interacts_with DB00887?
Is phentermine an inhibitor of monoamine oxidase ? A critical appraisal . DB00191 produces a spectrum of concentration-dependent biochemical effects . It interacts with NE transporters at 0.1 microM , DA transporters at about 1 microM , 5-HT transporters at 15 microM and P21397 at about 100 microM . When administered at typical anorectic doses , phentermine primarily interacts with DA and NE transporters and does not produce biochemical or neurochemical effects which would occur if it were inhibiting P21397 . Some other explanation other than MAO inhibition must be sought to explain how oral phentermine increases platelet 5-HT , since platelet P27338 does not metabolize platelet 5-HT , and since amphetamine-type drugs are even weaker inhibitors of P27338 than P21397 . Clinical studies in humans have shown that amphetamine , which is a more potent inhibitor of P21397 than phentermine , does not inhibit P21397 at therapeutic doses . Neither phentermine alone , fluoxetine alone or their combined use have been associated with cardiac valvulopathy , and clinical experience has shown their combined use to be free of significant adverse effects . Viewed collectively , there appears to be no data to support the hypothesis that phentermine inhibits MAO at typical therapeutic doses . DB02527 inhibition of murine intestinal Na/H exchange requires P13569 -mediated cell shrinkage of villus epithelium . BACKGROUND AND AIMS : Unlike the intestine of normal subjects , small-intestinal epithelia of cystic fibrosis patients and cystic fibrosis transmembrane conductance regulator protein-null ( P13569 (-) ) mice do not respond to stimulation of intracellular cyclic adenosine monophosphate with inhibition of electroneutral NaCl absorption . Because P13569 -mediated anion secretion has been associated with changes in crypt cell volume , we hypothesized that P13569 -mediated cell volume reduction in villus epithelium is required for intracellular cyclic adenosine monophosphate inhibition of Na(+)/H(+) exchanger ( primarily P48764 ) activity in the proximal small intestine . METHODS : Transepithelial (22)Na flux across the jejuna of P13569 (+) , P13569 (-) , the basolateral membrane Na(+)/K(+)/2Cl(-) co-transporter protein P55011 (+) , and P55011 (-) mice were correlated with changes in epithelial cell volume of the midvillus region . RESULTS : Stimulation of intracellular cyclic adenosine monophosphate resulted in cessation of Na(+)/H(+) exchanger-mediated Na(+) absorption ( J(ms)(NHE) ) in P13569 (+) jejunum but had no effect on J(ms)(NHE) across P13569 (-) jejunum . Cell volume indices indicated an approximately 30 % volume reduction of villus epithelial cells in P13569 (+) jejunum but no changes in P13569 (-) epithelium after intracellular cyclic adenosine monophosphate stimulation . In contrast , cell shrinkage induced by hypertonic medium inhibited J(ms)(NHE) in both P13569 (+) and P13569 (-) mice . DB00887 treatment to inhibit Cl(-) secretion by blockade of the Na(+)/K(+)/2Cl(-) co-transporter , P55011 , of stimulated P13569 (+) jejunum prevented maximal volume reduction of villus epithelium and recovered approximately 40 % of J(ms)(NHE) . Likewise , J(ms)(NