular accumulation assays and recognized lipophilicity as an important factor in selecting a suitable transport assay. PMID:26708294

  8. Impedance spectroscopy for the detection and identification of unknown toxins

    NASA Astrophysics Data System (ADS)

    Riggs, B. C.; Plopper, G. E.; Paluh, J. L.; Phamduy, T. B.; Corr, D. T.; Chrisey, D. B.

    2012-06-01

    Advancements in biological and chemical warfare has allowed for the creation of novel toxins necessitating a universal, real-time sensor. We have used a function-based biosensor employing impedance spectroscopy using a low current density AC signal over a range of frequencies (62.5 Hz-64 kHz) to measure the electrical impedance of a confluent epithelial cell monolayer at 120 sec intervals. Madin Darby canine kidney (MDCK) epithelial cells were grown to confluence on thin film interdigitated gold electrodes. A stable impedance measurement of 2200 Ω was found after 24 hrs of growth. After exposure to cytotoxins anthrax lethal toxin and etoposide, the impedance decreased in a linear fashion resulting in a 50% drop in impedance over 50hrs showing significant difference from the control sample (~20% decrease). Immunofluorescent imaging showed that apoptosis was induced through the addition of toxins. Similarities of the impedance signal shows that the mechanism of cellular death was the same between ALT and etoposide. A revised equivalent circuit model was employed in order to quantify morphological changes in the cell monolayer such as tight junction integrity and cell surface area coverage. This model showed a faster response to cytotoxin (2 hrs) compared to raw measurements (20 hrs). We demonstrate that herein that impedance spectroscopy of epithelial monolayers serves as a real-time non-destructive sensor for unknown pathogens.

  9. Measurements and modeling of water transport and osmoregulation in a single kidney cell using optical tweezers and videomicroscopy

    NASA Astrophysics Data System (ADS)

    Lúcio, A. D.; Santos, R. A.; Mesquita, O. N.

    2003-10-01

    With an optical tweezer installed in our optical microscope we grab a single Madin Darby Canine kidney cell and keep it suspended in the medium without touching the glass substrate or other cells. Since the optically trapped cell remains with a closely round shape, we can directly measure its volume by using videomicroscopy with digital image analysis. We submit this cell to a hyperosmotic shock (up-shock) and video record the process: the cell initially shrinks due to osmotic efflux of water and after a while, due to regulatory volume increase (RVI), an osmoregulation response, it inflates again (water influx) until it reaches a new volume (the regulatory volume VR). In addition to considering standard osmotic water transport, we model RVI using a simple phenomenological model. We obtain an expression for cell volume variation as a function of time that fits very well with our experimental data, where two characteristic times appear naturally: one related to water transport and the other related to RVI. From the fit we obtain water permeability, osmolyte influx rate for RVI, and regulatory volume. With the addition of the hormone vasopressin, water permeability increases while the regulatory volume decreases until inhibition of RVI. In summary, we present a technique to measure directly volume changes of a single isolated kidney cell under osmotic shock and a phenomenological analysis of water transport that takes into account osmoregulation.

  10. Susceptibility of influenza viruses circulating in Western Saudi Arabia to neuraminidase inhibitors

    PubMed Central

    Tolah, Ahmed M.; Azhar, Esam I.; Hashem, Anwar M.

    2016-01-01

    Objectives: To investigate the sensitivity of circulating influenza viruses in Western Saudi Arabia to neuraminidase inhibitors (NAIs); mainly, zanamivir and oseltamivir. Methods: Respiratory samples were collected from patients presenting with respiratory symptoms to King Abdulaziz University Hospital, Jeddah, Kingdom of Saudi Arabia (KSA) between September 2013 and October 2014. All samples were tested prospectively by real-time reverse-transcription polymerase chain reaction for influenza A and B viruses. Positive samples were then inoculated on Madin-Darby Canine Kidney (MDCK) cells and isolated viruses were examined for their sensitivity to NAIs using fluorescent neuraminidase inhibition assay. Results: Out of 406 tested samples, 25 samples (6.2%) were positive for influenza A/pdmH1N1 virus, one sample (0.25%) was positive for influenza A/H3N2 virus, and 7 samples (1.7%) were positive for influenza B Yamagata-like virus. Screening of isolated influenza A and B viruses (9 out of 33) for their sensitivity to NAIs showed no significant resistance to available NAIs. Conclusion: Our results show that circulating influenza viruses in Jeddah are still sensitive to NAIs. PMID:27052292

  11. Establishment of MDCK Stable Cell Lines Expressing TMPRSS2 and MSPL and Their Applications in Propagating Influenza Vaccine Viruses in Absence of Exogenous Trypsin

    PubMed Central

    Wen, Zhiyuan; Wu, Chao; Chen, Weiye; Zeng, Xianying; Shi, Jianzhong; Ge, Jinying; Chen, Hualan; Bu, Zhigao

    2015-01-01

    We established two Madin-Darby canine kidney (MDCK) cell lines stably expressing human airway transmembrane protease: transmembrane protease, serine 2 (TMPRSS2) and mosaic serine protease large form (MSPL) which support multicycle growth of two H5 highly pathogenic avian influenza viruses (HPAIV) recombinant vaccines (Re-5 and Re-6) and an H9 avian influenza virus (AIV) recombinant vaccine (Re-9) in the absence of trypsin. Data showed that the cell lines stably expressed TMPRSS2 and MSPL after 20 serial passages. Both MDCK-TMPRSS2 and MDCK-MSPL could proteolytically cleave the HA of Re-5, Re-6, and Re-9 and supported high-titer growth of the vaccine without exogenous trypsin. Re-5, Re-6, and Re-9 efficiently infected and replicated within MDCK-TMPRSS2 and MDCK-MSPL cells and viral titer were comparable to the virus grown in MDCK cells with TPCK-trypsin. Thus, our results indicate a potential application for these cell lines in cell-based influenza vaccine production and may serve as a useful tool for HA proteolytic cleavage-related studies. PMID:25918647

  12. Regulation of Myosin II Dynamics by Phosphorylation and Dephosphorylation of Its Light Chain in Epithelial Cells

    PubMed Central

    Watanabe, Toshiyuki; Hosoya, Hiroshi

    2007-01-01

    Nonmuscle myosin II, an actin-based motor protein, plays an essential role in actin cytoskeleton organization and cellular motility. Although phosphorylation of its regulatory light chain (MRLC) is known to be involved in myosin II filament assembly and motor activity in vitro, it remains unclear exactly how MRLC phosphorylation regulates myosin II dynamics in vivo. We established clones of Madin Darby canine kidney II epithelial cells expressing MRLC-enhanced green fluorescent protein or its mutants. Time-lapse imaging revealed that both phosphorylation and dephosphorylation are required for proper dynamics of myosin II. Inhibitors affecting myosin phosphorylation and MRLC mutants indicated that monophosphorylation of MRLC is required and sufficient for maintenance of stress fibers. Diphosphorylated MRLC stabilized myosin II filaments and was distributed locally in regions of stress fibers where contraction occurs, suggesting that diphosphorylation is involved in the spatial regulation of myosin II assembly and contraction. We further found that myosin phosphatase or Zipper-interacting protein kinase localizes to stress fibers depending on the activity of myosin II ATPase. PMID:17151359

  13. Reduction of high-energy shock-wave-induced renal tubular injury by selenium.

    PubMed

    Strohmaier, W L; Lahme, S; Weidenbach, P M; Bichler, K H

    1999-10-01

    In shock-wave-induced renal injury cavitation-generated free radicals play an important role. Using an in vitro model with Madin-Darby canine kidney (MDCK) cells, we investigated the influence of selenium, a free radical scavenger, in shock-wave-induced tubular cell injury. Suspensions of MDCK cells (33 x 10(6) cells/ml) were placed in small containers (volume 1.1 ml) for shock wave exposure. Two groups of 12 containers each were examined: (1) control (no medication), (2) selenium (0.4 microg/ml nutrient medium). Six containers in each group were exposed to shock waves (impulse rate 256, frequency 60 Hz, generator voltage 18 kV), while the other six containers in each group served as a control. After shock wave exposure, the concentration of cellular enzymes such as lactate dehydrogenase (LDH), N-acetyl-beta-glucosaminidase (NAG), glutamate oxaloacetate transaminase (GOT) and glutamate lactate dehydrogenase (GLDH) in the nutrient medium was examined. Following shock wave exposure there was a significant rise in LDH, NAG, GOT and GLDH concentrations. Selenium reduced this enzyme leakage significantly. Thus we conclude that selenium protects renal tubular cells against shock-wave-induced injury. Since selenium is an essential part of glutathione peroxidase, this effect seems to be mediated by a reduction in reactive oxygen species. PMID:10550528

  14. A Novel I221L Substitution in Neuraminidase Confers High-Level Resistance to Oseltamivir in Influenza B Viruses

    PubMed Central

    Escuret, Vanessa; Collins, Patrick J.; Casalegno, Jean-Sébastien; Vachieri, Sebastien G.; Cattle, Nicholas; Ferraris, Olivier; Sabatier, Murielle; Frobert, Emilie; Caro, Valérie; Skehel, John J.; Gamblin, Steve; Valla, Frédéric; Valette, Martine; Ottmann, Michèle; McCauley, John W.; Daniels, Rodney S.; Lina, Bruno

    2014-01-01

    Influenza B viruses with a novel I221L substitution in neuraminidase (NA) conferring high-level resistance to oseltamivir were isolated from an immunocompromised patient after prolonged oseltamivir treatment. Methods. Enzymatic characterization of the NAs (Km, Ki) and the in vitro fitness of viruses carrying wild-type or mutated (I221L) NA genes were evaluated. Proportions of wild-type and mutated NA genes were directly quantified in the patient samples. Structural characterizations by X-ray crystallography of a wild-type and I221L variant NA were performed. Results. The Km and Ki revealed that the I221L variant NA had approximately 84 and 51 times lower affinity for oseltamivir carboxylate and zanamivir, respectively, compared with wild-type NA. Viruses with a wild-type or I221L variant NA had similar growth kinetics in Madin-Darby canine kidney (MDCK) cells, and 5 passages in MDCK cells revealed no reversion of the I221L substitution. The crystal structure of the I221L NA and oseltamivir complex showed that the leucine side chain protrudes into the hydrophobic pocket of the active site that accommodates the pentyloxy substituent of oseltamivir. Conclusions. Enzyme kinetic and NA structural analyses provide an explanation for the high level of resistance to oseltamivir while retaining good fitness of viruses carrying I221L variant NA. PMID:24795482

  15. New Findings on the Mechanism of Perspiration Including Aquaporin-5 Water Channel.

    PubMed

    Inoue, Risako

    2016-01-01

    Aquaporin-5 (AQP5) is a member of the water channel protein family. Although AQP5 has been shown to be present in sweat glands, the presence or absence of regulated intracellular translocation of AQP5 in sweat glands remains to be determined. In this article, recent findings on AQP5 in sweat glands are presented. (1) Immunoreactive AQP5 was detected in the apical membranes and the intercellular canaliculi of secretory coils, and in the basolateral membranes of the clear cells in human eccrine sweat glands. (2) AQP5 rapidly concentrated at the apical membranes during sweating in mouse sweat glands. (3) Treatment of human AQP5-expressing Madin-Darby canine kidney cells with calcium ionophore A23187 resulted in a twofold increase in the AQP5 level in the apical membranes within 5 min. (4) Anoctamin-1, a calcium-activated chloride channel was detected in the apical membranes and it completely colocalized with AQP5 in the apical membranes in mouse sweat glands. AQP5 may be involved in sweating and its translocation may help to increase the water permeability of the apical membranes of sweat glands. AQP5 is a potential target molecule for the design of a sweat-modulating drug. PMID:27584958

  16. Host Adaptation and the Alteration of Viral Properties of the First Influenza A/H1N1pdm09 Virus Isolated in Japan

    PubMed Central

    Ainai, Akira; Hasegawa, Hideki; Obuchi, Masatsugu; Odagiri, Takato; Ujike, Makoto; Shirakura, Masayuki; Nobusawa, Eri; Tashiro, Masato; Asanuma, Hideki

    2015-01-01

    A/Narita/1/2009 (A/N) was the first H1N1 virus from the 2009 pandemic (H1pdm) to be isolated in Japan. To better understand and predict the possible development of this virus strain, the effect of passaging A/N was investigated in Madin-Darby canine kidney cells, chicken eggs and mice. A/N that had been continuously passaged in cells, eggs, or mice obtained the ability to grow efficiently in each host. Moreover, A/N grown in mice had both a high level of pathogenicity in mice and an increased growth rate in cells and eggs. Changes in growth and pathogenicity were accompanied by amino acid substitutions in viral hemagglutinin (HA) and PB2. In addition, the adapted viruses exhibited a reduced ability to react with ferret antisera against A/N. In conclusion, prolonged passaging allowed influenza A/N to adapt to different hosts, as indicated by a high increase in proliferative capacity that was accompanied by an antigenic alteration leading to amino acid substitutions. PMID:26079133

  17. Meprin A impairs epithelial barrier function, enhances monocyte migration, and cleaves the tight junction protein occludin

    PubMed Central

    Bao, Jialing; Yura, Renee E.; Matters, Gail L.; Bradley, S. Gaylen; Shi, Pan; Tian, Fang

    2013-01-01

    Meprin metalloproteases are highly expressed at the luminal interface of the intestine and kidney and in certain leukocytes. Meprins cleave a variety of substrates in vitro, including extracellular matrix proteins, adherens junction proteins, and cytokines, and have been implicated in a number of inflammatory diseases. The linkage between results in vitro and pathogenesis, however, has not been elucidated. The present study aimed to determine whether meprins are determinative factors in disrupting the barrier function of the epithelium. Active meprin A or meprin B applied to Madin-Darby canine kidney (MDCK) cell monolayers increased permeability to fluorescein isothiocyanate-dextran and disrupted immunostaining of the tight junction protein occludin but not claudin-4. Meprin A, but not meprin B, cleaved occludin in MDCK monolayers. Experiments with recombinant occludin demonstrated that meprin A cleaves the protein between Gly100 and Ser101 on the first extracellular loop. In vivo experiments demonstrated that meprin A infused into the mouse bladder increased the epithelium permeability to sodium fluorescein. Furthermore, monocytes from meprin knockout mice on a C57BL/6 background were less able to migrate through an MDCK monolayer than monocytes from their wild-type counterparts. These results demonstrate the capability of meprin A to disrupt epithelial barriers and implicate occludin as one of the important targets of meprin A that may modulate inflammation. PMID:23804454

  18. Meprin Metalloproteases Inactivate Interleukin 6*

    PubMed Central

    Keiffer, Timothy R.; Bond, Judith S.

    2014-01-01

    Meprins have been implicated in the pathogenesis of several inflammatory diseases, including inflammatory bowel disease, in which the cytokine IL-6 is a prominent effector molecule. Because IL-6 levels are elevated markedly in meprin α and α/β knockout mice in an experimental model of inflammatory bowel disease, the interaction between meprins and IL-6 was studied. The results demonstrate that rodent and human meprin A and B cleave IL-6 to a smaller product and, subsequently, are capable of extensive degradation of the cytokine. Analysis of the limited degradation product formed by meprin A indicated that three to five amino acids are removed from the C terminus of the cytokine. Meprin A and meprin B cleaved IL-6 with micromolar affinities (Km of 4.7 and 12.0 μm, respectively) and with high efficiencies (kcat/Km of 0.2 and 2.5 (m−1/s−1) × 106, respectively). These efficiency constants are among the highest for known meprin substrates. Madin-Darby canine kidney cells transiently transfected with meprin α or meprin β constructs also cleave exogenous IL-6. Both human and murine IL-6 cleaved by meprin A or B are inactivated, as demonstrated by their decreased capability to stimulate proliferation of B9 cells. These results are consistent with the proposition that one function of meprin metalloproteases is to modulate inflammation by inactivating IL-6. PMID:24474695

  19. An autocrine TGF-β/ZEB/miR-200 signaling network regulates establishment and maintenance of epithelial-mesenchymal transition

    PubMed Central

    Gregory, Philip A.; Bracken, Cameron P.; Smith, Eric; Bert, Andrew G.; Wright, Josephine A.; Roslan, Suraya; Morris, Melanie; Wyatt, Leila; Farshid, Gelareh; Lim, Yat-Yuen; Lindeman, Geoffrey J.; Shannon, M. Frances; Drew, Paul A.; Khew-Goodall, Yeesim; Goodall, Gregory J.

    2011-01-01

     Epithelial-mesenchymal transition (EMT) is a form of cellular plasticity that is critical for embryonic development and tumor metastasis. A double-negative feedback loop involving the miR-200 family and ZEB (zinc finger E-box-binding homeobox) transcription factors has been postulated to control the balance between epithelial and mesenchymal states. Here we demonstrate using the epithelial Madin Darby canine kidney cell line model that, although manipulation of the ZEB/miR-200 balance is able to repeatedly switch cells between epithelial and mesenchymal states, the induction and maintenance of a stable mesenchymal phenotype requires the establishment of autocrine transforming growth factor-β (TGF-β) signaling to drive sustained ZEB expression. Furthermore, we show that prolonged autocrine TGF-β signaling induced reversible DNA methylation of the miR-200 loci with corresponding changes in miR-200 levels. Collectively, these findings demonstrate the existence of an autocrine TGF-β/ZEB/miR-200 signaling network that regulates plasticity between epithelial and mesenchymal states. We find a strong correlation between ZEBs and TGF-β and negative correlations between miR-200 and TGF-β and between miR-200 and ZEBs, in invasive ductal carcinomas, consistent with an autocrine TGF-β/ZEB/miR-200 signaling network being active in breast cancers. PMID:21411626

  20. Lumen formation by epithelial cell lines in response to collagen overlay: a morphogenetic model in culture

    SciTech Connect

    Hall, H.G.; Farson, D.A.; Bissel, M.J.

    1982-08-01

    Two cells lines--Madin-Darby canine kidney (MDCK) and normal murine mammary gland (NMuMG)--growing as monolayers on collagen gels were overlaid with another collagen gel. The cells responded to the overlay by undergoing reorganization resulting in the creation of lumina. MDCK cells formed lumina that coalesced to form large cavities comparable in size with a tubule. NMuMG cells formed clusters surrounding small lumina, which appeared similar to acini of glandular tissue. The characteristic arrangements, described here by light and electron microscopy, resembled the morphology of the tissues of cell line origin. MDCK cells, grown in the presence of serum, formed lumina whether or not serum was removed at the time of overlay, whereas NMuMG cells required either a nondialyzable component of serum or hormonal supplements in serum-free defined media. Lumen formation was delayed by MDCK cells in the presence of the glutamine analog 6-diazo-5-oxo-L-norleucine, but this compound did not affect NMuMG lumen formation. In both cell lines, lumen formation was unaffected by the absence of sulfate, the presence of an inhibitor of sulfate glycosaminoglycan synthesis, or an inhibitor of collagen synthesis. DNA synthesis accompanied lumen formation but was not required.

  1. Bioisosteric approach in designing new monastrol derivatives: an investigation on their ADMET prediction using in silico derived parameters.

    PubMed

    Hassan, Syed Fahad; Rashid, Umer; Ansari, Farzana Latif; Ul-Haq, Zaheer

    2013-09-01

    Medicinal chemists are facing an increasing challenge to deliver safer and more effective medicines. An appropriate balance between drug-like properties such as solubility, permeability, metabolic stability, efficacy and toxicity is one of the most challenging problems during lead optimization of a potential drug candidate. Insoluble and impermeable compounds can result in erroneous biological data and unreliable SAR in enzyme and cell-based assays. The weak inhibitory activity and non-drug-like properties of monastrol, the first small mitotic kinesin Eg5 inhibitor, has hampered its further development. In this investigation, a bioisosteric approach was applied that resulted in the replacement of C-5 carbonyl of monastrol with thio-carbonyl. Further lead optimization of drug-like properties was evaluated through in silico predictions by using ADMET predictor software. This minor structural modification resulted in upgraded human effective jejunal permeability (Peff) and improved permeability in Madin-Darby canine kidney (MDCK) cells. Furthermore, C-5 thiocarbonyl analogue of monastrol (named as Special-2) was found safe to administer orally with no phospholipidosis toxicity, no raised levels of serum glutamate oxaloacetate transaminase (SGOT) and no potential towards cardiotoxicity. Molecular docking study was also carried out to understand the binding modes of these compounds. The docking study showed high binding affinity of the designed compounds against KSP. Hence a combination of in silico ADMET studies and molecular docking can help to improve prediction success and these compounds might be act as potential candidate for KSP inhibition. PMID:24080467

  2. Genomic structure, gene expression, and promoter analysis of human multidrug resistance-associated protein 7

    SciTech Connect

    Kao, Hsin-Hsin; Chang, Ming-Shi; Cheng, Jan-Fang; Huang, Jin-Ding

    2002-03-15

    The multidrug resistance-associated protein (MRP) subfamily transporters associated with anticancer drug efflux are attributed to the multidrug-resistance of cancer cells. The genomic organization of human multidrug resistance-associated protein 7 (MRP7) was identified. The human MRP7 gene, consisting of 22 exons and 21 introns, greatly differs from other members of the human MRP subfamily. A splicing variant of human MRP7, MRP7A, expressed in most human tissues, was also characterized. The 1.93-kb promoter region of MRP7 was isolated and shown to support luciferase activity at a level 4- to 5-fold greater than that of the SV40 promoter. Basal MRP7 gene expression was regulated by 2 regions in the 5-flanking region at 1,780 1,287 bp, and at 611 to 208 bp. In Madin-Darby canine kidney (MDCK) cells, MRP7 promoter activity was increased by 226 percent by genotoxic 2-acetylaminofluorene and 347 percent by the histone deacetylase inhibitor, trichostatin A. The protein was expressed in the membrane fraction of transfected MDCK cells.

  3. Temporal expression of transporters and receptors in a rat primary co-culture blood-brain barrier model.

    PubMed

    Liu, Houfu; Li, Yang; Lu, Sijie; Wu, Yiwen; Sahi, Jasminder

    2014-10-01

    1. The more relevant primary co-cultures of brain microvessel endothelial cells and astrocytes (BMEC) are less utilized for screening of potential CNS uptake when compared to intestinal and renal cell lines. 2. In this study, we characterized the temporal mRNA expression of major CNS transporters and receptors, including the transporter regulators Pxr, Ahr and Car in a rat BMEC co-cultured model. Permeability was compared with the Madin-Darby canine kidney (MDCKII)-MDR1 cell line and rat brain in situ perfusion model. 3. Our data demonstrated differential changes in expression of individual transporters and receptors over the culture period. Expression of ATP-binding cassette transporters was better retained than that of solute carrier transporters. The insulin receptor (IR) was best maintained among investigated receptors. AhR demonstrated high mRNA expression in rat brain capillaries and expression was better retained than Pxr or Car in culture. Mdr1b expression was up-regulated during primary culture, albeit Mdr1a mRNA levels were much higher. P-gp and Bcrp-1 were highly expressed and functional in this in vitro system. 4. Permeability measurements with 18 CNS marketed drugs demonstrated weak correlation between rBMEC model and rat in situ permeability and moderate correlation with MDCKII-MDR1 cells. 5. We have provided appropriate methodologies, as well as detailed and quantitative characterization data to facilitate improved understanding and rational use of this in vitro rat BBB model. PMID:24827375

  4. Grainyhead-like 2 inhibits the coactivator p300, suppressing tubulogenesis and the epithelial-mesenchymal transition.

    PubMed

    Pifer, Phillip M; Farris, Joshua C; Thomas, Alyssa L; Stoilov, Peter; Denvir, James; Smith, David M; Frisch, Steven M

    2016-08-01

    Developmental morphogenesis and tumor progression require a transient or stable breakdown of epithelial junctional complexes to permit programmed migration, invasion, and anoikis resistance, characteristics endowed by the epithelial-mesenchymal transition (EMT). The epithelial master-regulatory transcription factor Grainyhead-like 2 (GRHL2) suppresses and reverses EMT, causing a mesenchymal-epithelial transition to the default epithelial phenotype. Here we investigated the role of GRHL2 in tubulogenesis of Madin-Darby canine kidney cells, a process requiring transient, partial EMT. GRHL2 was required for cystogenesis, but it suppressed tubulogenesis in response to hepatocyte growth factor. Surprisingly, GRHL2 suppressed this process by inhibiting the histone acetyltransferase coactivator p300, preventing the induction of matrix metalloproteases and other p300-dependent genes required for tubulogenesis. A 13-amino acid region of GRHL2 was necessary for inhibition of p300, suppression of tubulogenesis, and interference with EMT. The results demonstrate that p300 is required for partial or complete EMT occurring in tubulogenesis or tumor progression and that GRHL2 suppresses EMT in both contexts through inhibition of p300. PMID:27251061

  5. Anoikis-resistant MDCK cells carrying susceptibilities to TNF-alpha and verotoxin that are suitable for influenza virus cultivation.

    PubMed

    Tsutsumi, Reiko; Fujisaki, Shigemi; Shozushima, Masanori; Saito, Koichi; Sato, Shigehiro

    2006-10-01

    Madin-Darby canine kidney (MDCK) cells were originally anchorage-dependent epithelial cells. Here, we have isolated a novel MDCK-derived cell population, termed 6 M-4, by means of culturing MDCK cells in suspension for nearly 6 months in the presence of Streptomyces griseus metalloendopeptidase (MEP). The isolated cells showed unique proliferation characteristics, which differed from parental MDCK cells. They proliferated adherently on a polystyrene matrix, but proliferated non-adherently both in the presence of MEP and on a non-adhesive matrix coated with poly 2-methacryloyloxyethyl phosphorylcholine (MPC). The 6 M-4 cells consisted of at least two cell types. One type, termed 6 M-4-TR7, would not grow in soft agar and showed a novel phenotype in that the cells were susceptible to both TNF-alpha and verotoxin 1 (VT1). In addition, the isolated adhesion-independent cells sustained epithelial traits of parental MDCK cells. We further show that these MDCK-derivative cells are suitable for influenza virus cultivation. Hemagglutination (HA) titers of influenzaviruses A and B were increased in the suspension culture of 6 M-4-TR7 cells supplemented with the MEP in comparison to adherently growing cells in the presence of trypsin. PMID:19002866

  6. Sphingomyelin metabolism is involved in the differentiation of MDCK cells induced by environmental hypertonicity

    PubMed Central

    Favale, Nicolás Octavio; Santacreu, Bruno Jaime; Pescio, Lucila Gisele; Marquez, Maria Gabriela; Sterin-Speziale, Norma Beatriz

    2015-01-01

    Sphingolipids (SLs) are relevant lipid components of eukaryotic cells. Besides regulating various cellular processes, SLs provide the structural framework for plasma membrane organization. Particularly, SM is associated with detergent-resistant microdomains. We have previously shown that the adherens junction (AJ) complex, the relevant cell-cell adhesion structure involved in cell differentiation and tissue organization, is located in an SM-rich membrane lipid domain. We have also demonstrated that under hypertonic conditions, Madin-Darby canine kidney (MDCK) cells acquire a differentiated phenotype with changes in SL metabolism. For these reasons, we decided to evaluate whether SM metabolism is involved in the acquisition of the differentiated phenotype of MDCK cells. We found that SM synthesis mediated by SM synthase 1 is involved in hypertonicity-induced formation of mature AJs, necessary for correct epithelial cell differentiation. Inhibition of SM synthesis impaired the acquisition of mature AJs, evoking a disintegration-like process reflected by the dissipation of E-cadherin and β- and α-catenins from the AJ complex. As a consequence, MDCK cells did not develop the hypertonicity-induced differentiated epithelial cell phenotype. PMID:25670801

  7. Chirally Pure Prodrugs and Their Converting Enzymes Lead to High Supersaturation and Rapid Transcellular Permeation of Benzodiazepines.

    PubMed

    Kapoor, Mamta; Cheryala, Narsihmulu; Rautiola, Davin; Georg, Gunda I; Cloyd, James C; Siegel, Ronald A

    2016-08-01

    Water-soluble prodrugs can be rapidly converted by enzymes to hydrophobic drugs, whose aqueous thermodynamic solubilities are low, but are maintained in aqueous solution at supersaturated concentrations due to slow precipitation kinetics. Recently, we investigated avizafone (AVF) in combination with Aspergillus oryzae protease as a prodrug/enzyme system intended to produce supersaturated diazepam (DZP). Several fold enhancement of permeation of supersaturated DZP across Madin-Darby canine kidney II-wild type (MDCKII-wt) monolayers was observed, compared to saturated DZP solutions. However, prodrug conversion was incomplete, putatively due to partial racemization of AVF and stereoselectivity of A oryzae protease. Here we report synthesis of chirally pure AVF, and demonstrate complete conversion to supersaturated DZP followed by complete DZP permeation at enhanced rates across MDCKII-wt cell monolayers. We also synthesized, for the first time, a chirally pure prodrug of midazolam (MDZ-pro) and carried out the same sequence of studies. A oryzae protease was identified as a benign and efficient activating enzyme for MDZ-pro. The MDZ-pro/A oryzae protease system showed greater than 25-fold increase in absorption rate of MDZ across MDCKII-wt monolayers, compared to saturated MDZ. Such chirally pure prodrug/enzyme systems are promising candidates for efficient intranasal delivery of benzodiazepine drugs used in the treatment of seizure emergencies. PMID:27342435

  8. Preparation, characterization and pharmacokinetics of fluorescence labeled propylene glycol alginate sodium sulfate

    NASA Astrophysics Data System (ADS)

    Li, Pengli; Li, Chunxia; Xue, Yiting; Zhang, Yang; Liu, Hongbing; Zhao, Xia; Yu, Guangli; Guan, Huashi

    2014-08-01

    A rapid and sensitive fluorescence labeling method was developed and validated for the microanalysis of a sulfated polysaccharide drug,namely propylene glycol alginate sodium sulfate (PSS), in rat plasma. Fluorescein isothiocyanate (FITC) was selected to label PSS, and 1, 6-diaminohexane was used to link PSS and FITC in order to prepare FITC-labeled PSS (F-PSS) through a reductive amination reaction. F-PSS was identified by UV-Vis, FT-IR and 1H-NMR spectrum. The cell stability and cytotoxicity of F-PSS were tested in Madin-Darby canine kidney (MDCK) cells. The results indicated that the labeling efficiency of F-PSS was 0.522% ± 0.0248% and the absolute bioavailability was 8.39%. F-PSS was stable in MDCK cells without obvious cytotoxicity. The method was sensitive and reliable; it showed a good linearity, precision, recovery and stability. The FITC labeling method can be applied to investigating the absorption and metabolism of PSS and other polysaccharides in biological samples.

  9. Prophylactic effect of herbal-marine compound (HESA-A) on influenza A virus infectivity

    PubMed Central

    2014-01-01

    Background Influenza virus is still a severe respiratory disease affecting human and other species. As conventional drugs are not recommended for long time because of side effects and drug resistance occurrence, traditional medication has been focused as alternative remedy. HESA-A is a natural compound from herbal-marine origin. Previous studies have reported the therapeutic properties of HESA-A on psoriasis vulgaris and different types of cancers and we also showed its anti-inflammatory effects against influenza A infection. Methods This study was designed to investigate the potential properties of HESA-A as prophylaxis or treatment. To investigate the prophylaxis or treatment activities of HESA-A, Madin-Darby Canine Kidney (MDCK) cells were exposed to HESA-A and influenza A virus in different manners of exposure and different time intervals. The results were evaluated by MTT and HA assays. Results It was found that HESA-A is much more effective against influenza cytopathic effects when it is applied for prophylaxis and also in concurrent treatment (p ≤ 0.05) but not in post-infection treatment (p ≥ 0.05). Conclusion In conclusion, HESA-A is significantly effective against influenza replication in prophylaxis application affecting the virus penetration/adsorption to the cell without any toxic effect on the cell viability. PMID:24708698

  10. Contact inhibition of locomotion determines cell-cell and cell-substrate forces in tissues.

    PubMed

    Zimmermann, Juliane; Camley, Brian A; Rappel, Wouter-Jan; Levine, Herbert

    2016-03-01

    Cells organized in tissues exert forces on their neighbors and their environment. Those cellular forces determine tissue homeostasis as well as reorganization during embryonic development and wound healing. To understand how cellular forces are generated and how they can influence the tissue state, we develop a particle-based simulation model for adhesive cell clusters and monolayers. Cells are contractile, exert forces on their substrate and on each other, and interact through contact inhibition of locomotion (CIL), meaning that cell-cell contacts suppress force transduction to the substrate and propulsion forces align away from neighbors. Our model captures the traction force patterns of small clusters of nonmotile cells and larger sheets of motile Madin-Darby canine kidney (MDCK) cells. In agreement with observations in a spreading MDCK colony, the cell density in the center increases as cells divide and the tissue grows. A feedback between cell density, CIL, and cell-cell adhesion gives rise to a linear relationship between cell density and intercellular tensile stress and forces the tissue into a nonmotile state characterized by a broad distribution of traction forces. Our model also captures the experimentally observed tissue flow around circular obstacles, and CIL accounts for traction forces at the edge. PMID:26903658

  11. Functional Characterization of Sodium-dependent Multivitamin Transporter (SMVT) in MDCK-MDR1 cells and its Utilization as a Target for Drug Delivery

    PubMed Central

    Luo, Shuanghui; Kansara, Viral S.; Zhu, Xiaodong; Pal, Dhananjay; Mitra, Ashim. K.

    2008-01-01

    The objective of this research is to characterize a sodium-dependent multivitamin transporter (SMVT) in MDCK-MDR1 cells (Madin-Darby canine kidney cells transfected with the human MDR1 gene) and to investigate the feasibility of utilizing MDCK-MDR1 cell line as an in vitro model to study the permeability of biotin-conjugated prodrugs of anti-HIV protease inhibitors. Mechanism of [3H] biotin uptake and transport was delineated. Transepithelial permeability of the biotin conjugated prodrug i.e. biotin-saquinavir was also studied. Reverse transcription-polymerase chain reaction (RT-PCR) was carried out to confirm the existence of SMVT in MDCK-MDR1 cells. Biotin uptake was Na+, pH, and temperature dependent, but energyindependent. Transepithelial transport studies of biotin-saquinavir in MDCK-MDR1, wild type MDCK, and Caco-2 cells revealed that permeability of biotin-saquinavir was similar in all three cell lines. A band of SMVT mRNA at 862 bp was identified by RT-PCR. A sodium-dependent multivitamin transporter, SMVT, responsible for biotin uptake and transport, was identified and functionally characterized in MDCK-MDR1 cells. Therefore, MDCK-MDR1 cell line may be utilized as an in vitro model to study the permeability of biotin conjugated prodrugs such as HIV protease inhibitors. PMID:16749865

  12. Fusion-Related Host Proteins Are Actively Regulated by NA during Influenza Infection as Revealed by Quantitative Proteomics Analysis

    PubMed Central

    Sui, Zhiwei; Wen, Bo; Gao, Zhimin; Chen, Quanjiao

    2014-01-01

    Three recombinant influenza A viruses with different neuraminidases (NAs) in the background of A/PR/8/34 (PR8), named rPR8-H5N1NA, rPR8-H9N2NA, and rPR8-H1N1NA, derived from H5N1, H9N2, H1N1 (swine) viruses, respectively, were constructed. We performed a quantitative proteomics analysis to investigate differential protein expression in Madin-Darby canine kidney (MDCK) cells infected with recombinant and wild-type influenza viruses to determine whether NA replacement would alter host cell gene expression. Using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-TOF MS) and two-dimensional gel electrophoresis (2-DE), we identified 12 up-regulated and 49 down-regulated protein spots, including cytoskeletal proteins, molecular biosynthesis proteins, ubiquitin-proteasome pathway proteins, and heat shock proteins. The most significant changes in infected cells were observed for molecular biosynthesis proteins. We found more differentially expressed protein spots in cells infected with rPR8-H5N1NA or rPR8-H9N2NA viruses than cells infected with wild-type virus. Many of those proteins are postulated to be involved in cell-cell fusion, but the full mechanism remains to be explored. Meanwhile, our data demonstrate that the wild-type virus has evolutionary advantages over recombinant viruses. PMID:25153908

  13. Human RON receptor tyrosine kinase induces complete epithelial-to-mesenchymal transition but causes cellular senescence

    SciTech Connect

    Cote, Marceline; Miller, A. Dusty; Liu, Shan-Lu . E-mail: shan-lu.liu@mcgill.ca

    2007-08-17

    The RON receptor tyrosine kinase is a member of the MET proto-oncogene family and is important for cell proliferation, differentiation, and cancer development. Here, we created a series of Madin-Darby canine kidney (MDCK) epithelial cell clones that express different levels of RON, and have investigated their biological properties. While low levels of RON correlated with little morphological change in MDCK cells, high levels of RON expression constitutively led to morphological scattering or complete and stabilized epithelial-to-mesenchymal transition (EMT). Unexpectedly, MDCK clones expressing higher levels of RON exhibited retarded proliferation and senescence, despite increased motility and invasiveness. RON was constitutively tyrosine-phosphorylated in MDCK cells expressing high levels of RON and undergoing EMT, and the MAPK signaling pathway was activated. This study reveals for the first time that RON alone is sufficient to induce complete and stabilized EMT in MDCK cells, and overexpression of RON does not cause cell transformation but rather induces cell cycle arrest and senescence, leading to impaired cell proliferation.

  14. Amiloride interacts with renal. cap alpha. - and. beta. -adrenergic receptors

    SciTech Connect

    Howard, M.J.; Mullen, M.D.; Insel, P.A.

    1987-07-01

    The authors have used radioligand binding techniques to assess whether amiloride and certain analogues of amiloride (ethylisopropyl amiloride and benzamil) can bind to adrenergic receptors in the kidney. They found that amiloride could compete for (/sup 3/H)rauwolscine (..cap alpha../sub 2/-adrenergic receptors), (/sup 3/H)prazosin (..cap alpha../sub 1/-adrenergic receptors), and (/sup 125/I)iodocyanopindolol (..beta..-adrenergic receptors) binding in rat renal cortical membranes with inhibitor constants of 13.6 /plus minus/ 5.7, 24.4 /plus minus/ 7.4, and 8.36 /plus minus/ 13.5 ..mu..M, respectively. Ethylisopropyl amiloride and benzamil were from 2- to 25-fold more potent than amiloride in competing for radioligand binding sites in studies with these membranes. In addition, amiloride and the two analogues competed for (/sup 3/H)prazosin sites on intact Madin-Darby canine kidney cells and amiloride blocked epinephrine-stimulated prostaglandin E/sub 2/ production in these cells. They conclude that amiloride competes for binding to several classes of renal adrenergic receptors with a rank order of potency of ..cap alpha../sub 2/ > ..cap alpha../sub 1/ > ..beta... Binding to, and antagonism of, adrenergic receptors occurs at concentrations of amiloride that are lower than previously observed nonspecific interactions of this agent.

  15. Non-Brownian dynamics and strategy of amoeboid cell locomotion

    NASA Astrophysics Data System (ADS)

    Nishimura, Shin I.; Ueda, Masahiro; Sasai, Masaki

    2012-04-01

    Amoeboid cells such as Dictyostelium discoideum and Madin-Darby canine kidney cells show the non-Brownian dynamics of migration characterized by the superdiffusive increase of mean-squared displacement. In order to elucidate the physical mechanism of this non-Brownian dynamics, a computational model is developed which highlights a group of inhibitory molecules for actin polymerization. Based on this model, we propose a hypothesis that inhibitory molecules are sent backward in the moving cell to accumulate at the rear of cell. The accumulated inhibitory molecules at the rear further promote cell locomotion to form a slow positive feedback loop of the whole-cell scale. The persistent straightforward migration is stabilized with this feedback mechanism, but the fluctuation in the distribution of inhibitory molecules and the cell shape deformation concurrently interrupt the persistent motion to turn the cell into a new direction. A sequence of switching behaviors between persistent motions and random turns gives rise to the superdiffusive migration in the absence of the external guidance signal. In the complex environment with obstacles, this combined process of persistent motions and random turns drives the simulated amoebae to solve the maze problem in a highly efficient way, which suggests the biological advantage for cells to bear the non-Brownian dynamics.

  16. Investigation of Enantioselective Membrane Permeability of α-Lipoic Acid in Caco-2 and MDCKII Cell.

    PubMed

    Uchida, Ryota; Okamoto, Hinako; Ikuta, Naoko; Terao, Keiji; Hirota, Takashi

    2016-01-01

    α-Lipoic acid (LA) contains a chiral carbon and exists as two enantiomers (R-α-lipoic acid (RLA) and S-α-lipoic acid (SLA)). We previously demonstrated that oral bioavailability of RLA is better than that of SLA. This difference arose from the fraction absorbed multiplied by gastrointestinal availability (F(a) × F(g)) and hepatic availability (F(h)) in the absorption phase. However, it remains unclear whether F(a) and/or F(g) are involved in enantioselectivity. In this study, Caco-2 cells and Madin-Darby canine kidney strain II cells were used to assess the enantioselectivity of membrane permeability. LA was actively transported from the apical side to basal side, regardless of the differences in its steric structure. Permeability rates were proportionally increased in the range of 10-250 µg LA/mL, and the permeability coefficient did not differ significantly between enantiomers. Hence, we conclude that enantioselective pharmacokinetics arose from the metabolism (F(h) or F(g) × F(h)), and definitely not from the membrane permeation (F(a)) in the absorption phase. PMID:26821014

  17. Feasibility Study of the Permeability and Uptake of Mesoporous Silica Nanoparticles across the Blood-Brain Barrier.

    PubMed

    Baghirov, Habib; Karaman, Didem; Viitala, Tapani; Duchanoy, Alain; Lou, Yan-Ru; Mamaeva, Veronika; Pryazhnikov, Evgeny; Khiroug, Leonard; de Lange Davies, Catharina; Sahlgren, Cecilia; Rosenholm, Jessica M

    2016-01-01

    Drug delivery into the brain is impeded by the blood-brain-barrier (BBB) that filters out the vast majority of drugs after systemic administration. In this work, we assessed the transport, uptake and cytotoxicity of promising drug nanocarriers, mesoporous silica nanoparticles (MSNs), in in vitro models of the BBB. RBE4 rat brain endothelial cells and Madin-Darby canine kidney epithelial cells, strain II, were used as BBB models. We studied spherical and rod-shaped MSNs with the following modifications: bare MSNs and MSNs coated with a poly(ethylene glycol)-poly(ethylene imine) (PEG-PEI) block copolymer. In transport studies, MSNs showed low permeability, whereas the results of the cellular uptake studies suggest robust uptake of PEG-PEI-coated MSNs. None of the MSNs showed significant toxic effects in the cell viability studies. While the shape effect was detectable but small, especially in the real-time surface plasmon resonance measurements, coating with PEG-PEI copolymers clearly facilitated the uptake of MSNs. Finally, we evaluated the in vivo detectability of one of the best candidates, i.e. the copolymer-coated rod-shaped MSNs, by two-photon in vivo imaging in the brain vasculature. The particles were clearly detectable after intravenous injection and caused no damage to the BBB. Thus, when properly designed, the uptake of MSNs could potentially be utilized for the delivery of drugs into the brain via transcellular transport. PMID:27547955

  18. Cadmium is more toxic to LLC-PK1 cells than to MDCK cells acting on the cadherin-catenin complex.

    PubMed

    Zimmerhackl, L B; Momm, F; Wiegele, G; Brandis, M

    1998-07-01

    Cadmium toxicity to renal cells was investigated in Madin-Darby canine kidney (MDCK) and LLC-PK1 cells as models of the distal tubule/collecting duct and proximal tubule, respectively. Cells were grown on two-compartment filters and exposed to 0.1-50 microM Cd2+. In MDCK cells, Cd2+ was more toxic from the basolateral than from the apical side and dependent on the extracellular Ca2+ concentration. Toxicity was evident within 24 h, as shown by a decrease in transepithelial resistance (TER), reduced proliferation (bromodeoxyuridine incorporation), reduction in ATP concentration, and morphological changes. On confocal microscopy, E-cadherin and alpha-catenin staining patterns indicated interference with the cadherin-catenin complex. LLC-PK1 cells showed a similar toxicity pattern, which was evident at lower Cd2+ concentrations. An increase of E-cadherin and alpha-catenin molecules in the Triton X-100-insoluble fraction was detectable at high Cd2+ concentrations in LLC-PK1 cells but not in MDCK cells. Lactate dehydrogenase release indicated membrane leakage in LLC-PK1 cells. Rhodamine-phalloidin staining, a probe for F-actin filaments, demonstrated alterations of the actin cytoskeleton in both cell lines. In conclusion, cadmium caused ATP depletion and interfered with the cadherin-catenin complex and probably the tight junctions changing renal cell morphology and function. PMID:9689016

  19. Transport Mechanism of Coumarin 6 Nanocrystals with Two Particle Sizes in MDCKII Monolayer and Larval Zebrafish.

    PubMed

    Miao, Xiaoqing; Li, Ye; Wang, Xueqing; Lee, Simon Ming-Yuen; Zheng, Ying

    2016-05-25

    Nanocrystals (NCs) were utilized as oral formulations in commercial products to deliver lipophilic drug, but their transport mechanisms are not fully understood. This study aimed to explore the transport mechanism of NCs using in vitro Madin-Darby canine kidney II (MDCK II) cells and in vivo larval zebrafish models. Coumarin 6 (C6) was formulated into NCs with particle size of 67.5 ± 5.2 and 190 ± 9.2 nm. In vitro studies showed that 70 nm NCs accumulated in lysosome and endoplasmic reticulum (ER) as destinations. Lipid raft pathways mediated the endocytosis, while lipid raft, ER/Golgi, and Golgi/plasma membrane pathways were involved in exocytosis and transcytosis process. However, 200 nm NCs accumulated more in a lysosome, where lipid raft pathways were also involved in the endocytosis process. In vivo studies in larval zebrafish model further confirmed that the above network plays an important role in the absorption and distribution of C6-NCs. PMID:27159431

  20. 9 CFR 113.316 - Canine Parainfluenza Vaccine.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... serum neutralization test using 50 to 300 TCID50 of canine parainfluenza virus. (2) A geometric mean... neutralization titers of 1:4 or greater; or, if there is not a significant reduction in virus isolation rate...

  1. 9 CFR 113.316 - Canine Parainfluenza Vaccine.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... serum neutralization test using 50 to 300 TCID50 of canine parainfluenza virus. (2) A geometric mean... neutralization titers of 1:4 or greater; or, if there is not a significant reduction in virus isolation rate...

  2. 9 CFR 113.316 - Canine Parainfluenza Vaccine.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... serum neutralization test using 50 to 300 TCID50 of canine parainfluenza virus. (2) A geometric mean... neutralization titers of 1:4 or greater; or, if there is not a significant reduction in virus isolation rate...

  3. 9 CFR 113.316 - Canine Parainfluenza Vaccine.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... serum neutralization test using 50 to 300 TCID50 of canine parainfluenza virus. (2) A geometric mean... neutralization titers of 1:4 or greater; or, if there is not a significant reduction in virus isolation rate...

  4. 9 CFR 113.316 - Canine Parainfluenza Vaccine.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... serum neutralization test using 50 to 300 TCID50 of canine parainfluenza virus. (2) A geometric mean... neutralization titers of 1:4 or greater; or, if there is not a significant reduction in virus isolation rate...

  5. Death of a wild wolf from canine parvovirus enteritis

    USGS Publications Warehouse

    Mech, L.D.; Kurtz, H.J.; Goyal, S.

    1997-01-01

    A 9-mo-old female wolf (Canis lupus) in the Superior National Forest of Minnesota (USA) died from a canine parvovirus (CPV) infection. This is the first direct evidence that this infection effects free-ranging wild wolves.

  6. Polar Landforms

    NASA Technical Reports Server (NTRS)

    2005-01-01

    10 August 2005 This Mars Global Surveyor (MGS) Mars Orbiter Camera (MOC) image shows eroded remnants of carbon dioxide ice in the south polar residual cap of Mars. The scarps that outline each small mesa have retreated about 3 meters (10 feet) per Mars year since MGS began orbiting the red planet in 1997.

    Location near: 87.0oS, 31.9oW Image width: width: 3 km (1.9 mi) Illumination from: upper left Season: Southern Spring

  7. Dento-Alveolar Distraction Osteogenesis for rapid Orthodontic Canine Retraction

    PubMed Central

    Kumar, Naveen; Prashantha, GS; Raikar, Sudhir; Ranganath, Krishnappa; Mathew, Silju; Nambiar, Sandeep

    2013-01-01

    Background: The objectives of this study were to evaluate the rate of canine distalization by segmental alveolar distraction method in first premolar extraction cases, to evaluate the displacement of the canine and first molar teeth, to assess the effects of the procedure on the pulpal vitality of the canines, and to determine the amount of root resorption in retracted canines. Materials & Methods: The sample of the study consisted of 20 teeth in 7 patients (five females and two males, mean age 18.5 years). After the osteotomy procedure distractor was fixed. After 3 days of consolidation period, the distractor was activated 3 quarter turns per day(0.75 mm/day) till the canines comes in contact with second premolar. An electrical vitality test was applied before and after the distraction procedure and during the follow-up period. Results: The mean distal retraction of canines was 7.262 ± 0.4864 mm. The distal displacement of the canine was mainly a combination of tipping and translation. The mean distraction procedure was completed in 14.60 ±1.536 days. The duration of retraction was less for mandibular canine compared to maxillary canine. The mean posterior anchorage loss was mean 0.50±0.688 mm. The amount of root resorption that occurred during distraction was clinically insignificant. None of the teeth reacted negatively to the electrical vitality test that was performed 6 months after the completion of the distraction procedure. There was no clinical sign of discoloration or pulpal pain in any tooth. Conclusion: With dentoalveolar distraction, as canines can be fully retracted in 12 to 16 days, the non-compliance patients, patients with root-shape malformations, periodontal problems, or ankylosed teeth will benefit from this technique. The anchorage teeth can withstand the retraction forces better with no anchorage loss, and without clinical or radiographic evidence of root resorption, ankylosis, periodontal problems, and soft tissue dehiscence. This

  8. Management of an Unusual Maxillary Canine: A Rare Entity

    PubMed Central

    Muppalla, Jaya Nagendra Krishna; Kavuda, Krishnamurthy; Punna, Rajani; Vanapatla, Amulya

    2015-01-01

    Clinicians need to have intimate knowledge and thorough understanding of both pulp chamber and root canal anatomy. They should be aware of possibility of anatomical variations in the root canal system during endodontic treatment. Maxillary canines usually have single root and root canal but rarely may have single root with two root canals. This case describes a lengthier maxillary canine with two root canals. PMID:26779354

  9. Nutraceuticals for canine liver disease: assessing the evidence.

    PubMed

    Vandeweerd, Jean-Michel; Cambier, Carole; Gustin, Pascal

    2013-09-01

    Nutraceuticals, or nutritional supplements, have been promoted for the ancillary treatment of liver disease in dogs. However, minimal information is available in the scientific literature about commonly used nutraceuticals, such as S-adenosylmethionine, silymarin, and vitamin E. No strong clinical evidence exists regarding the efficacy of these compounds as hepatoprotectants in canine liver disease. Until this evidence exists, individual veterinarians must assume responsibility for their decision to use nutritional supplements in their canine patients with liver disease. PMID:23890245

  10. Genomic instability and telomere fusion of canine osteosarcoma cells.

    PubMed

    Maeda, Junko; Yurkon, Charles R; Fujisawa, Hiroshi; Kaneko, Masami; Genet, Stefan C; Roybal, Erica J; Rota, Garrett W; Saffer, Ethan R; Rose, Barbara J; Hanneman, William H; Thamm, Douglas H; Kato, Takamitsu A

    2012-01-01

    Canine osteosarcoma (OSA) is known to present with highly variable and chaotic karyotypes, including hypodiploidy, hyperdiploidy, and increased numbers of metacentric chromosomes. The spectrum of genomic instabilities in canine OSA has significantly augmented the difficulty in clearly defining the biological and clinical significance of the observed cytogenetic abnormalities. In this study, eight canine OSA cell lines were used to investigate telomere fusions by fluorescence in situ hybridization (FISH) using a peptide nucleotide acid probe. We characterized each cell line by classical cytogenetic studies and cellular phenotypes including telomere associated factors and then evaluated correlations from this data. All eight canine OSA cell lines displayed increased abnormal metacentric chromosomes and exhibited numerous telomere fusions and interstitial telomeric signals. Also, as evidence of unstable telomeres, colocalization of γ-H2AX and telomere signals in interphase cells was observed. Each cell line was characterized by a combination of data representing cellular doubling time, DNA content, chromosome number, metacentric chromosome frequency, telomere signal level, cellular radiosensitivity, and DNA-PKcs protein expression level. We have also studied primary cultures from 10 spontaneous canine OSAs. Based on the observation of telomere aberrations in those primary cell cultures, we are reasonably certain that our observations in cell lines are not an artifact of prolonged culture. A correlation between telomere fusions and the other characteristics analyzed in our study could not be identified. However, it is important to note that all of the canine OSA samples exhibiting telomere fusion utilized in our study were telomerase positive. Pending further research regarding telomerase negative canine OSA cell lines, our findings may suggest telomere fusions can potentially serve as a novel marker for canine OSA. PMID:22916246

  11. Three-year serologic immunity against canine parvovirus type 2 and canine adenovirus type 2 in dogs vaccinated with a canine combination vaccine.

    PubMed

    Larson, L J; Schultz, R D

    2007-01-01

    A group of client-owned dogs and a group of dogs at a commercial kennel were evaluated for duration of antibody responses against canine parvovirus type 2 (CPV-2) and canine adenovirus type 1 (CAV-1) after receiving a combination vaccine containing recombinant canarypox-vectored canine distemper virus (CDV) and modified-live CPV-2, CAV-2, and canine parainfluenza virus, with (C6) or without (C4) two serovars of Leptospira (Recombitek C4 or C6, Merial). Duration of antibody, which correlates with protective immunity, was found to be at least 36 months in both groups. Recombitek combination vaccines can confidently be given every 3 years with assurance of protection in immunocompetent dogs against CPV-2 and CAV-1 as well as CDV. This allows this combination vaccine, like other, similar modified- live virus combination products containing CDV, CAV-2, and CPV-2, to be administered in accordance with the recommendations of the American Animal Hospital Association Canine Vaccine Task Force. PMID:18183549

  12. Polar ozone

    NASA Technical Reports Server (NTRS)

    Solomon, S.; Grose, W. L.; Jones, R. L.; Mccormick, M. P.; Molina, Mario J.; Oneill, A.; Poole, L. R.; Shine, K. P.; Plumb, R. A.; Pope, V.

    1990-01-01

    The observation and interpretation of a large, unexpected ozone depletion over Antarctica has changed the international scientific view of stratospheric chemistry. The observations which show the veracity, seasonal nature, and vertical structure of the Antarctic ozone hole are presented. Evidence for Arctic and midlatitude ozone loss is also discussed. The chemical theory for Antarctic ozone depletion centers around the occurrence of polar stratospheric clouds (PSCs) in Antarctic winter and spring; the climatology and radiative properties of these clouds are presented. Lab studies of the physical properties of PSCs and the chemical processes that subsequently influence ozone depletion are discussed. Observations and interpretation of the chemical composition of the Antarctic stratosphere are described. It is shown that the observed, greatly enhanced abundances of chlorine monoxide in the lower stratosphere are sufficient to explain much if not all of the ozone decrease. The dynamic meteorology of both polar regions is given, interannual and interhemispheric variations in dynamical processes are outlined, and their likely roles in ozone loss are discussed.

  13. Polar Markings

    NASA Technical Reports Server (NTRS)

    2006-01-01

    [figure removed for brevity, see original site] Context image for PIA02155 Polar Markings

    These bright and dark markings occurred near the end of summer in the south polar region. The dark material is likely dust that has been freed of frost cover.

    Image information: VIS instrument. Latitude -76.3N, Longitude 84.9E. 17 meter/pixel resolution.

    Note: this THEMIS visual image has not been radiometrically nor geometrically calibrated for this preliminary release. An empirical correction has been performed to remove instrumental effects. A linear shift has been applied in the cross-track and down-track direction to approximate spacecraft and planetary motion. Fully calibrated and geometrically projected images will be released through the Planetary Data System in accordance with Project policies at a later time.

    NASA's Jet Propulsion Laboratory manages the 2001 Mars Odyssey mission for NASA's Office of Space Science, Washington, D.C. The Thermal Emission Imaging System (THEMIS) was developed by Arizona State University, Tempe, in collaboration with Raytheon Santa Barbara Remote Sensing. The THEMIS investigation is led by Dr. Philip Christensen at Arizona State University. Lockheed Martin Astronautics, Denver, is the prime contractor for the Odyssey project, and developed and built the orbiter. Mission operations are conducted jointly from Lockheed Martin and from JPL, a division of the California Institute of Technology in Pasadena.

  14. Polar Layers

    NASA Technical Reports Server (NTRS)

    2006-01-01

    [figure removed for brevity, see original site] Context image for PIA02153 Polar Layers

    This image of the south polar region shows layered material. It is not known if the layers are formed yearly or if they form over the period of 10s to 100s of years or more.

    Image information: VIS instrument. Latitude -80.3N, Longitude 296.2E. 17 meter/pixel resolution.

    Note: this THEMIS visual image has not been radiometrically nor geometrically calibrated for this preliminary release. An empirical correction has been performed to remove instrumental effects. A linear shift has been applied in the cross-track and down-track direction to approximate spacecraft and planetary motion. Fully calibrated and geometrically projected images will be released through the Planetary Data System in accordance with Project policies at a later time.

    NASA's Jet Propulsion Laboratory manages the 2001 Mars Odyssey mission for NASA's Office of Space Science, Washington, D.C. The Thermal Emission Imaging System (THEMIS) was developed by Arizona State University, Tempe, in collaboration with Raytheon Santa Barbara Remote Sensing. The THEMIS investigation is led by Dr. Philip Christensen at Arizona State University. Lockheed Martin Astronautics, Denver, is the prime contractor for the Odyssey project, and developed and built the orbiter. Mission operations are conducted jointly from Lockheed Martin and from JPL, a division of the California Institute of Technology in Pasadena.

  15. Polar Ridges

    NASA Technical Reports Server (NTRS)

    2006-01-01

    [figure removed for brevity, see original site] Context image for PIA03662 Polar Ridges

    This ridge system is located in the south polar region.

    Image information: VIS instrument. Latitude -81.7N, Longitude 296.5E. 17 meter/pixel resolution.

    Note: this THEMIS visual image has not been radiometrically nor geometrically calibrated for this preliminary release. An empirical correction has been performed to remove instrumental effects. A linear shift has been applied in the cross-track and down-track direction to approximate spacecraft and planetary motion. Fully calibrated and geometrically projected images will be released through the Planetary Data System in accordance with Project policies at a later time.

    NASA's Jet Propulsion Laboratory manages the 2001 Mars Odyssey mission for NASA's Office of Space Science, Washington, D.C. The Thermal Emission Imaging System (THEMIS) was developed by Arizona State University, Tempe, in collaboration with Raytheon Santa Barbara Remote Sensing. The THEMIS investigation is led by Dr. Philip Christensen at Arizona State University. Lockheed Martin Astronautics, Denver, is the prime contractor for the Odyssey project, and developed and built the orbiter. Mission operations are conducted jointly from Lockheed Martin and from JPL, a division of the California Institute of Technology in Pasadena.

  16. Polar Textures

    NASA Technical Reports Server (NTRS)

    2005-01-01

    [figure removed for brevity, see original site] Context image for PIA03638 Polar Textures

    This image illustrates the variety of textures that appear in the south polar region during late summer.

    Image information: VIS instrument. Latitude 80.5S, Longitude 57.9E. 17 meter/pixel resolution.

    Note: this THEMIS visual image has not been radiometrically nor geometrically calibrated for this preliminary release. An empirical correction has been performed to remove instrumental effects. A linear shift has been applied in the cross-track and down-track direction to approximate spacecraft and planetary motion. Fully calibrated and geometrically projected images will be released through the Planetary Data System in accordance with Project policies at a later time.

    NASA's Jet Propulsion Laboratory manages the 2001 Mars Odyssey mission for NASA's Office of Space Science, Washington, D.C. The Thermal Emission Imaging System (THEMIS) was developed by Arizona State University, Tempe, in collaboration with Raytheon Santa Barbara Remote Sensing. The THEMIS investigation is led by Dr. Philip Christensen at Arizona State University. Lockheed Martin Astronautics, Denver, is the prime contractor for the Odyssey project, and developed and built the orbiter. Mission operations are conducted jointly from Lockheed Martin and from JPL, a division of the California Institute of Technology in Pasadena.

  17. Polar Terrains

    NASA Technical Reports Server (NTRS)

    2005-01-01

    [figure removed for brevity, see original site] Context image for PIA03577 Polar Terrains

    The region surrounding the South Polar Cap contains many different terrain types. This image shows both etched terrain and a region of 'mounds'.

    Image information: VIS instrument. Latitude 75S, Longitude 286.5E. 17 meter/pixel resolution.

    Note: this THEMIS visual image has not been radiometrically nor geometrically calibrated for this preliminary release. An empirical correction has been performed to remove instrumental effects. A linear shift has been applied in the cross-track and down-track direction to approximate spacecraft and planetary motion. Fully calibrated and geometrically projected images will be released through the Planetary Data System in accordance with Project policies at a later time.

    NASA's Jet Propulsion Laboratory manages the 2001 Mars Odyssey mission for NASA's Office of Space Science, Washington, D.C. The Thermal Emission Imaging System (THEMIS) was developed by Arizona State University, Tempe, in collaboration with Raytheon Santa Barbara Remote Sensing. The THEMIS investigation is led by Dr. Philip Christensen at Arizona State University. Lockheed Martin Astronautics, Denver, is the prime contractor for the Odyssey project, and developed and built the orbiter. Mission operations are conducted jointly from Lockheed Martin and from JPL, a division of the California Institute of Technology in Pasadena.

  18. Why segment the maxilla between laterals and canines?

    PubMed Central

    Esteves, Lucas Senhorinho; dos Santos, Jean Nunes; Sullivan, Steven M.; Martins, Luana Maria Rosário; Ávila, Carolina

    2016-01-01

    Introduction: Maxillary surgery on a bone segment enables movement in the sagittal and vertical planes. When performed on multiple segments, it further provides movement in the transverse plane. Typical sites for interdental osteotomies are between laterals and canines, premolars and canines, or between incisors. Additionally, osteotomies can be bilateral, unilateral or asymmetric. The ability to control intercanine width, buccolingual angulation of incisors, and correct Bolton discrepancy are some of the advantages of maxillary segmentation between laterals and canines. Objective: This article describes important features to be considered in making a clinical decision to segment the maxilla between laterals and canines when treating a dentoskeletal deformity. It further discusses the history of this surgical approach, the indications for its clinical use, the technique used to implement it, as well as its advantages, disadvantages, complications and stability. It is therefore hoped that this paper will contribute to disseminate information on this topic, which will inform the decision-making process of those professionals who wish to make use of this procedure in their clinical practice. Conclusions: Segmental maxillary osteotomy between laterals and canines is a versatile technique with several indications. Furthermore, it offers a host of advantages compared with single-piece osteotomy, or between canines and premolars. PMID:27007769

  19. Chlamydia in canine or feline coronary arteriosclerotic lesions

    PubMed Central

    2011-01-01

    Background There are numerous reports linking Chlamydia infection to human coronary atherosclerosis. However, there is a lack of data regarding this correlation in dogs and cats, and there are no reports investigating coronary arteriosclerosis and Chlamydia in these species. The aim of the present study was to examine whether there is a correlation between canine and feline spontaneous atherosclerosis or arteriosclerosis and the presence of Chlamydia. Archived histopathological samples of dogs (n = 16) and cats (n = 13) with findings of atherosclerosis or arteriosclerosis in heart tissue were examined for the presence of Chlamydiaceae using real-time PCR, ArrayTube Microarray and immunohistochemistry. Additionally, arteriosclerotic lesions of all cases were histologically classified and graded. Results Both canine atherosclerotic cases, and all 14 canine arteriosclerotic cases were negative for Chlamydia. Only one of the 13 arteriosclerotic feline cases was positive for Chlamydia by real-time PCR, revealing C. abortus by ArrayTube Microarray. To our knowledge, this is the first description of C. abortus in a cat. Overall, the type and grade of canine and feline arteriosclerotic lesions revealed similarities, and were predominantly moderate and hyperplastic. Conclusions These findings suggest that there is no obvious correlation between canine and feline coronary arteriosclerosis and the presence of Chlamydia. In order to draw final conclusions about the correlation between Chlamydia and canine atherosclerosis, examination of more samples is required. PMID:21906306

  20. Canine vector-borne diseases in Brazil

    PubMed Central

    Dantas-Torres, Filipe

    2008-01-01

    Canine vector-borne diseases (CVBDs) are highly prevalent in Brazil and represent a challenge to veterinarians and public health workers, since some diseases are of great zoonotic potential. Dogs are affected by many protozoa (e.g., Babesia vogeli, Leishmania infantum, and Trypanosoma cruzi), bacteria (e.g., Anaplasma platys and Ehrlichia canis), and helminths (e.g., Dirofilaria immitis and Dipylidium caninum) that are transmitted by a diverse range of arthropod vectors, including ticks, fleas, lice, triatomines, mosquitoes, tabanids, and phlebotomine sand flies. This article focuses on several aspects (etiology, transmission, distribution, prevalence, risk factors, diagnosis, control, prevention, and public health significance) of CVBDs in Brazil and discusses research gaps to be addressed in future studies. PMID:18691408

  1. Canine Distemper in Endangered Ethiopian Wolves

    PubMed Central

    Gordon, Christopher H.; Hussein, Alo; Laurenson, M. Karen; Malcolm, James R.; Marino, Jorgelina; Regassa, Fekede; Stewart, Anne-Marie E.; Fooks, Anthony R.; Sillero-Zubiri, Claudio

    2015-01-01

    The Ethiopian wolf (Canis simensis) is the world’s rarest canid; ≈500 wolves remain. The largest population is found within the Bale Mountains National Park (BMNP) in southeastern Ethiopia, where conservation efforts have demonstrated the negative effect of rabies virus on wolf populations. We describe previously unreported infections with canine distemper virus (CDV) among these wolves during 2005–2006 and 2010. Death rates ranged from 43% to 68% in affected subpopulations and were higher for subadult than adult wolves (83%–87% vs. 34%–39%). The 2010 CDV outbreak started 20 months after a rabies outbreak, before the population had fully recovered, and led to the eradication of several focal packs in BMNP’s Web Valley. The combined effect of rabies and CDV increases the chance of pack extinction, exacerbating the typically slow recovery of wolf populations, and represents a key extinction threat to populations of this highly endangered carnivore. PMID:25898177

  2. Canine distemper in endangered Ethiopian wolves.

    PubMed

    Gordon, Christopher H; Banyard, Ashley C; Hussein, Alo; Laurenson, M Karen; Malcolm, James R; Marino, Jorgelina; Regassa, Fekede; Stewart, Anne-Marie E; Fooks, Anthony R; Sillero-Zubiri, Claudio

    2015-05-01

    The Ethiopian wolf (Canis simensis) is the world's rarest canid; ≈500 wolves remain. The largest population is found within the Bale Mountains National Park (BMNP) in southeastern Ethiopia, where conservation efforts have demonstrated the negative effect of rabies virus on wolf populations. We describe previously unreported infections with canine distemper virus (CDV) among these wolves during 2005-2006 and 2010. Death rates ranged from 43% to 68% in affected subpopulations and were higher for subadult than adult wolves (83%-87% vs. 34%-39%). The 2010 CDV outbreak started 20 months after a rabies outbreak, before the population had fully recovered, and led to the eradication of several focal packs in BMNP's Web Valley. The combined effect of rabies and CDV increases the chance of pack extinction, exacerbating the typically slow recovery of wolf populations, and represents a key extinction threat to populations of this highly endangered carnivore. PMID:25898177

  3. Canine infectious cyclic thrombocytopenia found in Taiwan.

    PubMed

    Chang, A C; Chang, W L; Lin, C T; Pan, M J; Lee, S C

    1996-05-01

    Here were report the first canine infectious cyclic thrombocytopenia (CICT) found in Taiwan. Platelet-specific inclusions were detected in the blood smear of a military working dog. To identify the etiologic agent, the patient's blood was transmitted to three six-month-old German Shepherd dogs. The Ehrlichia platys-like inclusions were observed six to eight days after inoculation. Indirect fluorescent antibody test showed that the serum from the patient reacted specifically with the microorganisms within the platelets. Typical hematologic manifestations of E. platys infection, cyclic parasitemia and concomitant thrombocytopenia, were observed in these dogs. The prevalence of CICT in north Taiwan was also studied, and the incidence was 8.9% (4 out of 45) in civilian dogs and 97.1% (34 out of 35) in dogs from a heavily tick infested kennel. PMID:8741613

  4. A canine model of multiple portosystemic shunting.

    PubMed

    Howe, L M; Boothe, H W; Miller, M W; Boothe, D M

    2000-01-01

    The objective of this study was to develop and describe an experimental canine model of multiple acquired portosystemic shunts (PSS) similar in nature to spontaneously occurring PSS. Sixteen dogs were used and were divided into a control (n = 6) and a diseased group (n = 10). Dogs of the diseased group were administered dimethylnitrosamine (2 mg/kg of body weight, po) twice weekly, and clinicopathologic, ultrasonographic, and hepatic scintigraphic findings were recorded during the development of hepatic disease and PSS. Surgery was then performed to permit visual verification of multiple shunts, catheter placement for portography examination, and biopsy of the liver. All diseased dogs developed severe hepatic disease and multiple PSS as documented visually at surgery and on portography. Based on this study, dimethylnitrosamine-induced portosystemic shunting appears to be an appropriate model for spontaneously occurring multiple PSS secondary to portal hypertension. PMID:10741951

  5. European Surveillance for Pantropic Canine Coronavirus

    PubMed Central

    Cordonnier, Nathalie; Demeter, Zoltan; Egberink, Herman; Elia, Gabriella; Grellet, Aurélien; Le Poder, Sophie; Mari, Viviana; Martella, Vito; Ntafis, Vasileios; von Reitzenstein, Marcela; Rottier, Peter J.; Rusvai, Miklos; Shields, Shelly; Xylouri, Eftychia; Xu, Zach; Buonavoglia, Canio

    2013-01-01

    Highly virulent pantropic canine coronavirus (CCoV) strains belonging to subtype IIa were recently identified in dogs. To assess the distribution of such strains in Europe, tissue samples were collected from 354 dogs that had died after displaying systemic disease in France (n = 92), Hungary (n = 75), Italy (n = 69), Greece (n = 87), The Netherlands (n = 27), Belgium (n = 4), and Bulgaria (n = 1). A total of 124 animals tested positive for CCoV, with 33 of them displaying the virus in extraintestinal tissues. Twenty-four CCoV strains (19.35% of the CCoV-positive dogs) detected in internal organs were characterized as subtype IIa and consequently assumed to be pantropic CCoVs. Sequence and phylogenetic analyses of the 5′ end of the spike protein gene showed that pantropic CCoV strains are closely related to each other, with the exception of two divergent French viruses that clustered with enteric strains. PMID:23100349

  6. European surveillance for pantropic canine coronavirus.

    PubMed

    Decaro, Nicola; Cordonnier, Nathalie; Demeter, Zoltan; Egberink, Herman; Elia, Gabriella; Grellet, Aurélien; Le Poder, Sophie; Mari, Viviana; Martella, Vito; Ntafis, Vasileios; von Reitzenstein, Marcela; Rottier, Peter J; Rusvai, Miklos; Shields, Shelly; Xylouri, Eftychia; Xu, Zach; Buonavoglia, Canio

    2013-01-01

    Highly virulent pantropic canine coronavirus (CCoV) strains belonging to subtype IIa were recently identified in dogs. To assess the distribution of such strains in Europe, tissue samples were collected from 354 dogs that had died after displaying systemic disease in France (n = 92), Hungary (n = 75), Italy (n = 69), Greece (n = 87), The Netherlands (n = 27), Belgium (n = 4), and Bulgaria (n = 1). A total of 124 animals tested positive for CCoV, with 33 of them displaying the virus in extraintestinal tissues. Twenty-four CCoV strains (19.35% of the CCoV-positive dogs) detected in internal organs were characterized as subtype IIa and consequently assumed to be pantropic CCoVs. Sequence and phylogenetic analyses of the 5' end of the spike protein gene showed that pantropic CCoV strains are closely related to each other, with the exception of two divergent French viruses that clustered with enteric strains. PMID:23100349

  7. Refinement of the canine CD1 locus topology and investigation of antibody binding to recombinant canine CD1 isoforms.

    PubMed

    Schjaerff, Mette; Keller, Stefan M; Fass, Joseph; Froenicke, Lutz; Grahn, Robert A; Lyons, Leslie; Affolter, Verena K; Kristensen, Annemarie T; Moore, Peter F

    2016-03-01

    CD1 molecules are antigen-presenting glycoproteins primarily found on dendritic cells (DCs) responsible for lipid antigen presentation to CD1-restricted T cells. Despite their pivotal role in immunity, little is known about CD1 protein expression in dogs, notably due to lack of isoform-specific antibodies. The canine (Canis familiaris) CD1 locus was previously found to contain three functional CD1A genes: canCD1A2, canCD1A6, and canCD1A8, where two variants of canCD1A8, canCD1A8.1 and canCD1A8.2, were assumed to be allelic variants. However, we hypothesized that these rather represented two separate genes. Sequencing of three overlapping bacterial artificial chromosomes (BACs) spanning the entire canine CD1 locus revealed canCD1A8.2 and canCD1A8.1 to be located in tandem between canCD1A7 and canCD1C, and canCD1A8.1 was consequently renamed canCD1A9. Green fluorescent protein (GFP)-fused canine CD1 transcripts were recombinantly expressed in 293T cells. All proteins showed a highly positive GFP expression except for canine CD1d and a splice variant of canine CD1a8 lacking exon 3. Probing with a panel of anti-CD1 monoclonal antibodies (mAbs) showed that Ca13.9H11 and Ca9.AG5 only recognized canine CD1a8 and CD1a9 isoforms, and Fe1.5F4 mAb solely recognized canine CD1a6. Anti-CD1b mAbs recognized the canine CD1b protein, but also bound CD1a2, CD1a8, and CD1a9. Interestingly, Ca9.AG5 showed allele specificity based on a single nucleotide polymorphism (SNP) located at position 321. Our findings have refined the structure of the canine CD1 locus and available antibody specificity against canine CD1 proteins. These are important fundamentals for future investigation of the role of canine CD1 in lipid immunity. PMID:26687789

  8. Wilson disease and canine copper toxicosis.

    PubMed

    Brewer, G J

    1998-05-01

    In this article we review the current clinical and research status of Wilson disease and canine copper toxicosis. One of the main clinical challenges in Wilson disease is for clinicians to recognize the possibility of Wilson disease when young patients present with liver disease, psychiatric disease, or a movement-disorder type of neurologic disease. Once the possibility of the disease is recognized, many copper-related tests are available that are quite accurate in making the diagnosis or ruling it out. It is important to remember that this is an inherited disease and that family members at risk should be screened, particularly siblings. The cloning of the Wilson disease gene opened up the possibility that a direct DNA test could be developed, allowing convenient screening of certain patients and family members. However, the large number of mutations already found, with no small set of mutations dominating the picture, have thwarted this approach. Once the diagnosis has been made, a variety of treatments are available. For maintenance therapy, therapy of presymptomatic patients, and therapy of pregnant patients, we use zinc. For initial therapy of patients with liver disease, we use a combination of zinc and trientine. For initial therapy of patients with neurologic disease we use tetrathiomolybdate. Canine copper toxicosis in Bedlington terriers is due to a gene different from the gene for Wilson disease. However, the disease is treatable with the same array of anticopper therapies that work in humans. Recently, we established linkage of the copper toxicosis gene to a microsatellite marker, which has made available a linkage test to breeders of Bedlington terriers. PMID:9587157

  9. Canine Hip Dysplasia is Predictable by Genotyping

    PubMed Central

    Guo, Gang; Zhou, Zhengkui; Wang, Yachun; Zhao, Keyan; Zhu, Lan; Lust, George; Hunter, Linda; Friedenberg, Steven; Li, Junya; Zhang, Yuan; Harris, Stephen; Jones, Paul; Sandler, Jody; Krotscheck, Ursula; Todhunter, Rory; Zhang, Zhiwu

    2011-01-01

    Summary Objective To establish a predictive method using whole genome genotyping for early intervention in canine hip dysplasia (CHD) risk management, for the prevention of the progression of secondary osteoarthritis (OA), and for selective breeding. Design Two sets of dogs (6 breeds) were genotyped with dense SNPs covering the entire canine genome. The first set contained 359 dogs upon which a predictive formula for genomic breeding value (GBV) was derived by using their estimated breeding value (EBV) of the Norberg angle (a measure of CHD) and their genotypes. To investigate how well the formula would work for an individual dog with genotype only (without using EBV or phenotype), a cross validation was performed by masking the EBV of one dog at a time. The genomic data and the EBV of the remaining dogs were used to predict the GBV for the single dog that was left out. The second set of dogs included 38 new Labrador retriever dogs, which had no pedigree relationship to the dogs in the first set. Results The cross validation showed a strong correlation (r>0.7) between the EBV and the GBV. The independent validation showed a strong correlation (r=0.5) between GBV for the Norberg angle and the observed Norberg angle (no EBV was available for the new 38 dogs). Sensitivity, specificity, positive, and negative predictive value of the genomic data were all above 70%. Conclusions Prediction of CHD from genomic data is feasible, and can be applied for risk management of CHD and early selection for genetic improvement to reduce the prevalence of CHD in breeding programs. The prediction can be implemented before maturity, at which age current radiographic screening programs are traditionally applied, and as soon as DNA is available. PMID:21215318

  10. Dental anomalies in first-degree relatives of transposed canine probands.

    PubMed

    Bartolo, Adriana; Calleja, Neville; McDonald, Fraser; Camilleri, Simon

    2015-09-01

    The aim of this study was to investigate and compare the inheritance pattern and prevalence of inheritable dental anomalies in a sample of patients with maxillary canine-first premolar transposition and their first-degree relatives with a sample of palatally displaced canine families. Thirty-five consecutive maxillary canine-first premolar transposition probands and 111 first-degree relatives were matched to 35 consecutive palatally displaced canine probands and 115 first-degree relatives. These were assessed for palatally displaced canines and incisor-premolar hypodontia. Parental age at birth of the proband was also noted. The results revealed that (i) there is no difference in the overall prevalence of palatally displaced canine or incisor-premolar hypodontia between the groups of relatives; (ii) first-degree relatives of bilateral palatally displaced canine probands have a higher prevalence of palatally displaced canine and incisor-premolar hypodontia than those with unilateral palatally displaced canine; and (iii) maternal age at birth of the maxillary canine-first premolar transposition probands was significantly higher than that of the palatally displaced canine probands. The results suggest that maxillary canine-first premolar transposition and palatally displaced canine are unlikely to be different genetic entities and also indicate environmental or epigenetic influences on dental development. PMID:25634123

  11. Production of antibodies to canine IL-1beta and canine TNF to assess the role of proinflammatory cytokines.

    PubMed

    Baggio, Vannozza; Ott, Fabienne; Fischer, René W; Gram, Hermann; Peele, John; Spreng, David; Schmökel, Hugo; Jungi, Thomas W

    2005-08-15

    IL-1 and TNF are important proinflammatory cytokines implicated in both antimicrobial host defense and pathogenesis of diseases with an immune-mediated and/or inflammatory component. Respective studies in the dog have been hampered by the unavailability of reagents allowing the specific measurement of canine cytokine proteins and the effect of canine cytokine neutralization by Ab. Starting with recombinant canine (rcan) IL-1beta and rcanTNF, four polyclonal antisera and 22 mAb specific for rcanIL-1beta and rcanTNF were generated. Their usefulness in neutralization assays was determined. Using cytokine-containing supernatants of canine cells in bioassays, polyclonal antisera neutralized either canine IL-1beta or TNF. TNF was also neutralized by three antibodies developed in this study and one commercial mAb. The usefulness of monoclonal and polyclonal Ab in canine cytokine-specific Ab capture ELISA's was assessed. This resulted in the identification of a commercial mAb combination and one pair developed in this study allowing low levels of TNF to be detected by antibody capture ELISA. The detection limit was 141 pg/ml rcanTNF for both combinations. Using rcanIL-1beta as an antigen allowed the detection of lower concentrations of rcanIL-1beta (20 pg/ml, on the average) by a pair of polyclonal antisera than when monoclonals were used. By using such IL-1beta-specific and TNF-specific ELISA's, the respective cytokines were detected in supernatants of canine PBMC stimulated with LPS or heat-killed Listeria monocytogenes and interferon-gamma combined. Thus, monoclonal and polyclonal reagents were identified allowing the quantitation of canine IL-1beta and TNF production in vitro, and the neutralization of these cytokines. PMID:15982477

  12. Polar Barchans

    NASA Technical Reports Server (NTRS)

    2004-01-01

    20 July 2004 This Mars Global Surveyor (MGS) Mars Orbiter Camera (MOC) image shows dark, barchan sand dunes of the north polar region of Mars. Barchan dunes are simple, rounded forms with two horns that extend downwind. Inequalities in local wind patterns may result in one horn being extended farther than the other, as is the case for several dunes in this image. The image also shows several barchans may merge to form a long dune ridge. The horns and attendant slip faces on these dunes indicate wind transport of sand from the upper left toward the lower right. The image is located near 77.6oN, 103.6oW. The picture covers an area about 3 km (1.9 mi) wide; sunlight illuminates the scene from the lower left.

  13. Polar Polygons

    NASA Technical Reports Server (NTRS)

    2004-01-01

    26 December 2003 This Mars Global Surveyor (MGS) Mars Orbiter Camera (MOC) picture displays polygons outlined by cracks in the martian south polar region. This southern summer view was acquired in October 2003 and is located near 86.9oS, 170.6oW. Polygons similar in size and shape to these are common in the arctic and antarctic regions of Earth. On Earth, they indicate the presence (or the past presence) of ground ice and the freeze-thaw cycles that accompany this ice. On Mars, whether ground ice was responsible for these landforms is uncertain, but their presence is suggestive that ground ice may exist or may once have existed in this region. The picture covers an area 3 km (1.9 mi) wide. Sunlight illuminates the scene from the upper left.

  14. Canine Distemper Virus Infects Canine Keratinocytes and Immune Cells by Using Overlapping and Distinct Regions Located on One Side of the Attachment Protein▿

    PubMed Central

    Langedijk, Johannes P. M.; Janda, Jozef; Origgi, Francesco C.; Örvell, Claes; Vandevelde, Marc; Zurbriggen, Andreas; Plattet, Philippe

    2011-01-01

    The morbilliviruses measles virus (MeV) and canine distemper virus (CDV) both rely on two surface glycoproteins, the attachment (H) and fusion proteins, to promote fusion activity for viral cell entry. Growing evidence suggests that morbilliviruses infect multiple cell types by binding to distinct host cell surface receptors. Currently, the only known in vivo receptor used by morbilliviruses is CD150/SLAM, a molecule expressed in certain immune cells. Here we investigated the usage of multiple receptors by the highly virulent and demyelinating CDV strain A75/17. We based our study on the assumption that CDV-H may interact with receptors similar to those for MeV, and we conducted systematic alanine-scanning mutagenesis on CDV-H throughout one side of the β-propeller documented in MeV-H to contain multiple receptor-binding sites. Functional and biochemical assays performed with SLAM-expressing cells and primary canine epithelial keratinocytes identified 11 residues mutation of which selectively abrogated fusion in keratinocytes. Among these, four were identical to amino acids identified in MeV-H as residues contacting a putative receptor expressed in polarized epithelial cells. Strikingly, when mapped on a CDV-H structural model, all residues clustered in or around a recessed groove located on one side of CDV-H. In contrast, reported CDV-H mutants with SLAM-dependent fusion deficiencies were characterized by additional impairments to the promotion of fusion in keratinocytes. Furthermore, upon transfer of residues that selectively impaired fusion induction in keratinocytes into the CDV-H of the vaccine strain, fusion remained largely unaltered. Taken together, our results suggest that a restricted region on one side of CDV-H contains distinct and overlapping sites that control functional interaction with multiple receptors. PMID:21849439

  15. Canine retraction: A systematic review of different methods used

    PubMed Central

    Kulshrestha, Rohit S; Tandon, Ragni; Chandra, Pratik

    2015-01-01

    Canine retraction is a very important step in treatment of patients with crowding, or first premolar extraction cases. In severe crowding cases until, the canines have been distilized to relive the crowding, space to correctly align the incisors will not be available. Correct positioning of the canines after retraction is of great importance for the function, stability, and esthetics. The aim of this systematic review was to examine, in an evidence-based way, which kinds of canine retraction methods/techniques are most effective and which have the least side effects. A literature survey was performed by applying the Medline Database (Entrez PubMed) and Science Direct database covering the period from 1985 to 2014, to find out efficient ways to accomplish canine retraction. Randomized controlled trials (RCTs), prospective and retrospective controlled studies, and clinical trials were included. Two reviewers selected and extracted the data independently and assessed the quality of the retrieved studies. The search strategy resulted in 324 articles, of which 22 met the inclusion criteria. Due to the vast heterogeneity in study methods, the scientific evidence was too weak to evaluate retraction efficiency during space closure. The data so far reviewed proved that elastomeric power chains, elastic threads, magnets, NiTi coil springs, corticotomies, distraction osteogenesis, and laser therapy, all are able to provide optimum rate of tooth movements. All the methods were nearly similar to each other for retraction of canines Most of the techniques lead to anchorage loss in various amounts depending on the methods used. Most of the studies had serious problems with small sample size, confounding factors, lack of method error analysis, and no blinding in measurements. To obtain reliable scientific evidence, controlled RCT's with sufficient sample sizes are needed to determine which method/technique is the most effective in the respective retraction situation. Further

  16. Canine detection of free-ranging brown treesnakes on Guam

    USGS Publications Warehouse

    Savidge, J.A.; Stanford, J.W.; Reed, R.N.; Haddock, G.R.; Adams, A.A.Y.

    2011-01-01

    We investigated canine teams (dogs and their handlers) on Guam as a potential tool for finding invasive brown treesnakes (Boiga irregularis) in the wild. Canine teams searched a 40 ?? 40 m forested area for a snake that had consumed a dead mouse containing a radio-transmitter. To avoid tainting the target or target area with human scent, no snake was handled or closely approached prior to searches. Trials were conducted during the morning when these nocturnal snakes were usually hidden in refugia. A radiotracker knew the snake's location, but dog handlers and search navigators did not. Of 85 trials conducted over four months, the two canine teams had an average success rate of 35% of correctly defining an area ??? 5 ?? 5 m that contained the transmittered snake; the team with more experience prior to the trials had a success rate of 44% compared with 26% for the less experienced team. Canine teams also found 11 shed skins from wild snakes. Although dogs alerted outside the vicinity of transmittered snakes, only one wild, non-transmittered snake was found during the trials, possibly reflecting the difficulty humans have in locating non-transmittered brown treesnakes in refugia. We evaluated success at finding snakes as a function of canine team, number of prior trials (i.e. experience gained during the trials), recent canine success at finding a target snake, various environmental conditions, snake perch height, and snake characteristics (snout-vent length and sex). Success rate increased over the course of the trials. Canine team success also increased with increasing average humidity and decreased with increasing average wind speed. Our results suggest dogs could be useful at detecting brown treesnakes in refugia, particularly when compared to daytime visual searches by humans, but techniques are needed to help humans find and extract snakes once a dog has alerted. ?? New Zealand Ecological Society.

  17. The anti-canine distemper virus activities of ex vivo-expanded canine natural killer cells.

    PubMed

    Park, Ji-Yun; Shin, Dong-Jun; Lee, Soo-Hyeon; Lee, Je-Jung; Suh, Guk-Hyun; Cho, Duck; Kim, Sang-Ki

    2015-04-17

    Natural killer (NK) cells play critical roles in induction of antiviral effects against various viruses of humans and animals. However, few data on NK cell activities during canine distemper virus (CDV) infections are available. Recently, we established a culture system allowing activation and expansion of canine non-B, non-T, large granular NK lymphocytes from PBMCs of normal dogs. In the present study, we explored the ability of such expanded NK cells to inhibit CDV infection in vitro. Cultured CD3-CD5-CD21- NK cells produced large amounts of IFN-γ, exhibited highly upregulated expression of mRNAs encoding NK-cell-associated receptors, and demonstrated strong natural killing activity against canine tumor cells. Although the expanded NK cells were dose-dependently cytotoxic to both normal and CDV-infected Vero cells, CDV infection rendered Vero cells more susceptible to NK cells. Pretreatment with anti-CDV serum from hyperimmunized dogs enhanced the antibody-dependent cellular cytotoxicity (ADCC) of NK cells against CDV-infected Vero cells. The culture supernatants of NK cells, added before or after infection, dose-dependently inhibited both CDV replication and development of CDV-induced cytopathic effects (CPEs) in Vero cells. Anti-IFN-γ antibody neutralized the inhibitory effects of NK cell culture supernatants on CDV replication and CPE induction in Vero cells. Such results emphasize the potential significance of NK cells in controlling CDV infection, and indicate that NK cells may play roles both during CDV infection and in combating such infections, under certain conditions. PMID:25680810

  18. Unusual Cases of the Transmigrated Mandibular Canines: Report of 4 Cases

    PubMed Central

    Aktan, Ali Murat; Kara, Sami; Akgunlu, Faruk; Isman, Eren; Malkoc, Siddik

    2008-01-01

    The canine impaction is not uncommon in dental literature, but the transmigration of mandibular canine is a rare phenomenon, and some of them are far more extreme than others. In this study, we aimed to present the more extreme cases of the four transmigrated mandibular canine cases with special emphasis on their classification. PMID:19212522

  19. Citizen science: a new direction in canine behavior research.

    PubMed

    Hecht, Julie; Spicer Rice, Eleanor

    2015-01-01

    Researchers increasingly rely on members of the public to contribute to scientific projects-from collecting or identifying, to analyzing and disseminating data. The "citizen science" model proves useful to many thematically distinctive fields, like ornithology, astronomy, and phenology. The recent formalization of citizen science projects addresses technical issues related to volunteer participation--like data quality--so that citizen scientists can make longstanding, meaningful contributions to scientific projects. Since the late 1990s, canine science research has relied with greater frequency on the participation of the general public, particularly dog owners. These researchers do not typically consider the methods and technical issues that those conducting citizen science projects embrace and continue to investigate. As more canine science studies rely on public input, an in-depth knowledge of the benefits and challenges of citizen science can help produce relevant, high-quality data while increasing the general public's understanding of canine behavior and cognition as well as the scientific process. We examine the benefits and challenges of current citizen science models in an effort to enhance canine citizen science project preparation, execution, and dissemination. This article is part of a Special Issue entitled: Canine Behavior. PMID:25444773

  20. Permanent Maxillary Canine Agenesis: A Rare Case Report.

    PubMed

    Kambalimath, Halaswamy V; Jain, Somya; Patil, Raju Umaji; Asokan, Alexander; Kambalimath, Deepashri

    2015-01-01

    Congenitally missing teeth (CMT) are among one of the commonly known dental anomalies. The most frequently missing teeth in the permanent dentition, excluding the third molars, are mandibular second premolars and maxillary lateral incisors. Exclusive agenesis of both maxillary canines is an extremely rare occurrence and only a few cases have been reported. Previous studies showed that the prevalence of maxillary canine agenesis varies between 0.07 and 0.13%. In recent studies on Indian population, no cases of maxillary canine agenesis have been documented. This paper reports a case of non-syndromic bilateral agenesis of permanent maxillary canines, along with agenesis of both mandibular central incisors in a healthy 13-year-old Indian female patient; and a brief literature review on prevalence, etiology and treatment modalities of the condition. How to cite this article: Kambalimath HV, Jain S, Patil RU, Asokan A, Kambalimath D. Permanent Maxillary Canine Agenesis: A Rare Case Report. Int J Clin Pediatr Dent 2015; 8(3):242-246. PMID:26604546

  1. Biological effects on canine bladder by Nd:YAP laser

    NASA Astrophysics Data System (ADS)

    Zhang, Hui-Guo; Zhang, Mei-Jue; Zhu, Jing

    2005-07-01

    Objective: To observe the difference of biological effects on canine bladder by Nd:YAP laser with different power and different irradiation time. Methods: The canine bladder was irradiated with different power and different irradiation time. The effects of ablation and thermal coagulation in different laser settings were observed. The damage scale was evaluated macroscopically, with microscope and with electroscope. Results: The thermal coagulation effects is mostly and ablation effects is subordinate on the canine bladder by irradiation of Nd:YAP laser on. Pathology vision shows the thermal coagulation dose on perforation is 10W、6s;20W、4s;30W、3s;40W、2s;50-60W、1s;the dose of whole audience wear through is 10W、6s 20W、4s 30W、3s;40W、2s;50-60W、1s. Conclusions: The thermal coagulation effects is mostly and ablation effects is subordinate on biological effect of Nd:YAP laser on canine bladder. The better safety dose is power 10W、duration time less than 6s; power 20W、duration time less than 4s. power 30W、time less than 3s. power 40W、time less than 2s. The ablation and thermal coagulation effects of Nd:YAP laser on canine bladder is homocercal of power and time.

  2. Permanent Maxillary Canine Agenesis: A Rare Case Report

    PubMed Central

    Jain, Somya; Patil, Raju Umaji; Asokan, Alexander; Kambalimath, Deepashri

    2015-01-01

    ABSTRACT Congenitally missing teeth (CMT) are among one of the commonly known dental anomalies. The most frequently missing teeth in the permanent dentition, excluding the third molars, are mandibular second premolars and maxillary lateral incisors. Exclusive agenesis of both maxillary canines is an extremely rare occurrence and only a few cases have been reported. Previous studies showed that the prevalence of maxillary canine agenesis varies between 0.07 and 0.13%. In recent studies on Indian population, no cases of maxillary canine agenesis have been documented. This paper reports a case of non-syndromic bilateral agenesis of permanent maxillary canines, along with agenesis of both mandibular central incisors in a healthy 13-year-old Indian female patient; and a brief literature review on prevalence, etiology and treatment modalities of the condition. How to cite this article: Kambalimath HV, Jain S, Patil RU, Asokan A, Kambalimath D. Permanent Maxillary Canine Agenesis: A Rare Case Report. Int J Clin Pediatr Dent 2015; 8(3):242-246. PMID:26604546

  3. Discriminant canine index – a novel approach in sex determination

    PubMed Central

    Kiran, Chennoju Sai; Ramaswamy, Pachigolla; Swathi, Erva; Smitha, Balla; Sudhakar, Shankaran

    2015-01-01

    Summary Context Assessment of sex has significant contribution in construction of a physical profile of the decedent along with other parameters like race, stature and age. Sex determination with aid of skeletal remains is difficult procedure when, only a part of the body is obtainable. To solve this difficulty, tooth size standards based on odontometric data can be used in age and sex determination. The present study was undertaken with the objective to evaluate the reliability of sex determination using discriminant canine index (DCI). Methods A total of 120 subjects, with healthy periodontium and between the age groups of 15 to 40 years were selected randomly. Subjects with hard tissue abnormalities were excluded from the study. The maximum mesiodistal widths of left mandibular canines were measured intraorally with the help of divider and digital vernier caliper. Data was collected and analyzed statistically. Results A significant increase in the mesiodistal width of canines in males (7.21 ± 0.45 mm) when compared to females (6.77 ± 0.29 mm) was observed. The discriminant canine index (DCI) has identified 68.3% of males and 76.7% of females correctly with an overall accuracy rate of 72.5%. Conclusion The present study indicated that the DCI can produce reliable results and can be used as an alternative for mandibular canine index (MCI), for sex determination. PMID:26330903

  4. Posterior Cricoarytenoid Muscle Dynamics in Canines and Humans

    PubMed Central

    Chhetri, Dinesh K.; Neubauer, Juergen; Sofer, Elazar

    2015-01-01

    Objective The posterior cricoarytenoid (PCA) muscle is the sole abductor of the glottis and serves important functions during respiration, phonation, cough, and sniff. The present study examines vocal fold abduction dynamics during PCA muscle activation. Study Design Basic science study using an in vivo canine model and human subjects. Methods In four canines and five healthy humans vocal fold abduction time was measured using high speed video recording. In the canines, PCA muscle activation was achieved using graded stimulation of the PCA nerve branch. The human subjects performed coughing and sniffing tasks. High speed video and audio signals were concurrently recorded. Results In the canines the vocal fold moved posteriorly, laterally, and superiorly during abduction. Average time to reach 10%, 50% and 90% abduction was 23, 50, and 100 ms with low stimulation, 24, 58, and 129 ms with medium stimulation, and 21, 49, and 117 ms with high level stimulation. In the humans, 100% abduction times for coughing and sniffing tasks were 79 and 193 ms, respectively. Conclusion The PCA abduction times in canines are within the range in humans. The results also further support the notion that PCA muscles are fully active during cough. Level of Evidence N/A (Animal studies and basic research) PMID:24781959

  5. Transnasal endoscopic approach to the impacted maxillary canine.

    PubMed

    Marianetti, Tito Matteo; Torroni, Andrea; Gasparini, Giulio; Moro, Alessandro S; Pelo, Sandro

    2014-09-01

    The inclusion of maxillary canines is a very common condition. The intraoral approach to the canine extraction can be buccal or palatal depending on the position of the tooth. However, in some cases, the proximity to the nasal floor or the side wall of the nose makes the transoral approach rather invasive. The aim of this article was to describe a novel transnasal endoscopically assisted approach for the extraction of high palatal/paranasal impacted canines. Thirty-seven maxillary canines have been extracted in 29 patients. The surgical approaches were buccal in 5 cases, palatal in 24 cases, and transnasal endoscopically assisted in 8 cases. Patients treated with the transnasal approach required the least amount of pain killers in the postoperative period, and the average of the operative time was shorter than that of the transoral extraction. In our opinion, the transnasal endoscopically assisted approach is a safe and effective procedure for the extraction of highly impacted maxillary canines located within 2 cm from the piriform aperture. PMID:25102392

  6. Accuracy of Trained Canines for Detecting Bed Bugs (Hemiptera: Cimicidae).

    PubMed

    Cooper, Richard; Wang, Changlu; Singh, Narinderpal

    2014-12-01

    Detection of low-level bed bug, Cimex lectularius L. (Hemiptera: Cimicidae), infestations is essential for early intervention, confirming eradication of infestations, and reducing the spread of bed bugs. Despite the importance of detection, few effective tools and methods exist for detecting low numbers of bed bugs. Scent dogs were developed as a tool for detecting bed bugs in recent years. However, there are no data demonstrating the reliability of trained canines under natural field conditions. We evaluated the accuracy of 11 canine detection teams in naturally infested apartments. All handlers believed their dogs could detect infestations at a very high rate (≥95%). In three separate experiments, the mean (min, max) detection rate was 44 (10-100)% and mean false-positive rate was 15 (0-57)%. The false-positive rate was positively correlated with the detection rate. The probability of a bed bug infestation being detected by trained canines was not associated with the level of bed bug infestations. Four canine detection teams evaluated on multiple days were inconsistent in their ability to detect bed bugs and exhibited significant variance in accuracy of detection between inspections on different days. There was no significant relationship between the team's experience or certification status of teams and the detection rates. These data suggest that more research is needed to understand factors affecting the accuracy of canine teams for bed bug detection in naturally infested apartments. PMID:26470083

  7. Removal of deciduous canine tooth buds in Kenyan rural Maasai.

    PubMed

    Hassanali, J; Amwayi, P; Muriithi, A

    1995-04-01

    The removal of deciduous canine tooth buds in early childhood is a practice that has been documented in Kenya and in neighboring countries. This paper describes the occurrence, rationale and method of this practice amongst rural Kenyan Maasai. In a group of 95 children aged between six months and two years, who were examined in 1991/92, 87% were found to have undergone the removal of one or more deciduous canine tooth buds. In an older age group (3-7 years of age), 72% of the 111 children examined exhibited missing mandibular or maxillary deciduous canines. It was found that the actual removal of a deciduous tooth bud is often performed by middle-aged Maasai women who enucleate the developing tooth using a pointed pen-knife. There exists a strong belief among the Maasai that diarrhoea, vomiting and other febrile illnesses of early childhood are caused by the gingival swelling over the canine region, and which is thought to contain 'worms' or 'nylon' teeth. The immediate and long-term hazards of this practice include profuse bleeding, infection and damage to the developing permanent canines. A multi-disciplinary approach involving social anthropologists in addition to dental and medical personnel, is recommend in order to discourage this harmful operation that appears to be on the increase. PMID:7621751

  8. Polarized internal target apparatus

    DOEpatents

    Holt, R.J.

    1984-10-10

    A polarized internal target apparatus with a polarized gas target of improved polarization and density (achieved by mixing target gas atoms with a small amount of alkali metal gas atoms, and passing a high intensity polarized light source into the mixture to cause the alkali metal gas atoms to become polarized which interact in spin exchange collisions with target gas atoms yielding polarized target gas atoms) is described.

  9. Polarized internal target apparatus

    DOEpatents

    Holt, Roy J.

    1986-01-01

    A polarized internal target apparatus with a polarized gas target of improved polarization and density achieved by mixing target gas atoms with a small amount of alkali metal gas atoms, and passing a high intensity polarized light source into the mixture to cause the alkali metal gas atoms to become polarized which interact in spin exchange collisions with target gas atoms yielding polarized target gas atoms.

  10. Finite element analysis of rapid canine retraction through reducing resistance and distraction

    PubMed Central

    XUE, Junjie; YE, Niansong; YANG, Xin; WANG, Sheng; WANG, Jing; WANG, Yan; LI, Jingyu; MI, Congbo; LAI, Wenli

    2014-01-01

    Objective The aims of this study were to compare different surgical approaches to rapid canine retraction by designing and selecting the most effective method of reducing resistance by a three-dimensional finite element analysis. Material and Methods Three-dimensional finite element models of different approaches to rapid canine retraction by reducing resistance and distraction were established, including maxillary teeth, periodontal ligament, and alveolar. The models were designed to dissect the periodontal ligament, root, and alveolar separately. A 1.5 N force vector was loaded bilaterally to the center of the crown between first molar and canine, to retract the canine distally. The value of total deformation was used to assess the initial displacement of the canine and molar at the beginning of force loading. Stress intensity and force distribution were analyzed and evaluated by Ansys 13.0 through comparison of equivalent (von Mises) stress and maximum shear stress. Results The maximum value of total deformation with the three kinds of models occurred in the distal part of the canine crown and gradually reduced from the crown to the apex of the canine; compared with the canines in model 3 and model 1, the canine in model 2 had the maximum value of displacement, up to 1.9812 mm. The lowest equivalent (von Mises) stress and the lowest maximum shear stress were concentrated mainly on the distal side of the canine root in model 2. The distribution of equivalent (von Mises) stress and maximum shear stress on the PDL of the canine in the three models was highly concentrated on the distal edge of the canine cervix. Conclusions Removal of the bone in the pathway of canine retraction results in low stress intensity for canine movement. Periodontal distraction aided by surgical undermining of the interseptal bone would reduce resistance and effectively accelerate the speed of canine retraction. PMID:24626249

  11. Effect of simulated stages of the canine oestrous cycle on Escherichia coli binding to canine endometrium.

    PubMed

    Krekeler, N; Lodge, K M; Anderson, G A; Browning, G F; Charles, J A; Wright, P J

    2012-12-01

    Pyometra, a prevalent infectious uterine disease that affects intact middle-aged bitches, is typically associated with Escherichia coli. Our hypotheses were (i) that bacterial adhesion to canine endometrium differs between different stages of the oestrous cycle and (ii) that the adhesin FimH facilitates this adhesion. Twelve post-pubertal, ovariectomized greyhound bitches were treated with exogenous hormones to simulate different stages of the oestrous cycle. Tissue samples from each uterus were incubated with a pathogenic E. coli strain carrying the fimH gene, but no other adhesin genes (P4-wt)--or an E. coli strain in which fimH was insertionally inactivated (P4-∆fimH::kan)--or with phosphate-buffered saline as a negative control. After washing, tissue samples were homogenized for quantification of adherent bacteria. The differences in binding to canine endometrium at different stages of the oestrous cycle were not significant. However, the mean difference in binding of the P4-wt and the P4-∆fimH::kan across all stages of the simulated oestrous cycle was significant (p < 0.001 by paired t-test on geometric means). Individual differences in numbers of P4-wt bacteria bound between dogs might suggest genetic variations or epigenetic differences in FimH receptor expression by the endometrium, unrelated to the stage of the oestrous cycle. PMID:23279531

  12. Canine urolithiasis: retrospective analysis of 438 cases.

    PubMed

    Brown, N O; Parks, J L; Greene, R W

    1977-02-15

    In a retrospective analysis of 438 cases of canine urolithiasis, a total of 561 urolithic episodes were found to have occurred in a 6 1/2-year period. The hospital incidence of urolithiasis during that period, defined as the proportion of dogs hospitalized with urolithiasis to the total number of dogs hospitalized, was 2.8%. The major chemical component of the calculus in 307 dogs was phosphate; in 95 dogs, cystine; in 21 dogs, urate; in 12 dogs, oxalate; and in 3 dogs, carbonate. The Miniature Schnauzer, Dachsund, Dalmatian, Pug, Bulldog, Welsh Corgi, Beagle, and Bassett Hound were breeds that had a significantly higher (P less than 0.05) incidence of calculi than did breeds of other dogs hospitalized. Predisposition for calculi, by sex, was not found. Most dogs with calculi were between 3 and 7 years old. Most calculi were radiopaque and were located in the bladder or in the bladder and urethra. Specimens for bacteriologic culture were obtained by catheterization or by swabbing of tissue at the surgical site. Of 259 specimens obtained, 181 were culture-positive. The most common organisms isolated were Staphylococcus spp, Escherichia coli, Proteus spp, Streptococcus spp, and Klebsiella spp. Most of the bacteria were sensitive to gentamicin, chloramphenicol, nitrofurantoin, cephalothin, and methanamine mandelate. PMID:838615

  13. Canine Butterfly Glioblastomas: A Neuroradiological Review

    PubMed Central

    Rossmeisl, John H.; Clapp, Kemba; Pancotto, Theresa E.; Emch, Samantha; Robertson, John L.; Debinski, Waldemar

    2016-01-01

    In humans, high-grade gliomas may infiltrate across the corpus callosum resulting in bihemispheric lesions that may have symmetrical, winged-like appearances. This particular tumor manifestation has been coined a “butterfly” glioma (BG). While canine and human gliomas share many neuroradiological and pathological features, the BG morphology has not been previously reported in dogs. Here, we describe the magnetic resonance imaging (MRI) characteristics of BG in three dogs and review the potential differential diagnoses based on neuroimaging findings. All dogs presented for generalized seizures and interictal neurological deficits referable to multifocal or diffuse forebrain disease. MRI examinations revealed asymmetrical (2/3) or symmetrical (1/3), bihemispheric intra-axial mass lesions that predominantly affected the frontoparietal lobes that were associated with extensive perilesional edema, and involvement of the corpus callosum. The masses displayed heterogeneous T1, T2, and fluid-attenuated inversion recovery signal intensities, variable contrast enhancement (2/3), and mass effect. All tumors demonstrated classical histopathological features of glioblastoma multiforme (GBM), including glial cell pseudopalisading, serpentine necrosis, microvascular proliferation as well as invasion of the corpus callosum by neoplastic astrocytes. Although rare, GBM should be considered a differential diagnosis in dogs with an MRI evidence of asymmetric or symmetric bilateral, intra-axial cerebral mass lesions with signal characteristics compatible with glioma. PMID:27458589

  14. Canine Butterfly Glioblastomas: A Neuroradiological Review.

    PubMed

    Rossmeisl, John H; Clapp, Kemba; Pancotto, Theresa E; Emch, Samantha; Robertson, John L; Debinski, Waldemar

    2016-01-01

    In humans, high-grade gliomas may infiltrate across the corpus callosum resulting in bihemispheric lesions that may have symmetrical, winged-like appearances. This particular tumor manifestation has been coined a "butterfly" glioma (BG). While canine and human gliomas share many neuroradiological and pathological features, the BG morphology has not been previously reported in dogs. Here, we describe the magnetic resonance imaging (MRI) characteristics of BG in three dogs and review the potential differential diagnoses based on neuroimaging findings. All dogs presented for generalized seizures and interictal neurological deficits referable to multifocal or diffuse forebrain disease. MRI examinations revealed asymmetrical (2/3) or symmetrical (1/3), bihemispheric intra-axial mass lesions that predominantly affected the frontoparietal lobes that were associated with extensive perilesional edema, and involvement of the corpus callosum. The masses displayed heterogeneous T1, T2, and fluid-attenuated inversion recovery signal intensities, variable contrast enhancement (2/3), and mass effect. All tumors demonstrated classical histopathological features of glioblastoma multiforme (GBM), including glial cell pseudopalisading, serpentine necrosis, microvascular proliferation as well as invasion of the corpus callosum by neoplastic astrocytes. Although rare, GBM should be considered a differential diagnosis in dogs with an MRI evidence of asymmetric or symmetric bilateral, intra-axial cerebral mass lesions with signal characteristics compatible with glioma. PMID:27458589

  15. Viscoelastic and ultrasonic measurements of canine tissue

    NASA Astrophysics Data System (ADS)

    Kiss, Miklos; Varghese, Tomy

    2005-03-01

    Mechanical properties, of biological tissues, such as the complex modulus, are of interest for assessing the performance of elastographic methods that evaluate the stiffness characteristics of tissue. Determination of the mechanical properties of biological tissues is often limited by proper geometry of the sample, as well as homogeneity of the stress-strain relationship. Viscoelastic measurements were performed on in vitro canine liver tissue specimens, using a dynamic testing system, from 0.1 -- 100 Hz, and ultrasonic attenuation measurements were performed from 6 -- 9 MHz . Both normal tissues as well as thermal lesions prepared by immersion heating at several temperatures were tested. Experiments were conducted by uniaxially compressing tissue samples and measuring the load response. The resulting moduli spectra were then fit to both the Kelvin-Voigt model, as well as the Kelvin-Voigt fractional derivative model. The data agree well with the models and in comparing the results from the normal tissue with that of the thermal lesions, the concept of a complex modulus contrast is introduced and its applications to elastography are discussed.

  16. Molecular epizootiology of canine hepatozoonosis in Croatia.

    PubMed

    Vojta, Lea; Mrljak, Vladimir; Curković, Snjezana; Zivicnjak, Tatjana; Marinculić, Albert; Beck, Relja

    2009-08-01

    An epizootiological survey was conducted to investigate the prevalence of hepatozoonosis in a population of 924 apparently asymptomatic dogs from different regions of Croatia. DNA was isolated from canine blood and screening PCR on the 666 bp fragment of 18S rRNA revealed that 108 (11.8%) of dogs were infected. Positive samples were confirmed by partial sequencing of the 18S rRNA gene. The consensus sequences, derived from various sequence data sets, were compared with sequences of 18S ssrRNA of Hepatozoon spp. available in GenBank. The alignments revealed 106 Hepatozoon canis and two Hepatozoon sp. sequences. Among H. canis isolates, we found a certain amount of heterogeneity, while both Hepatozoon sp. isolates were identical to the Spanish isolate (Accession No. AY600625) from Clethrionomys glareolus. On the basis of eight commonly mutated nucleotide positions in the partial 18S rRNA gene sequence, we divided the H. canis isolates into five groups. The results obtained indicate a higher prevalence and significance of hepatozoonosis in Croatia than previously believed and demonstrate that the organisms belonging to H. canis that infect European dogs are genetically very heterogeneous. PMID:19249302

  17. Prevalence of canine dirofilariasis in Taiwan.

    PubMed

    Wu, Ching-Cheng; Fan, Ping-Chin

    2003-03-01

    Between 1993 and 1997, 837 stray dogs from North Taiwan were necropsied and examined for dog heartworm infections. A thick smear from 20 ml of peripheral blood from each dog was also prepared and examined for microfilariae (mf). The overall prevalence of adult worms in the dog population was 57%. The prevalence of mf in 1228 house dogs from different parts of Taiwan was also determined from 20 ml of peripheral blood in the same way. The overall prevalence of mf was 25%, with a value of 30% in the main island of Taiwan, this being 15 times higher than that in the offshore islands (2%). In Taiwan, the prevalence ranged from 4% in Hualien County, East Taiwan, to 41% in Nantou County, Central Taiwan. The mf prevalences on offshore islands were 1% on Liuchiu and 2% on Lanyu. The mf density per 20 ml blood in 82 house dogs was found to be 23 mf per dog, with a range of 3-97 mf per dog. A total of 477 stray dogs were found to be infected with adult worms of Dirofilaria immitis. The mean number of 7 worms per dog was obtained, with a range of 1-55 worms per dog. These results indicate that the prevalence of canine dirofilariasis has increased in Taiwan over the past 10 years. Moreover, the prevalence may be related to the wind speed, temperature, relative humidity, and altitude in the different areas surveyed. PMID:12590670

  18. Molecular epidemiology of canine parvovirus in Morocco.

    PubMed

    Amrani, Nadia; Desario, Costantina; Kadiri, Ahlam; Cavalli, Alessandra; Berrada, Jaouad; Zro, Khalil; Sebbar, Ghizlane; Colaianni, Maria Loredana; Parisi, Antonio; Elia, Gabriella; Buonavoglia, Canio; Malik, Jamal; Decaro, Nicola

    2016-07-01

    Since it first emergence in the mid-1970's, canine parvovirus 2 (CPV-2) has evolved giving rise to new antigenic variants termed CPV-2a, CPV-2b and CPV-2c, which have completely replaced the original strain and had been variously distributed worldwide. In Africa limited data are available on epidemiological prevalence of these new types. Hence, the aim of the present study was to determine circulating variants in Morocco. Through TaqMan-based real-time PCR assay, 91 samples, collected from symptomatic dogs originating from various cities between 2011 and 2015, were diagnosed. Positive specimens were characterised by means of minor groove binder (MGB) probe PCR. The results showed that all samples but one (98.9%) were CPV positive, of which 1 (1.1%) was characterised as CPV-2a, 43 (47.7%) as CPV-2b and 39 (43.3%) as CPV-2c. Interestingly, a co-infection with CPV-2b and CPV-2c was detected in 4 (4.4%) samples and 3 (3.3%) samples were not characterised. Sequencing of the full VP2 gene revealed these 3 uncharacterised strains as CPV-2c, displaying a change G4068A responsible for the replacement of aspartic acid with asparagine at residue 427, impacting the MGB probe binding. In this work we provide a better understanding of the current status of prevailing CPV strains in northern Africa. PMID:27083072

  19. Prognostic factors in canine mammary cancer.

    PubMed

    Misdorp, W; Hart, A A

    1976-04-01

    From a follow-up study of dogs surgically treated for mammary cancer, ten characteristics were analyzed statistically with special reference to their association with prognosis (expressed as survival for 2 years). The interrelations among five of the characteristics were also tested. The histologic type (descending range in malignancy: sarcomas greater than simple carcinomas greater than complex carcinomas), mode of growth (highly infiltrating greater than moderately infiltrating greater than expansive), clinical stage of complex carcinomas (large tumors and/or tumors involving the skin or underlying tissue greater than small, well-defined tumors), and size (greater than 15 cm greater than 11-15 cm greater than 5-10 cm greater than 0-5 cm) were of definite prognostic importance. The histologic grade was of possible prognostic importance. Localization, type of surgical therapy (mastectomy, block-dissection), growth in lymph vessels, involvement of regional lymph nodes, and duration of symptoms before treatment were not important to prognosis. A comparison between the factors associated with the prognosis of canine and human mammary cancer showed many similarities. However, the involvement of regional lymph nodes, important in women, was not so in bitches. PMID:1255797

  20. Canine mdr1 gene mutation in Japan.

    PubMed

    Kawabata, Akiko; Momoi, Yasuyuki; Inoue-Murayama, Miho; Iwasaki, Toshiroh

    2005-11-01

    Frequency of the 4-bp deletion mutant in canine mdr1 gene was examined in 193 dogs of eight breeds in Japan. The mutant allele was found in Collies, Australian Shepherds, and Shetland Sheepdogs, where its respective frequencies were 58.3%, 33.3%, and 1.2%. The MDR1 protein was detected on peripheral blood mononuclear cells (PBMC) from a MDR1/MDR1 dog, but not on PBMC from a mdr1-1Delta/mdr1-1Delta Collie. Rhodamine 123 was extruded from MDR1/MDR1 lymphocytes. That excretion was inhibited by a MDR1 inhibitor, verapamil. On the other hand, Rh123 excretion was not observed from lymphocytes derived from a mdr1-1Delta/mdr1-1Delta Collie. These results indicated that the mutant mdr1 allele also existed in Collie-breed dogs in Japan at high rates and that mdr1-1Delta /mdr1-1Delta dogs have no functional MDR1. PMID:16327220

  1. In vivo canine muscle function assay.

    PubMed

    Childers, Martin K; Grange, Robert W; Kornegay, Joe N

    2011-01-01

    We describe a minimally-invasive and reproducible method to measure canine pelvic limb muscle strength and muscle response to repeated eccentric contractions. The pelvic limb of an anesthetized dog is immobilized in a stereotactic frame to align the tibia at a right angle to the femur. Adhesive wrap affixes the paw to a pedal mounted on the shaft of a servomotor to measure torque. Percutaneous nerve stimulation activates pelvic limb muscles of the paw to either push (extend) or pull (flex) against the pedal to generate isometric torque. Percutaneous tibial nerve stimulation activates tibiotarsal extensor muscles. Repeated eccentric (lengthening) contractions are induced in the tibiotarsal flexor muscles by percutaneous peroneal nerve stimulation. The eccentric protocol consists of an initial isometric contraction followed by a forced stretch imposed by the servomotor. The rotation effectively lengthens the muscle while it contracts, e.g., an eccentric contraction. During stimulation flexor muscles are subjected to an 800 msec isometric and 200 msec eccentric contraction. This procedure is repeated every 5 sec. To avoid fatigue, 4 min rest follows every 10 contractions with a total of 30 contractions performed. PMID:21494224

  2. Secretory pattern of canine growth hormone

    SciTech Connect

    French, M.B.; Vaitkus, P.; Cukerman, E.; Sirek, A.; Sirek, O.V.

    1987-02-01

    The aim of this paper was to define the secretory pattern of growth hormone (GH) under basal conditions in fasted, conscious, male dogs accustomed to handling. Blood samples were withdrawn from a cephalic vein at 15-min intervals. In this way, any ultradian rhythms, if present, could be detected within the frequency range of 0.042-2 cycles/h. In addition, samples were drawn at either 1- or 2.5-min intervals for 2.5 or 5 h to determine whether frequency components greater than 2 cycles/h were present. GH was measured by radioimmunoassay and the raw data were submitted to time series analysis employing power spectral estimation by means of fast Fourier transformation techniques. Peak plasma levels were up to 12 times higher than the baseline concentration of approx. 1 ng/ml. Spectral analysis revealed an endogenous frequency of 0.22 cycles/h, i.e., a periodicity of 4.5 h/cycle. The results indicate that under basal conditions the secretory bursts of canine GH are limited to one peak every 4.5 h.

  3. Recombinant Canine Coronaviruses in Dogs, Europe

    PubMed Central

    Mari, Viviana; Elia, Gabriella; Addie, Diane D.; Camero, Michele; Lucente, Maria Stella; Martella, Vito; Buonavoglia, Canio

    2010-01-01

    Coronaviruses of potential recombinant origin with porcine transmissible gastroenteritis virus (TGEV), referred to as a new subtype (IIb) of canine coronavirus (CCoV), were recently identified in dogs in Europe. To assess the distribution of the TGEV-like CCoV subtype, during 2001–2008 we tested fecal samples from dogs with gastroenteritis. Of 1,172 samples, 493 (42.06%) were positive for CCoV. CCoV-II was found in 218 samples, and CCoV-I and CCoV-II genotypes were found in 182. Approximately 20% of the samples with CCoV-II had the TGEV-like subtype; detection rates varied according to geographic origin. The highest and lowest rates of prevalence for CCoV-II infection were found in samples from Hungary and Greece (96.87% and 3.45%, respectively). Sequence and phylogenetic analyses showed that the CCoV-IIb strains were related to prototype TGEV-like strains in the 5′ and the 3′ ends of the spike protein gene. PMID:20031041

  4. Genotyping of Canine parvovirus in western Mexico.

    PubMed

    Pedroza-Roldán, César; Páez-Magallan, Varinia; Charles-Niño, Claudia; Elizondo-Quiroga, Darwin; De Cervantes-Mireles, Raúl Leonel; López-Amezcua, Mario Alberto

    2015-01-01

    Canine parvovirus (CPV) is one of the most common infectious agents related to high morbidity rates in dogs. In addition, the virus is associated with severe gastroenteritis, diarrhea, and vomiting, resulting in high death rates, especially in puppies and nonvaccinated dogs. To date, there are 3 variants of the virus (CPV-2a, CPV-2b, and CPV-2c) circulating worldwide. In Mexico, reports describing the viral variants circulating in dog populations are lacking. In response to this deficiency, a total of 41 fecal samples of suspected dogs were collected from October 2013 through April 2014 in the Veterinary Hospital of the University of Guadalajara in western Mexico. From these, 24 samples resulted positive by polymerase chain reaction, and the viral variant was determined by restriction fragment length polymorphism. Five positive diagnosed samples were selected for partial sequencing of the vp2 gene and codon analysis. The results demonstrated that the current dominant viral variant in Mexico is CPV-2c. The current study describes the genotyping of CPV strains, providing valuable evidence of the dominant frequency of this virus in a dog population from western Mexico. PMID:25525144

  5. [Diagnostic tools for canine parvovirus infection].

    PubMed

    Proksch, A L; Hartmann, K

    2015-01-01

    Canine parvovirus (CPV) infection is one of the most important and common infectious diseases in dogs, in particular affecting young puppies when maternal antibodies have waned and vaccine-induced antibodies have not yet developed. The mortality rate remains high. Therefore, a rapid and safe diagnostic tool is essential to diagnose the disease to 1) provide intensive care treatment and 2) to identify virus-shedding animals and thus prevent virus spread. Whilst the detection of antibodies against CPV is considered unsuitable to diagnose the disease, there are several different methods to directly detect complete virus, virus antigen or DNA. Additionally, to test in commercial laboratories, rapid in-house tests based on ELISA are available worldwide. The specificity of the ELISA rapid in-house tests is reported to be excellent. However, results on sensitivity vary and high numbers of false-negative results are commonly reported, which potentially leads to misdiagnosis. Polymerase chain reaction (PCR) is a very sensitive and specific diagnostic tool. It also provides the opportunity to differentiate vaccine strains from natural infection when sequencing is performed after PCR. PMID:26403490

  6. Allergens causing atopic diseases in canine.

    PubMed

    Youn, Hwa-Young; Kang, Hyung-Seok; Bhang, Dong-Ha; Kim, Min-Kue; Hwang, Cheol-Yong; Han, Hong-Ryul

    2002-12-01

    Canine atopic skin disease is seasonal or sometimes non-seasonal immune-mediated skin disease which occurs commonly in Korea. The definite clinical sign is systemic pruritus, especially on periocular parts, external ear, interdigit spaces and lateral flank. For diagnosis of this dermatitis, complete history taking followed by intradermal skin test and serum in vitro IgE test needs to be performed. Allergen selection for the diagnosis and treatment of atopic dermatitis should be varied geographically. In this study, with intradermal skin test(IDST) the prevalence of atopic disease and what allergens are involved in are researched. Allergens used for IDST included 26 allergen extracts from six allergen groups: grasses, trees, weeds, molds, epidermal allergens and environmental allergens. The number of allergens was 42 in which the positive and negative controls are included. The most common positive allergen reaction was the house dust mites on IDST(22/35, 63%). The other positive allergen reactions were to flea(3/35, 9%), molds(1/35, 3%), house dusts(2/35, 6%), feathers (1/35, 3%), cedar/juniper(1/35, 3%), timothy grass(1/35, 3%) and dandelion(1/35, 3%). In this study, the most prevalent allergen causing atopic dermatitis in dogs in Korea was the house dust mites followed by the flea. PMID:12819384

  7. Experimental canine parvovirus infection in dogs.

    PubMed

    Pollock, R V

    1982-04-01

    In specific pathogen-free dogs, clinical signs of experimental canine parvovirus infection were mild, inconsistent and transient. Clinical signs were more pronounced in conventionally-raised dogs, but the severe disease reported in field cases was not reproduced in either group. A pronounced plasma viremia occurred on the 2nd to 4th day post-infection (d.p.i.) in dogs challenged oronasally. Antibody was detectable on the 5th d.p.i. Marked pyrexia was rare, but a significant temperature rise usually coincided with the appearance of antibody and the cessation of viremia. Significant lymphopenia, but not leukopenia, occurred on the 3rd to 7th d.p.i. Virus could be readily isolated from fecal matter on the 3rd to 8th d.p.i.; a few dogs continued to shed virus for up to 12 days. In dogs challenged parenterally, the onset of elevated temperatures, viral shed and antibody production occurred 24-48 hours sooner. Convalescent dogs were no longer contagious for susceptible contact animals 25 days or longer after challenge, although infectious virus persisted in feces for more than 6 months at room temperature. Active giardiasis seemed to exacerbate the clinical syndrome, although treatment with corticosteroids or anti-thymocyte serum did not. PMID:6211333

  8. A clinically relevant canine lung cancer model

    SciTech Connect

    Benfield, J.R.; Shors, E.C.; Hammond, W.G.; Paladugu, R.R.; Cohen, A.H.; Jensen, T.; Fu, P.C.; Pak, H.Y.; Teplitz, R.L.

    1981-12-01

    Research on early human lung cancer is difficult; we have sought a canine correlate. Regimens included endobronchial submucosal injections and topical focal applications of benzo(a)pyrene, nitrosomethylurea, dimethylbenzanthracene, and methylcholanthrene, singly or in combinations. Sustained-release discs were placed into lung parenchyma or sutured into major bronchi. Tracheal segments were isolated as cervical pedicle grafts. Gross and histological evolution was reproducible. Columnar and basal hyperplasia and squamous metaplasia were early changes. Atypia occurred within 6 weeks and was found in all dogs within 16 to 18 weeks. Invasive cancers occurred within 8 to 65 months. No tracheal graft developed cancer. Of 15 dogs with parenchymal sustained-release implants, 1 to date has developed cancer in 8 months. Four endobronchial regimens have produced 16 cancers in 56 lungs at risk for 18 to 65 months. No cancers developed in 23 lungs at risk from eight other regimens. Of 10 dogs at risk for unilateral endobronchial cancer, 5 have had cancer. Of 23 dogs with both lungs at risk, 9 developed cancer. We have shown focal carcinogenesis with well-defined pathogenesis and an extended preneoplastic period at predictable sites in a lung cancer model.

  9. Canine detection odor signatures for explosives

    NASA Astrophysics Data System (ADS)

    Williams, Marc; Johnston, J. M.; Cicoria, Matt; Paletz, E.; Waggoner, L. Paul; Edge, Cindy C.; Hallowell, Susan F.

    1998-12-01

    Dogs are capable of detecting and discriminating a number of compounds constituting a complex odor. However, they use only a few of these to recognize a substance. The focus of this research is to determine the compounds dogs learn to use in recognizing explosives. This is accomplished by training dogs under behavioral laboratory conditions to respond differentially on separate levers to 1) blank air, 2) a target odor, such as an explosive, and 3) all other odors (non-target odors). Vapor samples are generated by a serial dilution vapor generator whose operation and output is characterized by GC/MS. Once dogs learn this three-lever discrimination, testing sessions are conducted containing a number of probe trials in which vapor from constituent compounds of the target is presented. Which lever the dogs respond to on these probe trials indicates whether they can smell the compound at all (blank lever) or whether it smells like toe target odor (e.g., the explosive) or like something else. This method was conducted using TNT, C-4, and commercial dynamite. The data show the dogs' reactions to each of the constituent compounds tested for each explosive. Analysis of these data reveal the canine detection odor signature for these explosives.

  10. Recombinant vaccine for canine parvovirus in dogs.

    PubMed

    López de Turiso, J A; Cortés, E; Martínez, C; Ruiz de Ybáñez, R; Simarro, I; Vela, C; Casal, I

    1992-05-01

    VP2 is the major component of canine parvovirus (CPV) capsids. The VP2-coding gene was engineered to be expressed by a recombinant baculovirus under the control of the polyhedrin promoter. A transfer vector that contains the lacZ gene under the control of the p10 promoter was used in order to facilitate the selection of recombinants. The expressed VP2 was found to be structurally and immunologically indistinguishable from authentic VP2. The recombinant VP2 shows also the capability to self-assemble, forming viruslike particles similar in size and appearance to CPV virions. These viruslike particles have been used to immunize dogs in different doses and combinations of adjuvants, and the anti-CPV responses have been measured by enzyme-linked immunosorbent assay, monolayer protection assays, and an assay for the inhibition of hemagglutination. A dose of ca. 10 micrograms of VP2 was able to elicit a good protective response, higher than that obtained with a commercially available, inactivated vaccine. The results indicate that these viruslike particles can be used to protect dogs from CPV infection. PMID:1313899

  11. Comparative mapping of canine and human proximal Xq and genetic analysis of canine X-linked severe combined immunodeficiency

    SciTech Connect

    Deschenes, S.M.; Puck, J.M.; Dutra, A.S.

    1994-09-01

    Parallel genetic analysis of animal and human genetic diseases can facilitate the identification and characterization of the causative gene defects. For example, canine X-linked severe combined immunodeficiency (SCID) is characterized by clinical, pathological, and immunological manifestations similar to the most common form of human SCID. To derive a canine syntenic map including genes that in humans are located in proximal Xq, near human X-linked SCID, poly (TG) polymorphisms were identified at the canine phosphoglycerate kinase (PGK) and choroideremia (CHM) loci. These plus a polymorphic poly (CAG) sequence in exon 1 of the canine androgen receptor gene (AR) were used to genotype members of the colony informative for X-linked SCID. No recombinations among SCIDX1, AR, PGK, or CHM were observed. Fluorescence in situ hybridization localized PGK and CHM to proximal Xq in the dog, in the same chromosomal location occupied by the human genes. Somatic cell hybrid analysis and methylation differences at AR demonstrated that female dogs carrying X-linked SCID have the same lymphocyte-limited skewed X-chromosome inactivation patterns as human carriers. These genetic and phenotypic findings provide evidence that mutations in the same gene, now identified as the {gamma} chain of the IL-2 receptor, cause canine and human X-linked SCID. This approach is an efficient method for comparative gene mapping and disease identification. 35 refs., 4 figs., 1 tab.

  12. Correction of bilateral impacted mandibular canines with a lip bumper for anchorage reinforcement.

    PubMed

    Agarwal, Sachin; Yadav, Sumit; Shah, Neelesh V; Valiathan, Ashima; Uribe, Flavio; Nanda, Ravindra

    2013-03-01

    Multiple treatment options are available to patients with impacted manibular canines in addition to a retained deciduous canine. This article describes the treatment of a prepubertal girl, aged 10 years 6 months, with a skeletal Class I, dental Class II Division 1 malocclusion, retrognathic mandible, deep overbite, proclined maxillary incisors, midline diastema, and bilateral mandibular canine impaction. The orthodontic treatment plan included extraction of the deciduous canine and forced eruption of the impacted canines. A modified lip bumper appliance was used both for forced eruption and to reinforce anchorage. Through the collaborative efforts of an orthodontist and an oral surgeon, an excellent esthetic and functional outcome was achieved. PMID:23452974

  13. Immunopathogenic and Neurological Mechanisms of Canine Distemper Virus

    PubMed Central

    Carvalho, Otávio Valério; Botelho, Clarisse Vieira; Ferreira, Caroline Gracielle Torres; Scherer, Paulo Oldemar; Soares-Martins, Jamária Adriana Pinheiro; Almeida, Márcia Rogéria; Silva Júnior, Abelardo

    2012-01-01

    Canine distemper is a highly contagious viral disease caused by the canine distemper virus (CDV), which is a member of the Morbillivirus genus, Paramyxoviridae family. Animals that most commonly suffer from this disease belong to the Canidae family; however, the spectrum of natural hosts for CDV also includes several other families of the order Carnivora. The infectious disease presents worldwide distribution and maintains a high incidence and high levels of lethality, despite the availability of effective vaccines, and no specific treatment. CDV infection in dogs is characterized by the presentation of systemic and/or neurological courses, and viral persistence in some organs, including the central nervous system (CNS) and lymphoid tissues. An elucidation of the pathogenic mechanisms involved in canine distemper disease will lead to a better understanding of the injuries and clinical manifestations caused by CDV. Ultimately, further insight about this disease will enable the improvement of diagnostic methods as well as therapeutic studies. PMID:23193403

  14. Testosterone biotransformation by the isolated perfused canine pancreas

    SciTech Connect

    Fernandez-del Castillo, C.; Diaz-Sanchez, V.; Varela-Fascinetto, G.; Altamirano, A.; Odor-Morales, A.; Lopez-Medrano, R.M.; Robles-Diaz, G. )

    1991-01-01

    There is strong evidence indicating that the pancreas is under the influence of sex steroid hormones, and that it may even participate in their biosynthesis and metabolism. In the present study, (3H)testosterone was perfused into the isolated canine pancreas, and measured in the effluent with several of its metabolites (5 alpha-dihydrotestosterone, androstenedione, and estradiol). Results show that testosterone is readily transformed by the canine pancreas. The main product found in the effluent is androstenedione. The testis and spleen were also perfused with (3H)testosterone and used as controls. In both cases, this hormone appeared mostly unchanged in the effluent as compared to the pancreatic perfusion (p less than 0.0001). From our data, we conclude that the canine pancreas has the capacity to transform sex steroid hormones, and could be considered an extragonadal site of sex steroid biosynthesis.

  15. The Cost of Canine Rabies on Four Continents.

    PubMed

    Anderson, A; Shwiff, S A

    2015-08-01

    We estimated the economic impacts of canine rabies in Latin America, Africa and Asia. Direct and indirect costs of rabies post-exposure prophylaxis, dog vaccination and control, rabies diagnostic testing and cattle mortality-related costs were accounted for. The number of human deaths was updated from previous estimates based on population growth, and the costs associated with the risk of human mortality were incorporated. We accounted for uncertainty associated with the parameter estimates using a Monte Carlo simulation and estimated that the global burden of canine rabies is approximately $124 billion annually. This result illustrates the potential benefits that could be realized if canine rabies was eliminated and provides an important benchmark against which the cost of any potential elimination campaign can be compared. PMID:24112194

  16. Severe canine distemper outbreak in unvaccinated dogs in Mozambique.

    PubMed

    Zacarias, Julieta; Dimande, Alberto; Achá, Sara; Dias, Paula T; Leonel, Elisa M; Messa, Aurora; Macucule, Baltazar; Júnior, José L; Bila, Custódio G

    2016-01-01

    Although significant animal suffering caused by preventable diseases is frequently seen in developing countries, reports of this are scarce. This report describes avoidable animal suffering owing to a suspected canine distemper (CD) outbreak in unvaccinated dogs owned by low-income families in Mozambique that killed approximately 200 animals. Affected dogs exhibited clinical signs, and gross and microscopic lesions compatible with CD. Immunohistochemical staining confirmed the presence of canine distemper virus (CDV) in the kidney of one dog from the cohort. This brief communication again illustrates that large outbreaks of CDV in unvaccinated dogs occur and that large-scale avoidable suffering and threats to the health of dogs and wild canines continue. Mass vaccination supported by government and non-government organisations is recommended. PMID:27543040

  17. Canine dysautonomia in a litter of Havanese puppies.

    PubMed

    Hull, Noah C; O'Toole, Donal; Miller, Myrna M; Shoults, Hannah; Deck, Robin; Jones, Warren; Johnson, Gayle C; Shaw, Daniel P; Schumaker, Brant A

    2015-09-01

    Canine dysautonomia is a sporadic, generally fatal disease that rarely affects groups of related animals. Four 10-week-old Havanese puppies from a litter of 5 developed clinical signs of canine dysautonomia. The 4 affected dogs were exposed to an outdoor environment, whereas the fifth littermate was not exposed to the outdoors and remained clinically healthy. Clinical signs of dysautonomia developed 10-16 days after going outside the house. An unrelated dog also developed dysautonomia after exposure to 1 of the affected Havanese littermates. All 5 dogs had morphological changes consistent with dysautonomia (widespread neuronal degeneration in autonomic ganglia, select brainstem nuclei, and ventral horn motor neurons). Differential diagnoses were excluded through negative toxicological evaluation, fecal parasite screening, negative Canine distemper virus reverse transcription polymerase chain reaction, fluorescent antibody testing, attempted virus isolation, and electron microscopy. The 5 affected dogs were in the Kansas City, Missouri area, where there is a high incidence of dysautonomia. PMID:26179098

  18. Distribution of beta-amyloid in the canine brain.

    PubMed

    Hou, Y; White, R G; Bobik, M; Marks, J S; Russell, M J

    1997-03-01

    The distribution of amyloid-beta protein (A beta) in the canine brain was demonstrated by immunochemistry on serially sectioned tissues from 10 aged mixed breed dogs. Summation of quantitative data and relegation to anatomical sites for the 10 dogs showed A beta to be widely distributed in the cortex and hippocampus while completely absent in the brain stem and cerebellum. The highest density of A beta was in the dentate gyrus of the hippocampus. Cortical areas exhibiting the greatest A beta deposition were the posterior and medial suprasylvius gyrus and the proreus gyrus of the frontal lobe. Unlike humans the canine entorhinal cortex, amygdala, basal ganglia and olfactory bulbs were rarely affected. This suggested that the highly developed olfactory pathways of the canine are generally spared from A beta deposition. PMID:9141082

  19. The anatomy and internal aerodynamics of canine olfaction

    NASA Astrophysics Data System (ADS)

    Craven, Brent; Paterson, Eric; Settles, Gary

    2007-11-01

    High-resolution magnetic resonance imaging (MRI) scans of the nasal airway of a large dog reveal an intricate scrollwork of nasal conchae providing large surface area for heat, moisture, and odorant transfer. From these anatomical scans we reconstruct a 3-D surface model of the nasal passage and extract detailed morphometric data providing insight into the internal airflows of canine olfaction. A complicated airway network is revealed, wherein the branched maxilloturbinate and ethmoturbinate scrolls are structurally distinct. 3-D airway connectivity also reveals separate respiratory and olfactory flow paths. Knowing the approximate airflow rate and frequency of canine sniffing, we find Reynolds numbers that are, surprisingly, well below the turbulent-flow threshold. Finally, the internal aerodynamics and transport phenomena of canine olfaction are considered via non-dimensional analysis and initially-simple theoretical and computational models. (To appear in the Anatomical Record.)

  20. The innervation of canine hip joint capsule: an anatomic study.

    PubMed

    Huang, C H; Hou, S M; Yeh, L S

    2013-12-01

    To clarify the contributions of the nerves supplying the canine hip joint capsule for clinical application, cadaver study of six healthy mongrel dogs was performed. The pelvises and hindlimbs of cadavers were dissected and fixed in formaldehyde. Innervation of the joint capsule was investigated with the aid of an operative microscope. As a result, the canine hip joint capsule receives multiple innervations from articular branches of four nerves. They are articular nerve fibres of femoral, obturator, cranial gluteal and sciatic nerves from the cranioventral, caudoventral, craniolateral and dorsolateral directions of the joint, respectively. No branch originating from the caudal gluteal nerve was observed innervating the hip joint capsule. Our data provides useful information for research on the canine hip joint, including pain analysis with hip disorders and surgical nerve blockade to relieve pain. PMID:23410229

  1. Polar Color

    NASA Technical Reports Server (NTRS)

    2004-01-01

    [figure removed for brevity, see original site]

    Released 3 May 2004 This nighttime visible color image was collected on January 1, 2003 during the Northern Summer season near the North Polar Troughs.

    This daytime visible color image was collected on September 4, 2002 during the Northern Spring season in Vastitas Borealis. The THEMIS VIS camera is capable of capturing color images of the martian surface using its five different color filters. In this mode of operation, the spatial resolution and coverage of the image must be reduced to accommodate the additional data volume produced from the use of multiple filters. To make a color image, three of the five filter images (each in grayscale) are selected. Each is contrast enhanced and then converted to a red, green, or blue intensity image. These three images are then combined to produce a full color, single image. Because the THEMIS color filters don't span the full range of colors seen by the human eye, a color THEMIS image does not represent true color. Also, because each single-filter image is contrast enhanced before inclusion in the three-color image, the apparent color variation of the scene is exaggerated. Nevertheless, the color variation that does appear is representative of some change in color, however subtle, in the actual scene. Note that the long edges of THEMIS color images typically contain color artifacts that do not represent surface variation.

    Image information: VIS instrument. Latitude 79, Longitude 346 East (14 West). 19 meter/pixel resolution.

    Note: this THEMIS visual image has not been radiometrically nor geometrically calibrated for this preliminary release. An empirical correction has been performed to remove instrumental effects. A linear shift has been applied in the cross-track and down-track direction to approximate spacecraft and planetary motion. Fully calibrated and geometrically projected images will be released through the Planetary Data System in accordance with

  2. Penta prism laser polarizer.

    PubMed

    Lotem, H; Rabinovitch, K

    1993-04-20

    A novel type of laser prism polarizer is proposed. The polarizer is characterized by a high transmission efficiency, a high optical damage threshold, and a high extinction ratio. The polarizer is shaped like a regular penta prism and, thus, it is a constant deviation angle device. Polarization effects occur upon the two internal cascade reflections in the prism. Anisotropic and Isotropic types of the polarizer are discussed. The isotropic polarizer is a prism made of a high refractive-index glass coated by multilayer polarization-type dielectric coatings. Efficient s-state polarization is obtained because of p-state leakage upon the two internal cascade reflections. The anisotropic polarizer is made of a birefringent crystal in which angular polarization splitting is obtained by the bireflectance (double-reflection) effect. Fanning of a laser beam into up to eight polarized beams is possible in a prism made of a biaxial crystal. PMID:20820335

  3. Plasma free amino acid profiles of canine mammary gland tumors

    PubMed Central

    Azuma, Kazuo; Tsuka, Takeshi; Imagawa, Tomohiro; Minami, Saburo; Okamoto, Yoshiharu

    2012-01-01

    The purpose of this study was to elucidate the relationship between plasma free amino acid (PFAA) levels and the clinical stages of mammary gland tumors (MGT) in dogs. PFAA levels in canines with malignant mammary tumors were decreased compared to those of healthy animals. The levels of aspartate and ornithine, in the dogs with tumor metastasis were significantly decreased when compared to those of dogs that did not have metastases. Results of this study indicate that PFAA levels could be a risk factor or biomarker for canine MGT metastasis. PMID:23271187

  4. Cadherin Cell Adhesion System in Canine Mammary Cancer: A Review

    PubMed Central

    Gama, Adelina; Schmitt, Fernando

    2012-01-01

    Cadherin-catenin adhesion complexes play important roles by providing cell-cell adhesion and communication in different organ systems. Abnormal expression of cadherin adhesion molecules constitutes a common phenomenon in canine mammary cancer and has been frequently implicated in tumour progression. This paper summarizes the current knowledge on cadherin/catenin adhesion molecules (E-cadherin, β-catenin, and P-cadherin) in canine mammary cancer, focusing on the putative biological functions and clinical significance of these molecules in this disease. This paper highlights the need for further research studies in this setting in order to elucidate the role of these adhesion molecules during tumour progression and metastasis. PMID:22973534

  5. Comprehensive characterization of commercially available canine training aids.

    PubMed

    Tipple, Christopher A; Caldwell, Patricia T; Kile, Brian M; Beussman, Douglas J; Rushing, Blake; Mitchell, Natalie J; Whitchurch, Christian J; Grime, Martin; Stockham, Rex; Eckenrode, Brian A

    2014-09-01

    Effective and reliable training aids for victim recovery canine teams is essential for law enforcement and investigative purposes. Without adequate training aids, the rate of recovery for sub surface or surface human remains deposition using canine teams may be adversely affected and result in confusing information. The composition of three commercially available canine training aids that purportedly generate volatile components responsible for the odor of human decomposition is relatively simple and not closely related to those compounds experimentally determined to be present at the site of surface or sub-surface human remains. In this study, these different commercial formulations were chemically characterized using six different sampling approaches, including two applications of direct liquid injection, solid-phase microextraction (SPME), purge and trap, ambient preconcentration/thermal desorption, and cryogenic preconcentration/thermal desorption. Direct liquid injections resulted in the fewest number of detected compounds, while a cryogen based thermal desorption method detected the greatest number of compounds in each formulation. Based solely upon the direct liquid injection analysis, Pseudo™ Scent I was composed of approximately 29±4% and 71±5% of 2-pyrrolidinone and 4-aminobutanoic acid, respectively. This same analysis showed that Pseudo™ Scent II was composed of approximately 11±1, 11±1, 24±5, and 54±7% of putrescine, cadaverine, 2-pyrrolidinone, and 4-aminobutanoic acid, respectively. Headspace analysis was conducted to more closely simulate the process whereby a canine's nose would capture a volatiles profile. More compounds were detected using the headspace sampling method; however, the vast majority was not consistent with current data on human decomposition. Additionally, the three formulations were tested in outdoor and indoor scenarios by a double-blinded canine team, using a certified and specifically trained victim recovery canine

  6. Interdisciplinary approach for the management of bilaterally impacted maxillary canines

    PubMed Central

    Sukh, Ram; Singh, Gyan P.; Tandon, Pradeep

    2014-01-01

    Interdisciplinary approach for the management of malocclusion provides a holistic approach of patient management. Prudent treatment planning is necessary to achieve the various treatment goals. This case report describes the orthodontic management of a 16-year-old adolescent female patient with bilateral labially impacted maxillary canines. The problems associated with impacted maxillary canines and the biomechanical interventions used for this patient are discussed. The treatment protocol involved surgical intervention, followed by sequential traction of the impacted teeth. An interdisciplinary approach to treatment with different mechanical strategies led to the achievement of the desired esthetic, functional, and occlusal treatment goals. PMID:25395776

  7. Canine distemper spillover in domestic dogs from urban wildlife.

    PubMed

    Kapil, Sanjay; Yeary, Teresa J

    2011-11-01

    Canine distemper virus (CDV) causes a major disease of domestic dogs that develops as a serious systemic infection in unvaccinated or improperly vaccinated dogs. Domesticated dogs are the main reservoir of CDV, a multihost pathogen. This virus of the genus Morbillivirus in the family Paramyxoviridae occurs in other carnivorous species including all members of the Canidae and Mustelidae families and in some members of the Procyonidae, Hyaenidae, Ursidae, and Viverridae families. Canine distemper also has been reported in the Felidae family and marine mammals. The spread and incidences of CDV epidemics in dogs and wildlife here and worldwide are increasing. PMID:22041204

  8. Identification of quantitative trait loci (QTL) for canine hip dysplasia and canine elbow dysplasia in Bernese mountain dogs.

    PubMed

    Pfahler, Sophia; Distl, Ottmar

    2012-01-01

    A genome-wide association study for canine hip dysplasia (CHD) and canine elbow dysplasia (CED) using the Illumina canine high density bead chip had been performed for 174 Bernese mountain dogs. General and mixed linear model analysis identified two different regions with single nucleotide polymorphisms (SNPs) on dog chromosome (CFA) 14 significantly associated with CHD and a further significantly CHD-associated region on CFA37. For CED, four SNPs on CFA11 and 27 were significantly associated. The identified SNPs of four associated regions included nearby candidate genes. These possible positional candidates were the genes PON2 on CFA14 and FN1 on CFA37 for CHD and the genes LMNB1 on CFA11 and WNT10B on CFA27 for CED. PMID:23189162

  9. Identification of Quantitative Trait Loci (QTL) for Canine Hip Dysplasia and Canine Elbow Dysplasia in Bernese Mountain Dogs

    PubMed Central

    Pfahler, Sophia; Distl, Ottmar

    2012-01-01

    A genome-wide association study for canine hip dysplasia (CHD) and canine elbow dysplasia (CED) using the Illumina canine high density bead chip had been performed for 174 Bernese mountain dogs. General and mixed linear model analysis identified two different regions with single nucleotide polymorphisms (SNPs) on dog chromosome (CFA) 14 significantly associated with CHD and a further significantly CHD-associated region on CFA37. For CED, four SNPs on CFA11 and 27 were significantly associated. The identified SNPs of four associated regions included nearby candidate genes. These possible positional candidates were the genes PON2 on CFA14 and FN1 on CFA37 for CHD and the genes LMNB1 on CFA11 and WNT10B on CFA27 for CED. PMID:23189162

  10. Polarized Light in Astronomy.

    ERIC Educational Resources Information Center

    King, D. J.

    1983-01-01

    The application of very sensitive electronic detecting devices during the last decade has revolutionized and revitalized the study of polarization in celestial objects. The nature of polarization, how polaroids work, interstellar polarization, dichroic filters, polarization by scattering, and modern polarimetry are among the topics discussed. (JN)

  11. Linearly polarized fiber amplifier

    DOEpatents

    Kliner, Dahv A.; Koplow, Jeffery P.

    2004-11-30

    Optically pumped rare-earth-doped polarizing fibers exhibit significantly higher gain for one linear polarization state than for the orthogonal state. Such a fiber can be used to construct a single-polarization fiber laser, amplifier, or amplified-spontaneous-emission (ASE) source without the need for additional optical components to obtain stable, linearly polarized operation.

  12. Nondiffracting transversally polarized beam.

    PubMed

    Yuan, G H; Wei, S B; Yuan, X-C

    2011-09-01

    Generation of a nondiffracting transversally polarized beam by means of transmitting an azimuthally polarized beam through a multibelt spiral phase hologram and then highly focusing by a high-NA lens is presented. A relatively long depth of focus (∼4.84λ) of the electric field with only radial and azimuthal components is achieved. The polarization of the wavefront near the focal plane is analyzed in detail by calculating the Stokes polarization parameters. It is found that the polarization is spatially varying and entirely transversally polarized, and the polarization singularity disappears at the beam center, which makes the central bright channel possible. PMID:21886250

  13. Diode laser prostatectomy (VLAP): initial canine evaluation

    NASA Astrophysics Data System (ADS)

    Kopchok, George E.; Verbin, Chris; Ayres, Bruce; Peng, Shi-Kaung; White, Rodney A.

    1995-05-01

    This study evaluated the acute and chronic effects of diode laser (960 nm) prostatectomy using a Prolase II fiber in a canine model (n equals 5). The laser fiber consists of a 1000 um quartz fiber which reflects a cone of laser energy, at 45 degree(s) to the axis of the fiber, into the prostatic urethra (Visual Laser Ablation of Prostate). Perineal access was used to guide a 15.5 Fr cystoscope to the level of the prostate. Under visual guidance and continual saline irrigation, 60 watts of laser power was delivered for 60 seconds at 3, 9, and 12 o'clock and 30 seconds at the 6 o'clock (posterior) positions for a total energy fluence of 12,600 J. One prostate received an additional 60 second exposure at 3 and 9 o'clock for a total fluence of 19,800 J. The prostates were evaluated at one day (n equals 1) and 8 weeks (n equals 4). The histopathology of laser effects at one day show areas of necrosis with loss of glandular structures and stromal edema. Surrounding this area was a zone of degenerative glandular structures extending up to 17.5 mm (cross sectional diameter). The histopathology of the 8 week laser treated animals demonstrated dilated prostatic urethras with maximum cross- sectional diameter of 23.4 mm (mean equals 18.5 +/- 3.9 mm). This study demonstrates the effectiveness of diode laser energy for prostatic tissue coagulation and eventual sloughing. The results also demonstrate the safety of diode laser energy, with similar tissue response as seen with Nd:YAG laser, for laser prostatectomy.

  14. Canine Babesioses in Noninvestigated Areas of Serbia.

    PubMed

    Gabrielli, Simona; Otašević, Suzana; Ignjatović, Aleksandra; Savić, Sara; Fraulo, Maurizio; Arsić-Arsenijević, Valentina; Momčilović, Stefan; Cancrini, Gabriella

    2015-09-01

    During the years 2012-2014, a total of 158 outdoor dogs from Pančevo and Đurđevo (northern Serbia) and Niš and Prokuplje (southern Serbia) were submitted to molecular analyses (PCR and sequencing) for canine babesioses. An overall prevalence of 21.5% was found, due to the species Babesia sp. 'spanish dog' (10.1%), B. gibsoni (5.7%), B. canis vogeli (1.9%), B. caballi (1.9%), and B. microti (1.9%). In addition, sequence analysis showed the presence of Hepatozoon canis in a dog from Niš. No significant difference between infected and noninfected dogs was found by age, sex, and place of residence, whereas there was difference regarding the presence of ticks (p<0.005) and application of preventive measures such as applying of antitick drugs/devices. Moreover, a significant difference was established by area: Dogs from Prokuplje showed infection rates (59.1%) higher than dogs from Pančevo (11.9%), Niš (4.5), and Đurđevo (where infected dogs were not found), and a different geographical distribution of the species was found. The presence of so many Babesia species and the first identification of H. canis will allow investigations on the pathogenic role played by each one and suggests entomological studies on the tick species that are more suitable vectors for each of them. Finally, the presence of so many infected dogs offers the opportunity of evaluating the hypothesis of a possible zoonotic role of babesial species affecting dogs. PMID:26348245

  15. Chromatography purification of canine adenoviral vectors.

    PubMed

    Segura, María Mercedes; Puig, Meritxell; Monfar, Mercè; Chillón, Miguel

    2012-06-01

    Canine adenovirus vectors (CAV2) are currently being evaluated for gene therapy, oncolytic virotherapy, and as vectors for recombinant vaccines. Despite the need for increasing volumes of purified CAV2 preparations for preclinical and clinical testing, their purification still relies on the use of conventional, scale-limited CsCl ultracentrifugation techniques. A complete downstream processing strategy for CAV2 vectors based on membrane filtration and chromatography is reported here. Microfiltration and ultra/diafiltration are selected for clarification and concentration of crude viral stocks containing both intracellular and extracellular CAV2 particles. A DNase digestion step is introduced between ultrafiltration and diafiltration operations. At these early stages, concentration of vector stocks with good recovery of viral particles (above 80%) and removal of a substantial amount of protein and nucleic acid contaminants is achieved. The ability of various chromatography techniques to isolate CAV2 particles was evaluated. Hydrophobic interaction chromatography using a Fractogel propyl tentacle resin was selected as a first chromatography step, because it allows removal of the bulk of contaminating proteins with high CAV2 yields (88%). An anion-exchange chromatography step using monolithic supports is further introduced to remove the remaining contaminants with good recovery of CAV2 particles (58-69%). The main CAV2 viral structural components are visualized in purified preparations by electrophoresis analyses. Purified vector stocks contained intact icosahedral viral particles, low contamination with empty viral capsids (10%), and an acceptable total-to-infectious particle ratio (below 30). The downstream processing strategy that was developed allows preparation of large volumes of high-quality CAV2 stocks. PMID:22799886

  16. Detection of Bacteriuria by Canine Olfaction.

    PubMed

    Maurer, Maureen; McCulloch, Michael; Willey, Angel M; Hirsch, Wendi; Dewey, Danielle

    2016-03-01

    Background.  Urinary tract infections (UTIs) are a significant medical problem , particularly for patients with neurological conditions and the elderly. Detection is often difficult in these patients, resulting in delayed diagnoses and more serious infections such as pyelonephritis and life-threatening sepsis. Many patients have a higher risk of UTIs because of impaired bladder function, catheterization, and lack of symptoms. Urinary tract infections are the most common nosocomial infection; however, better strategies are needed to improve early detection of the disease. Methods.  In this double-blinded, case-control, validation study, we obtained fresh urine samples daily in a consecutive case series over a period of 16 weeks. Dogs were trained to distinguish urine samples that were culture-positive for bacteriuria from those of culture-negative controls, using reward-based clicker and treat methods. Results.  Samples were obtained from 687 individuals (from 3 months to 92 years of age; 86% female and 14% male; 34% culture-positive and 66% culture-negative controls). Dogs detected urine samples positive for 100 000 colony-forming units/mL Escherichia coli (N = 250 trials; sensitivity 99.6%, specificity 91.5%). Dilution of E coli urine with distilled water did not affect accuracy at 1% (sensitivity 100%, specificity 91.1%) or 0.1% (sensitivity 100%, specificity 93.6%) concentration. Diagnostic accuracy was similar to Enterococcus (n = 50; sensitivity 100%, specificity 93.9%), Klebsiella (n = 50; sensitivity 100%, specificity 95.1%), and Staphylococcus aureus (n = 50; sensitivity 100%, specificity 96.3%). All dogs performed with similarly high accuracy: overall sensitivity was at or near 100%, and specificity was above 90%. Conclusions.  Canine scent detection is an accurate and feasible method for detection of bacteriuria. PMID:27186578

  17. Detection of Bacteriuria by Canine Olfaction

    PubMed Central

    Maurer, Maureen; McCulloch, Michael; Willey, Angel M.; Hirsch, Wendi; Dewey, Danielle

    2016-01-01

    Background. Urinary tract infections (UTIs) are a significant medical problem , particularly for patients with neurological conditions and the elderly. Detection is often difficult in these patients, resulting in delayed diagnoses and more serious infections such as pyelonephritis and life-threatening sepsis. Many patients have a higher risk of UTIs because of impaired bladder function, catheterization, and lack of symptoms. Urinary tract infections are the most common nosocomial infection; however, better strategies are needed to improve early detection of the disease. Methods. In this double-blinded, case-control, validation study, we obtained fresh urine samples daily in a consecutive case series over a period of 16 weeks. Dogs were trained to distinguish urine samples that were culture-positive for bacteriuria from those of culture-negative controls, using reward-based clicker and treat methods. Results. Samples were obtained from 687 individuals (from 3 months to 92 years of age; 86% female and 14% male; 34% culture-positive and 66% culture-negative controls). Dogs detected urine samples positive for 100 000 colony-forming units/mL Escherichia coli (N = 250 trials; sensitivity 99.6%, specificity 91.5%). Dilution of E coli urine with distilled water did not affect accuracy at 1% (sensitivity 100%, specificity 91.1%) or 0.1% (sensitivity 100%, specificity 93.6%) concentration. Diagnostic accuracy was similar to Enterococcus (n = 50; sensitivity 100%, specificity 93.9%), Klebsiella (n = 50; sensitivity 100%, specificity 95.1%), and Staphylococcus aureus (n = 50; sensitivity 100%, specificity 96.3%). All dogs performed with similarly high accuracy: overall sensitivity was at or near 100%, and specificity was above 90%. Conclusions. Canine scent detection is an accurate and feasible method for detection of bacteriuria. PMID:27186578

  18. Microbiological and histopathological aspects of canine pyometra

    PubMed Central

    Coggan, Jennifer Anne; Melville, Priscilla Anne; de Oliveira, Clair Motos; Faustino, Marcelo; Moreno, Andréa Micke; Benites, Nilson Roberti

    2008-01-01

    As pyometra is recognized as one of the main causes of disease and death in the bitch the purposes of this study were to evaluate microbiological and histopathological aspects of canine pyometra and to research the virulence factors of the E. coli isolates identifying possible risks to human health. The microbiological isolation from the intrauterine contents of 100 dogs with pyometra was carried out and the virulence factors in the E. coli strains were identified using PCR method. This study also consisted of the counting of microorganisms colonies forming units in samples of intrauterine content, tests of antimicrobial susceptibility of the E. coli isolates and the histological examination of the uterus. E. coli was the most prevalent microorganism isolated (76.6%) and 120 strains (79.5%) were positive for sfa, 86 (56.9%) were positive for cnf, 87 (57.6%) were positive for pap, 52 (34.4%) were positive for hly, 51 (33.8%) were positive for iuc and 5 (3.3%) were positive for afa genes. One observed more sensitivity of E. coli to norfloxacin, polimixin B, sulphazotrin, chloranfenicol and enrofloxacin. In 42% of the samples of uterine walls where microorganisms were isolated, the sizes of the areas of the inflammatory responses corresponded to 39–56%. Virulence factors were identified in 98.0% of the strains evaluated, demonstrating a high frequency of potentially pathogenic E. coli. It must be considered that dogs are animals that are living in close proximity to man for thousands of years and have an important role in the transmission of E. coli to other animals and to man. PMID:24031249

  19. Crossed elliptical polarization undulator

    SciTech Connect

    Sasaki, Shigemi

    1997-05-01

    The first switching of polarization direction is possible by installing two identical helical undulators in series in a same straight section in a storage ring. By setting each undulator in a circular polarization mode in opposite handedness, one can obtain linearly polarized radiation with any required polarization direction depending on the modulator setting between two undulators. This scheme can be used without any major degradation of polarization degree in any low energy low emittance storage ring.

  20. Polarization-balanced beamsplitter

    DOEpatents

    Decker, D.E.

    1998-02-17

    A beamsplitter assembly is disclosed that includes several beamsplitter cubes arranged to define a plurality of polarization-balanced light paths. Each polarization-balanced light path contains one or more balanced pairs of light paths, where each balanced pair of light paths includes either two transmission light paths with orthogonal polarization effects or two reflection light paths with orthogonal polarization effects. The orthogonal pairing of said transmission and reflection light paths cancels polarization effects otherwise caused by beamsplitting. 10 figs.

  1. Polarization-balanced beamsplitter

    DOEpatents

    Decker, Derek E.

    1998-01-01

    A beamsplitter assembly that includes several beamsplitter cubes arranged to define a plurality of polarization-balanced light paths. Each polarization-balanced light path contains one or more balanced pairs of light paths, where each balanced pair of light paths includes either two transmission light paths with orthogonal polarization effects or two reflection light paths with orthogonal polarization effects. The orthogonal pairing of said transmission and reflection light paths cancels polarization effects otherwise caused by beamsplitting.

  2. Expression of prolactin receptors in normal canine mammary tissue, canine mammary adenomas and mammary adenocarcinomas

    PubMed Central

    2012-01-01

    Background Mammary tumors represent the most common neoplastic disease in female dogs. Recently, the promoting role of prolactin (PRL) in the development of human breast carcinoma has been shown. Possible proliferative, anti-apoptotic, migratory and angiogenic effects of PRL on human mammary cancer cells in vitro and in vivo were suggested. The effects of PRL are mediated by its receptor, and alterations in receptor expression are likely to play a role in tumor development. Currently, not much data is available about prolactin receptor (PRLR) expression in canine mammary tumors. To set the basis for investigations on the role of PRL in mammary tumorigenesis in this species, prolactin receptor expression was evaluated by semi-quantitative real time PCR and immunohistochemistry on 10 formalin-fixed, paraffin-embedded samples each of canine non-neoplastic mammary tissue, mammary adenomas and adenocarcinomas. Results The highest PRLR expression levels were found in normal mammary tissue, while adenomas, and to an even higher degree adenocarcinomas, showed a significant decrease in prolactin receptor expression. Compared to normal tissue, PRLR mRNA was reduced 2.4 fold (p = 0.0261) in adenomas and 4.8 fold (p = 0.008) in adenocarcinomas. PRLR mRNA expression was significantly lower in malignant than in benign lesions (p = 0.0165). Immunohistochemistry demonstrated PRLR expression in all three tissue types with signals mostly limited to epithelial cells. Conclusions Malignant transformation of mammary tissue was associated with a decline in prolactin receptor expression. Further studies are warranted to address the functional significance of this finding. PMID:22647582

  3. Rabies, canine distemper, and canine parvovirus exposure in large carnivore communities from two Zambian ecosystems.

    PubMed

    Berentsen, Are R; Dunbar, Mike R; Becker, Matthew S; M'soka, Jassiel; Droge, Egil; Sakuya, Nicholas M; Matandiko, Wigganson; McRobb, Rachel; Hanlon, Cathleen A

    2013-09-01

    Disease transmission within and among wild and domestic carnivores can have significant impacts on populations, particularly for threatened and endangered species. We used serology to evaluate potential exposure to rabies virus, canine distemper virus (CDV), and canine parvovirus (CPV) for populations of African lions (Panthera leo), African wild dogs (Lycaon pictus), and spotted hyenas (Crocuta crocuta) in Zambia's South Luangwa National Park (SLNP) and Liuwa Plain National Park (LPNP) as well as community lands bordering these areas. In addition, domestic dogs in the study region were evaluated for exposure to CDV and rabies. We provide the first comprehensive disease exposure data for these species in these ecosystems. Twenty-one lions, 20 hyenas, 13 wild dogs, and 38 domestic dogs were sampled across both regions from 2009 to 2011. Laboratory results show 10.5% of domestic dogs, 5.0% of hyenas, and 7.7% of wild dogs sampled were positive for CDV exposure. All lions were negative. Exposure to CPV was 10.0% and 4.8% for hyenas and lions, respectively. All wild dogs were negative, and domestic dogs were not tested due to insufficient serum samples. All species sampled were negative for rabies virus neutralizing antibodies except lions. Forty percent of lions tested positive for rabies virus neutralizing antibodies. Because these lions appeared clinically healthy, this finding is consistent with seroconversion following exposure to rabies antigen. To our knowledge, this finding represents the first ever documentation of rabies virus neutralizing antibodies consistent with rabies exposure that did not lead to clinical disease in free-ranging African lions from this region. With ever-increasing human pressure on these ecosystems, understanding disease transmission dynamics is essential for proper management and conservation of these carnivore species. PMID:23805791

  4. Neutron polarizers based on polarized ^3He

    NASA Astrophysics Data System (ADS)

    Gentile, T. R.; Jones, G. L.; Thompson, A. K.; Fei, X.; Keith, C. D.; Rich, D.; Snow, W. M.; Penttila, S.

    1997-10-01

    Research is underway at NIST, Indiana Univ., and LANL to develop neutron polarizers and analyzers based on polarized ^3He. Such devices, which rely on the strong spin dependence of the neutron capture cross section by polarized ^3He, have applications in weak interaction physics and materials science. In addition, the technology for polarized ^3He production is directly applicable to polarized gas MRI of lungs, and polarized targets. Our program, which includes both the spin-exchange and metastability-exchange optical pumping methods, will be reviewed. Spin-exchange has been used to analyze a polarized cold neutron beam at NIST, and also for lung imaging in collaboration with the Univ. of Pennsylvania. In the metastable method, the ^3He is polarized at low pressure, and must be substantially compressed. A piston compressor has been designed for this goal at Indiana Univ. and is under construction. At NIST we have compressed polarized gas using an apparatus that is based on a modified commercial diaphragm pump.

  5. Computed tomographic anatomy of the canine inner and middle ear.

    PubMed

    Russo, Marco; Covelli, Eugenio M; Meomartino, Leonardo; Lamb, Christopher R; Brunetti, Arturo

    2002-01-01

    A series of high-resolution computed x-ray tomography (CT) images of the normal canine middle and inner ear are presented to serve as a reference for optimal interpretation of clinical CT images of animals with diseases affecting this region. PMID:11866039

  6. Canine Gouging: A Taboo Resurfacing in Migrant Urban Population

    PubMed Central

    Noman, Anila Virani; Wong, Ferranti; Pawar, Ravikiran Ramakrishna

    2015-01-01

    Cosmopolitan cities have become a pool of migrants from different parts of the world, who carry their cultural beliefs and superstitions with them around the globe. Canine gouging is a kind of infant oral mutilation (IOM) which is widely practiced among rural population of Africa where the primary tooth bud of the deciduous canine is enucleated. The belief is that the life threatening illnesses in children like vomiting, diarrhoea, and fevers are caused by worms which infest on tooth buds. This case report is of a 15-year-old Somalian born boy, who presented at the dental institute with intermittent pain in his lower right permanent canine which was associated with a discharging intra oral buccal sinus. The tooth was endodontically treated and then restored with composite. General dental practitioners need to be vigilant when encountered with tooth presenting unusual morphology, unilateral missing tooth, and shift in the midline due to early loss of deciduous/permanent canines. Identification of any such dental mutilation practice will need further counselling of the individual and family members. It is the duty of every dental professional to educate and safeguard the oral and dental health of general public. PMID:26266057

  7. Characterization of canine dental pulp cells and their neuroregenerative potential.

    PubMed

    Naito, Eiji; Kudo, Daichi; Sekine, Shin-ichiro; Watanabe, Kazuhiro; Kobatake, Yui; Tamaoki, Naritaka; Inden, Masatoshi; Iida, Kazuki; Ito, Yusuke; Hozumi, Isao; Shibata, Toshiyuki; Maeda, Sadatoshi; Kamishina, Hiroaki

    2015-11-01

    Dental pulp cells (DPCs) of various species have been studied for their potentials of differentiation into functional neurons and secretion of neurotrophic factors. In canine, DPCs have only been studied for cell surface markers and differentiation, but there is little direct evidence for therapeutic potentials for neurological disorders. The present study aimed to further characterize canine DPCs (cDPCs), particularly focusing on their neuroregenerative potentials. It was also reported that superparamagnetic iron oxide (SPIO) particles were useful for labeling of MSCs and tracking with magnetic resonance imaging (MRI). Our data suggested that cDPCs hold higher proliferation capacity than bone marrow stromal cells, the other type of mesenchymal stem cells which have been the target of intensive research. Canine DPCs constitutively expressed neural markers, suggesting a close relationship to the nervous system in their developmental origin. Canine DPCs promoted neuritogenesis of PC12 cells, most likely through secretion of neurotrophic factors. Furthermore, SPIO nanoparticles could be effectively transported to cDPCs without significant cytotoxicity and unfavorable effects on neuritogenesis. SPIO-labeled cDPCs embedded in agarose spinal cord phantoms were successfully visualized with a magnetic resonance imaging arousing a hope for noninvasive cell tracking in transplantation studies. PMID:26170225

  8. When Reading Gets Ruff: Canine-Assisted Reading Programs

    ERIC Educational Resources Information Center

    Lane, Holly B.; Zavada, Shannon D. W.

    2013-01-01

    Canine-assisted reading programs show promise as an innovative method for engaging reluctant readers and motivating them to practice. In such programs, specially trained dogs visit classrooms and libraries, and children read to them. Children who struggle with reading may be motivated to read more because they find dogs to be calming and…

  9. Canine Gouging: A Taboo Resurfacing in Migrant Urban Population.

    PubMed

    Noman, Anila Virani; Wong, Ferranti; Pawar, Ravikiran Ramakrishna

    2015-01-01

    Cosmopolitan cities have become a pool of migrants from different parts of the world, who carry their cultural beliefs and superstitions with them around the globe. Canine gouging is a kind of infant oral mutilation (IOM) which is widely practiced among rural population of Africa where the primary tooth bud of the deciduous canine is enucleated. The belief is that the life threatening illnesses in children like vomiting, diarrhoea, and fevers are caused by worms which infest on tooth buds. This case report is of a 15-year-old Somalian born boy, who presented at the dental institute with intermittent pain in his lower right permanent canine which was associated with a discharging intra oral buccal sinus. The tooth was endodontically treated and then restored with composite. General dental practitioners need to be vigilant when encountered with tooth presenting unusual morphology, unilateral missing tooth, and shift in the midline due to early loss of deciduous/permanent canines. Identification of any such dental mutilation practice will need further counselling of the individual and family members. It is the duty of every dental professional to educate and safeguard the oral and dental health of general public. PMID:26266057

  10. Canine brachycephalic airway syndrome: pathophysiology, diagnosis, and nonsurgical management.

    PubMed

    Trappler, Michelle; Moore, Kenneth

    2011-05-01

    Canine brachycephalic airway syndrome is a progressive disease that affects many brachycephalic dogs. This article describes the components of this syndrome and focuses on acute emergency management and long-term conservative management of these patients. Surgical management is described in a companion article. PMID:21870353

  11. Dilated Canine Hearts: A Specimen for Teaching Cardiac Anatomy

    ERIC Educational Resources Information Center

    Cope, Lee Anne

    2008-01-01

    Dilated canine hearts were used to teach undergraduate students internal and external cardiac anatomy. The specimens were dilated using hydrostatic pressure and then fixed using 5% formalin. These specimens provided the students with an alternative to prepackaged embalmed hearts and anatomical models for studying the external and internal cardiac…

  12. Selection against canine hip dysplasia: success or failure?

    PubMed

    Wilson, Bethany; Nicholas, Frank W; Thomson, Peter C

    2011-08-01

    Canine hip dysplasia (CHD) is a multifactorial skeletal disorder which is very common in pedigree dogs and represents a huge concern for canine welfare. Control schemes based on selective breeding have been in operation for decades. The aim of these schemes is to reduce the impact of CHD on canine welfare by selecting for reduced radiographic evidence of CHD pathology as assessed by a variety of phenotypes. There is less information regarding the genotypic correlation between these phenotypes and the impact of CHD on canine welfare. Although the phenotypes chosen as the basis for these control schemes have displayed heritable phenotypic variation in many studies, success in achieving improvement in the phenotypes has been mixed. There is significant room for improvement in the current schemes through the use of estimated breeding values (EBVs), which can combine a dog's CHD phenotype with CHD phenotypes of relatives, other phenotypes as they are proven to be genetically correlated with CHD (especially elbow dysplasia phenotypes), and information from genetic tests for population-relevant DNA markers, as such tests become available. Additionally, breed clubs should be encouraged and assisted to formulate rational, evidenced-based breeding recommendations for CHD which suit their individual circumstances and dynamically to adjust the breeding recommendations based on continuous tracking of CHD genetic trends. These improvements can assist in safely and effectively reducing the impact of CHD on pedigree dog welfare. PMID:21727013

  13. Immunohistochemical vascular factor expression in canine inflammatory mammary carcinoma.

    PubMed

    Camacho, L; Peña, L; Gil, A González; Martín-Ruiz, A; Dunner, S; Illera, J C

    2014-07-01

    Human inflammatory breast carcinoma (IBC) and canine inflammatory mammary carcinoma (IMC) are considered the most malignant types of breast cancer. IMC has similar characteristics to IBC; hence, IMC has been suggested as a model to study the human disease. To compare the angiogenic and angioinvasive features of IMC with non-IMC, 3 canine mammary tumor xenograft models in female SCID mice were developed: IMC, comedocarcinoma, and osteosarcoma. Histopathological and immunohistochemical characterization of both primary canine tumors and xenografts using cellular markers pancytokeratin, cytokeratin 14, vimentin, and α-smooth muscle actin and vascular factors (VEGF-A, VEGF-D, VEGFR-3, and COX-2) was performed. Tumor cell proliferation index was measured by the Ki-67 marker. The xenograft models reproduced histological features found in the primary canine tumor and preserved the original immunophenotype. IMC xenografts showed a high invasive character with tumor emboli in the dermis, edema, and occasional observations of ulceration. In addition, compared with osteosarcoma and comedocarcinoma, the IMC model showed the highest vascular factor expression associated with a high proliferation index. Likewise, IMC xenografts showed higher COX-2 expression associated with VEGF-D and VEGFR-3, as well as a higher presence of dermal lymphatic tumor emboli, suggesting COX-2 participation in IMC lymphangiogenesis. These results provide additional evidence to consider vascular factors, their receptors, and COX-2 as therapeutic targets for IBC. PMID:24048323

  14. Expression of cytokeratins in the epithelium of canine odontogenic tumours.

    PubMed

    Arzi, B; Murphy, B; Nemec, A; Vapniarsky, N; Naydan, D K; Verstraete, F J M

    2011-11-01

    Odontogenic tumours are considered to be relatively rare; however, several histologically distinct types have been identified in dogs. The more common canine odontogenic tumours are peripheral odontogenic fibroma and canine acanthomatous ameloblastoma. The expression of cytokeratins (CKs) has been established for the human dental germ and odontogenic tumours. The aim of the present study was to describe the immunohistochemical expression of a panel of CKs in the epithelium of the canine dental germ, normal gingiva and odontogenic tumours arising in this species. Samples from 20 odontogenic tumours, 12 tooth germs and three normal gingival tissues were obtained. Each sample was stained with haematoxylin and eosin and subjected to immunohistochemistry for CK expression. The typical expression pattern of CKs in the odontogenic epithelium and gingiva of dogs was CK14 and CK5/6. CKs 7, 8, 18 and 20 were generally absent from the canine dental germ, gingiva and odontogenic tumours. Dogs and man therefore exhibit similar CK expression in the odontogenic epithelium. PMID:21511272

  15. Migration of canine neutrophils to chitin and chitosan.

    PubMed

    Usami, Y; Okamoto, Y; Minami, S; Matsuhashi, A; Kumazawa, N H; Tanioka, S; Shigemasa, Y

    1994-12-01

    Suspension of chitin and chitosan particles (mean size of 1 micron) were found to attract canine neutrophils chemotactically as determined by a checkerboard assay through polycarbonate filter with 5 microns pore size in Blind well chamber. Suspension of chitin induced chemokinetic migrations of the neutrophils. These evidences might reflect accumulation of neutrophils to chitin- and chitosan-implanted regions in dogs. PMID:7696425

  16. Molecular Characterization of Canine Rabies Virus, Mali, 2006–2013

    PubMed Central

    Traoré, Abdallah; Picard-Meyer, Evelyne; Mauti, Stephanie; Biarnais, Melanie; Balmer, Oliver; Samaké, Kassim; Kamissoko, Badian; Tembely, Saïdou; Sery, Amadou; Traoré, Abdel K.; Coulibaly, Amy P.; Robardet, Emmanuelle; Zinsstag, Jakob

    2016-01-01

    We genetically characterized 32 canine rabies viruses isolated in Mali during 2006–2013 and identified 3 subgroups that belonged to the Africa 2 lineage. We also detected subgroup F rabies virus. This information should be useful for development of mass vaccination campaigns for dogs and eventual large-scale control programs in this country. PMID:27089307

  17. Molecular Characterization of Canine Rabies Virus, Mali, 2006-2013.

    PubMed

    Traoré, Abdallah; Picard-Meyer, Evelyne; Mauti, Stephanie; Biarnais, Melanie; Balmer, Oliver; Samaké, Kassim; Kamissoko, Badian; Tembely, Saïdou; Sery, Amadou; Traoré, Abdel K; Coulibaly, Amy P; Robardet, Emmanuelle; Zinsstag, Jakob; Cliquet, Florence

    2016-05-01

    We genetically characterized 32 canine rabies viruses isolated in Mali during 2006-2013 and identified 3 subgroups that belonged to the Africa 2 lineage. We also detected subgroup F rabies virus. This information should be useful for development of mass vaccination campaigns for dogs and eventual large-scale control programs in this country. PMID:27089307

  18. The Human-Canine Bond: Closer than Family Ties?

    ERIC Educational Resources Information Center

    Barker, Sandra B.; Barker, Randolph T.

    1988-01-01

    Used Family Life Space Diagram to compare relationship between human family members with the human-canine relationship. Subjects were 29 dog enthusiasts, 66 typical pet owners, and 27 elementary school students with dogs. Results suggest that individuals may perceive their relationship with their pet dog as being as close as their relationship…

  19. Associations between neurologic dysfunction and lesions in canine fucosidosis.

    PubMed

    Fletcher, J L; Taylor, R M

    2016-04-01

    Canine fucosidosis in English Springer spaniels is the only animal model of the neurovisceral lysosomal storage disease fucosidosis available for preclinical therapeutic trials. For this reason, it is crucial to identify critical time points in disease progression, and if there are particular lesions associated with specific aspects of neurologic dysfunction. Historical records of 53 canine fucosidosis cases from 1979 to 2009 containing a neurologic dysfunction score assessing motor, behavioral and sensory dysfunction were interrogated by statistical analysis. Motor and behavioral dysfunction scores assessing gait deficits and apprehensive behavior first significantly increased at 12-17 months, and increased at each 6-month interval thereafter. Sensory dysfunction scores, assessing hearing loss, balance and vision deterioration, did not significantly increase until 18-23 months, and coincided with a rapid decline in neurologic function. Regression analysis incorporating published neuropathology data, measured by image analysis, identified neuroinflammation and apoptotic cell death as significant informative predictors of increasing neurologic dysfunction. These findings indicate that the level of neuropathology required to induce consistent and conspicuous clinical signs in canine fucosidosis is reached by approximately 12 months of age in the absence of other disease processes. Significant association between neuroinflammation and apoptotic cell death also suggests that specifically targeting these lesions combined with enzyme replacement in future studies may reduce disease burden in fucosidosis. Overall, examining this historical clinical data to identify associations between the extent of neuropathology and degree of clinical dysfunction provides a useful reference tool for monitoring disease and evaluating therapeutic trials conducted in canine fucosidosis. PMID:26711085

  20. The blood supply to the canine middle ear.

    PubMed

    Stevens-Sparks, Cathryn; Jarrett, Keith; McNulty, Margaret A; Strain, George M

    2016-07-01

    Current descriptions of the anatomy of the blood supply to the canine middle ear are either incomplete or inconsistent, particularly in regards to the vascular branches in close proximity to the temporomandibular articulation (TMJ). To further investigate this blood supply, dissections (n = 9), corrosion casts (n = 4), and computed tomography (n = 8) of canine temporal regions/ears were performed. The goal of this study was to identify and describe branches of the external carotid and maxillary arteries in close proximity to the TMJ that supply the middle ear of the dog. Specific focus was placed on the constancy and origin of the canine rostral tympanic artery since this artery was anticipated to arise from the maxillary artery and enter a foramen at the medial aspect of the mandibular fossa adjacent to the TMJ. New anatomical variations of three canine arteries are described in this study. (1) The rostral tympanic artery is a branch of the temporomandibular ramus and is accommodated by a small foramen located within a depression medial to the temporomandibular joint. (2) A pharyngeal branch of the caudal deep temporal artery was identified. (3) The origin of the caudal auricular artery occurred opposite the lingual artery in 25.8% of dissected specimens, contrary to published descriptions. Anat Rec, 299:907-917, 2016. © 2016 Wiley Periodicals, Inc. PMID:27082971

  1. The oncolytic effects of reovirus in canine solid tumor cell lines

    PubMed Central

    IGASE, Masaya; HWANG, Chung Chew; COFFEY, Matt; OKUDA, Masaru; NOGUCHI, Shunsuke; MIZUNO, Takuya

    2015-01-01

    Oncolytic virotherapy is a new strategy for cancer treatment for humans and dogs. Reovirus has been proven to be a potent oncolytic virus in human medicine. Our laboratory has previously reported that canine mast cell tumor and canine lymphoma were susceptible to reovirus. In this study, canine solid tumor cell lines (mammary gland tumor, osteosarcoma and malignant melanoma) were tested to determine their susceptibility towards reovirus. We demonstrated that reovirus induces more than 50% cell death in three canine mammary gland tumors and one canine malignant melanoma cell line. The reovirus-induced cell death occurred via the activation of caspase 3. Ras activation has been shown to be one of the important mechanisms of reovirus-susceptibility in human cancers. However, Ras activation was not related to the reovirus-susceptibility in canine solid tumor cell lines, which was similar to reports in canine mast cell tumor and canine lymphoma. The results of this study highly suggest that canine mammary gland tumor and canine malignant melanoma are also potential candidates for reovirus therapy in veterinary oncology. PMID:25648933

  2. The oncolytic effects of reovirus in canine solid tumor cell lines.

    PubMed

    Igase, Masaya; Hwang, Chung Chew; Coffey, Matt; Okuda, Masaru; Noguchi, Shunsuke; Mizuno, Takuya

    2015-05-01

    Oncolytic virotherapy is a new strategy for cancer treatment for humans and dogs. Reovirus has been proven to be a potent oncolytic virus in human medicine. Our laboratory has previously reported that canine mast cell tumor and canine lymphoma were susceptible to reovirus. In this study, canine solid tumor cell lines (mammary gland tumor, osteosarcoma and malignant melanoma) were tested to determine their susceptibility towards reovirus. We demonstrated that reovirus induces more than 50% cell death in three canine mammary gland tumors and one canine malignant melanoma cell line. The reovirus-induced cell death occurred via the activation of caspase 3. Ras activation has been shown to be one of the important mechanisms of reovirus-susceptibility in human cancers. However, Ras activation was not related to the reovirus-susceptibility in canine solid tumor cell lines, which was similar to reports in canine mast cell tumor and canine lymphoma. The results of this study highly suggest that canine mammary gland tumor and canine malignant melanoma are also potential candidates for reovirus therapy in veterinary oncology. PMID:25648933

  3. Colorectal cancer screening with odour material by canine scent detection

    PubMed Central

    Kohnoe, Shunji; Yamazato, Tetsuro; Satoh, Yuji; Morizono, Gouki; Shikata, Kentaro; Morita, Makoto; Watanabe, Akihiro; Morita, Masaru; Kakeji, Yoshihiro; Inoue, Fumio; Maehara, Yoshihiko

    2011-01-01

    Objective Early detection and early treatment are of vital importance to the successful treatment of various cancers. The development of a novel screening method that is as economical and non-invasive as the faecal occult blood test (FOBT) for early detection of colorectal cancer (CRC) is needed. A study was undertaken using canine scent detection to determine whether odour material can become an effective tool in CRC screening. Design Exhaled breath and watery stool samples were obtained from patients with CRC and from healthy controls prior to colonoscopy. Each test group consisted of one sample from a patient with CRC and four control samples from volunteers without cancer. These five samples were randomly and separately placed into five boxes. A Labrador retriever specially trained in scent detection of cancer and a handler cooperated in the tests. The dog first smelled a standard breath sample from a patient with CRC, then smelled each sample station and sat down in front of the station in which a cancer scent was detected. Results 33 and 37 groups of breath and watery stool samples, respectively, were tested. Among patients with CRC and controls, the sensitivity of canine scent detection of breath samples compared with conventional diagnosis by colonoscopy was 0.91 and the specificity was 0.99. The sensitivity of canine scent detection of stool samples was 0.97 and the specificity was 0.99. The accuracy of canine scent detection was high even for early cancer. Canine scent detection was not confounded by current smoking, benign colorectal disease or inflammatory disease. Conclusions This study shows that a specific cancer scent does indeed exist and that cancer-specific chemical compounds may be circulating throughout the body. These odour materials may become effective tools in CRC screening. In the future, studies designed to identify cancer-specific volatile organic compounds will be important for the development of new methods for early detection of CRC

  4. Migrastatin Analogues Inhibit Canine Mammary Cancer Cell Migration and Invasion

    PubMed Central

    Majchrzak, Kinga; Lo Re, Daniele; Gajewska, Małgorzata; Bulkowska, Małgorzata; Homa, Agata; Pawłowski, Karol; Motyl, Tomasz; Murphy, Paul V.; Król, Magdalena

    2013-01-01

    Background Cancer spread to other organs is the main cause of death of oncological patients. Migration of cancer cells from a primary tumour is the crucial step in the complex process of metastasis, therefore blocking this process is currently the main treatment strategy. Metastasis inhibitors derived from natural products, such as, migrastatin, are very promising anticancer agents. Thus, the aim of our study was to investigate the effect of six migrastatin analogues (MGSTA-1 to 6) on migration and invasion of canine mammary adenocarcinoma cell lines isolated from primary tumours and their metastases to the lungs. Canine mammary tumours constitute a valuable tool for studying multiple aspect of human cancer. Results Our results showed that two of six fully synthetic analogues of migrastatin: MGSTA-5 and MGSTA-6 were potent inhibitors of canine mammary cancer cells migration and invasion. These data were obtained using the wound healing test, as well as trans-well migration and invasion assays. Furthermore, the treatment of cancer cells with the most effective compound (MGSTA-6) disturbed binding between filamentous F-actin and fascin1. Confocal microscopy analyses revealed that treatment with MGSTA-6 increased the presence of unbound fascin1 and reduced co-localization of F-actin and fascin1 in canine cancer cells. Most likely, actin filaments were not cross-linked by fascin1 and did not generate the typical filopodial architecture of actin filaments in response to the activity of MGSTA-6. Thus, administration of MGSTA-6 results in decreased formation of filopodia protrusions and stress fibres in canine mammary cancer cells, causing inhibition of cancer migration and invasion. Conclusion Two synthetic migrastatin analogues (MGSTA-5 and MGSTA-6) were shown to be promising compounds for inhibition of cancer metastasis. They may have beneficial therapeutic effects in cancer therapy in dogs, especially in combination with other anticancer drugs. However, further in

  5. Current state of knowledge: the canine gastrointestinal microbiome.

    PubMed

    Hooda, Seema; Minamoto, Yasushi; Suchodolski, Jan S; Swanson, Kelly S

    2012-06-01

    Gastrointestinal (GI) microbes have important roles in the nutritional, immunological, and physiologic processes of the host. Traditional cultivation techniques have revealed bacterial density ranges from 10(4) to 10(5) colony forming units (CFU)/g in the stomach, from 10(5) to 10(7) CFU/g in the small intestine, and from 10(9) to 10(11) CFU/g in the colon of healthy dogs. As a small number of bacterial species can be grown and studied in culture, however, progress was limited until the recent emergence of DNA-based techniques. In recent years, DNA sequencing technology and bioinformatics have allowed for better phylogenetic and functional/metabolic characterization of the canine gut microbiome. Predominant phyla include Firmicutes, Bacteroidetes, Fusobacteria, Proteobacteria, and Actinobacteria. Studies using 16S ribosomal RNA (rRNA) gene pyrosequencing have demonstrated spatial differences along the GI tract and among microbes adhered to the GI mucosa compared to those in intestinal contents or feces. Similar to humans, GI microbiome dysbiosis is common in canine GI diseases such as chronic diarrhea and inflammatory bowel diseases. DNA-based assays have also identified key pathogens contributing to such conditions, including various Clostridium, Campylobacter, Salmonella, and Escherichia spp. Moreover, nutritionists have applied DNA-based techniques to study the effects of dietary interventions such as dietary fiber, prebiotics, and probiotics on the canine GI microbiome and associated health indices. Despite recent advances in the field, the canine GI microbiome is far from being fully characterized and a deeper characterization of the phylogenetic and functional/metabolic capacity of the GI microbiome in health and disease is needed. This paper provides an overview of recent studies performed to characterize the canine GI microbiome. PMID:22647637

  6. A canine minidystrophin is functional and therapeutic in mdx mice.

    PubMed

    Wang, B; Li, J; Qiao, C; Chen, C; Hu, P; Zhu, X; Zhou, L; Bogan, J; Kornegay, J; Xiao, X

    2008-08-01

    Duchenne muscular dystrophy (DMD) is the most common and lethal genetic muscle disorder lacking a curative treatment. We wish to use the dystrophin-deficient golden retriever muscular dystrophy (GRMD) dog, a canine model of DMD, to investigate adeno-associated virus (AAV) vector-mediated minidystrophin gene therapy. The dog model is useful in evaluating vector dose requirement and immunological consequences owing to its large size and outbred nature. In this study, we have cloned and constructed a canine minidystrophin gene vector. Owing to limited availability of the GRMD dogs, here we first examined the functions and therapeutic effects of the canine minidystrophin in the mdx mouse model. We observed efficient minigene expression without cellular immune responses in mdx mice after AAV1-cMinidys vector intramuscular injection. We also observed restoration of the missing dystrophin-associated protein complex (DPC) onto the sarcolemma, including sarcoglycans and dystrobrevin, and a partial restoration of alpha-syntrophin and neural nitric oxide synthase (nNOS). In addition, minidystrophin treatment ameliorated dystrophic pathology, such as fibrosis and myofiber central nucleation (CN). CN remained minimal (<2%) after AAV injection in the neonatal mdx mice and was reduced from more than 75% to about 25% after AAV injection in adult mdx mice. Finally, in vivo cell membrane leakage test with Evans blue dye showed that the canine minidystrophin could effectively protect the myofiber plasma membrane integrity. Our results, thus, demonstrated the functionality and therapeutic potential of the canine minidystrophin and paved its way for further testing in the GRMD dog model. PMID:18432277

  7. Establishment of Canine-Derived Giardia duodenalis Isolates in Culture.

    PubMed

    Tysnes, Kristoffer R; Robertson, Lucy J

    2016-06-01

    Researchers continue to rely on axenic cultivation of Giardia duodenalis trophozoites in vitro to study the life cycle and host-parasite interactions of G. duodenalis and to develop vaccines and drugs to prevent and treat giardiasis. The majority of in vitro studies of G. duodenalis have used a small subset of isolates, mostly of assemblage A, and these isolates are usually originally isolated from humans. The most commonly used isolate for lab studies is known as WB. Canine giardiasis is a disease of veterinary importance, but it may also be of relevance in zoonotic transmission. Few G. duodenalis isolates from dogs have been adapted to in vitro culture, probably because the methods used are not suitable for the canine-specific genotypes that tend to dominate in most dog populations. In the current study, an experimental approach to cultivating canine-derived isolates of G. duodenalis was attempted by modification of the standard protocol based on physiological differences between the human and canine digestive system. An adapted method is described for improving the rate of in vitro excystation of cysts isolated from dogs by chemically weakening the cyst wall. A new canine-derived assemblage A G. duodenalis isolate was successfully adapted to axenic culture by using this method; the dog apparently had a mixed infection of assemblages A and D, but the assemblage A successfully outcompeted the assemblage D under conditions of in vitro culture. Based on the results, reasons regarding why humans do not seem to be suitable hosts for G. duodenalis in assemblages C and D are discussed. PMID:26835750

  8. Headspace concentrations of explosive vapors in containers designed for canine testing and training: theory, experiment, and canine trials.

    PubMed

    Lotspeich, Erica; Kitts, Kelley; Goodpaster, John

    2012-07-10

    It is a common misconception that the amount of explosive is the chief contributor to the quantity of vapor that is available to trained canines. In fact, this quantity (known as odor availability) depends not only on the amount of explosive material, but also the container volume, explosive vapor pressure and temperature. In order to better understand odor availability, headspace experiments were conducted and the results were compared to theory. The vapor-phase concentrations of three liquid explosives (nitromethane, nitroethane and nitropropane) were predicted using the Ideal Gas Law for containers of various volumes that are in use for canine testing. These predictions were verified through experiments that varied the amount of sample, the container size, and the temperature. These results demonstrated that the amount of sample that is needed to saturate different sized containers is small, predictable and agrees well with theory. In general, and as expected, once the headspace of a container is saturated, any subsequent increase in sample volume will not result in the release of more vapors. The ability of canines to recognize and alert to differing amounts of nitromethane has also been studied. In particular, it was found that the response of trained canines is independent of the amount of nitromethane present, provided it is a sufficient quantity to saturate the container in which it is held. PMID:22421324

  9. Characterization of the canine mda-7 gene, transcripts and expression patterns

    PubMed Central

    Sandey, Maninder; Bird, R. Curtis; Das, Swadesh K.; Sarkar, Devanand; Curiel, David T.; Fisher, Paul B.; Smith, Bruce F.

    2014-01-01

    Human melanoma differentiation associated gene-7/interleukin-24 (mda-7/IL-24) displays potent growth suppressing and cell killing activity against a wide variety of human and rodent cancer cells. In this study, we identified a canine ortholog of the human mda-7/IL-24 gene located within a cluster of IL-10 family members on chromosome 7. The full-length mRNA sequence of canine mda-7 was determined, which encodes a 186-amino acid protein that has 66% similarity to human MDA-7/IL-24. Canine MDA-7 is constitutively expressed in cultured normal canine epidermal keratinocytes (NCEKs), and its expression levels are increased after lipopolysaccharide stimulation. In cultured NCEKs, the canine mda-7 pre-mRNA is differentially spliced, via exon skipping and alternate 5′-splice donor sites, to yield five splice variants (canine mda-7sv1, canine mda-7sv2, canine mda-7sv3, canine mda-7sv4 and canine mda-7sv5) that encode four protein isoforms of the canine MDA-7 protein. These protein isoforms have a conserved N-terminus (signal peptide sequence) and are dissimilar in amino acid sequences at their C-terminus. Canine MDA-7 is not expressed in primary canine tumor samples, and most tumor derived cancer cell lines tested, like its human counterpart. Unlike human MDA-7/IL-24, canine mda-7 mRNA is not expressed in unstimulated or lipopolysaccharide (LPS), concanavalin A (ConA) or phytohemagglutinin (PHA) stimulated canine peripheral blood mononuclear cells (PBMCs). Furthermore, in-silico analysis revealed that canonical canine MDA-7 has a potential 28 amino acid signal peptide sequence that can target it for active secretion. This data suggests that canine mda-7 is indeed an ortholog of human mda-7/IL-24, its protein product has high amino acid similarity to human MDA-7/IL-24 protein and it may possess similar biological properties to human MDA-7/IL-24, but its expression pattern is more restricted than its human ortholog. PMID:24865935

  10. The cloning and expression of matrix metalloproteinase-2 and tissue inhibitor of matrix metalloproteinase 2 in normal canine lymph nodes and in canine lymphoma.

    PubMed

    Newman, R G; Kitchell, B E; Wallig, M A; Paria, B

    2008-04-01

    Matrix metalloproteinase-2 (MMP-2) and its inhibitor, tissue inhibitor of matrix metalloproteinase 2 (TIMP2), are known to be important in cancer. The purposes of this study were to determine the cDNA sequence of canine MMP-2 and to investigate the expression patterns of MMP-2 and TIMP2 in normal canine lymph nodes and spontaneously arising canine lymphomas. We cloned and sequenced a PCR product containing most (1901 base pairs) of the coding sequence of canine MMP-2 that translates into a 623 amino acid protein. The cDNA and deduced amino acid sequences are highly homologous to those of other mammalian species. Canine MMP-2 and TIMP2 mRNAs were detectable in the majority of normal lymph node and lymphomatous samples evaluated. No statistical difference was identified when comparing the expression of either gene with regard to normal versus neoplastic nodes, nodal versus extranodal lymphoma, lymphoma grade, or B versus T cell immunophenotype. PMID:17604063

  11. Distribution and activity levels of matrix metalloproteinase 2 and 9 in canine and feline osteosarcoma.

    PubMed

    Gebhard, Christiane; Fuchs-Baumgartinger, Andrea; Razzazi-Fazeli, Ebrahim; Miller, Ingrid; Walter, Ingrid

    2016-01-01

    Overexpression of matrix metalloproteinases (MMPs) has been associated with increased tumor aggressiveness and metastasis dissemination. We investigated whether the contrasting metastatic behavior of feline and canine osteosarcoma is related to levels and activities of MMP2 and MMP9. Zymography and immunohistochemistry were used to determine expression levels of MMP2 and MMP9 in canine and feline osteosarcoma. Using immunohistochemistry, increased MMP9 levels were identified in most canine osteosarcomas, whereas cat samples more often displayed moderate levels. High levels of pro-MMP9, pro-MMP2, and active MMP2 were detected by gelatin zymography in both species, with significantly higher values for active MMP2 in canine osteosarcoma. These findings indicate that MMP2 is probably involved in canine and feline osteosarcoma and their expression and activity could be associated with the different metastatic behavior of canine and feline osteosarcoma. PMID:26733734

  12. Distribution and activity levels of matrix metalloproteinase 2 and 9 in canine and feline osteosarcoma

    PubMed Central

    Gebhard, Christiane; Fuchs-Baumgartinger, Andrea; Razzazi-Fazeli, Ebrahim; Miller, Ingrid; Walter, Ingrid

    2016-01-01

    Overexpression of matrix metalloproteinases (MMPs) has been associated with increased tumor aggressiveness and metastasis dissemination. We investigated whether the contrasting metastatic behavior of feline and canine osteosarcoma is related to levels and activities of MMP2 and MMP9. Zymography and immunohistochemistry were used to determine expression levels of MMP2 and MMP9 in canine and feline osteosarcoma. Using immunohistochemistry, increased MMP9 levels were identified in most canine osteosarcomas, whereas cat samples more often displayed moderate levels. High levels of pro-MMP9, pro-MMP2, and active MMP2 were detected by gelatin zymography in both species, with significantly higher values for active MMP2 in canine osteosarcoma. These findings indicate that MMP2 is probably involved in canine and feline osteosarcoma and their expression and activity could be associated with the different metastatic behavior of canine and feline osteosarcoma. PMID:26733734

  13. Polarized Light Corridor Demonstrations.

    ERIC Educational Resources Information Center

    Davies, G. R.

    1990-01-01

    Eleven demonstrations of light polarization are presented. Each includes a brief description of the apparatus and the effect demonstrated. Illustrated are strain patterns, reflection, scattering, the Faraday Effect, interference, double refraction, the polarizing microscope, and optical activity. (CW)

  14. Polarization at SLAC

    SciTech Connect

    Woods, M.

    1995-01-01

    A highly polarized electron beam is a key feature. for the Current physics program at SLAC. An electron beam polarization of 80% can now be routinely achieved for typically 5000 hours of machine operation per year. Two main Physics programs utilize the polarized beam. Fixed target experiments in End Station A study the collision of polarized electrons with polarized nuclear targets to elucidate the spin structure of the nucleon and to provide an important test of QCD. Using the SLAC Linear Collider, collisions of polarized electrons with unpolarized positrons allow precise measurements of parity violation in the Z-fermion couplings and provide a very precise measurement of tile weak mixing angle. This paper discusses polarized beam operation at SLAC, and gives an overview of the polarized physics program.

  15. A Translational Polarization Rotator

    NASA Technical Reports Server (NTRS)

    Chuss, David T.; Wollack, Edward J.; Pisano, Giampaolo; Ackiss, Sheridan; U-Yen, Kongpop; Ng, Ming wah

    2012-01-01

    We explore a free-space polarization modulator in which a variable phase introduction between right- and left-handed circular polarization components is used to rotate the linear polarization of the outgoing beam relative to that of the incoming beam. In this device, the polarization states are separated by a circular polarizer that consists of a quarter-wave plate in combination with a wire grid. A movable mirror is positioned behind and parallel to the circular polarizer. As the polarizer-mirror distance is separated, an incident liear polarization will be rotated through an angle that is proportional to the introduced phase delay. We demonstrate a prototype device that modulates Stokes Q and U over a 20% bandwidth.

  16. Comparison of canine parvovirus with mink enteritis virus by restriction site mapping.

    PubMed Central

    McMaster, G K; Tratschin, J D; Siegl, G

    1981-01-01

    The genomes of canine parvovirus and mink enteritis virus were compared by restriction enzyme analysis of their replicative-form DNAs. Of 79 mapped sites, 68, or 86%, were found to be common for both types of DNA, indicating that canine parvovirus and mink enteritis virus are closely related viruses. Whether they evolved from a common precursor or whether canine parvovirus is derived from mink enteritis virus, however, cannot be deduced from our present data. Images PMID:6264109

  17. Polar Ozone Workshop. Abstracts

    NASA Technical Reports Server (NTRS)

    Aikin, Arthur C.

    1988-01-01

    Results of the proceedings of the Polar Ozone Workshop held in Snowmass, CO, on May 9 to 13, 1988 are given. Topics covered include ozone depletion, ozonometry, polar meteorology, polar stratospheric clouds, remote sensing of trace gases, atmospheric chemistry and dynamical simulations.

  18. Hybrid polarization control

    NASA Astrophysics Data System (ADS)

    Gray, George R.; Ibragimov, Edem; Sluz, Joseph; Sova, Raymond

    2005-05-01

    We demonstrate a novel method of polarization control that combines rotatable waveplates (angle control) and variable retarders (retardance control). Such a "hybrid" polarization controller performs far better than conventional controllers, allowing nearly perfect arbitrary-to-arbitrary polarization transformations. We show theoretically that the two control parameters augment one another because they tend to result in orthogonal movements on the Poincaré sphere.

  19. Graphing Polar Curves

    ERIC Educational Resources Information Center

    Lawes, Jonathan F.

    2013-01-01

    Graphing polar curves typically involves a combination of three traditional techniques, all of which can be time-consuming and tedious. However, an alternative method--graphing the polar function on a rectangular plane--simplifies graphing, increases student understanding of the polar coordinate system, and reinforces graphing techniques learned…

  20. Playing with Polarizers.

    ERIC Educational Resources Information Center

    Hecht, Jeff

    1991-01-01

    Discussed is how polarized sunglasses block glare, help spot subtle differences in surfaces, and give a clearer view under water. Information on unpolarized and polarized light is provided. The reasons causing glare to occur and how polarizers decrease glare are discussed. (KR)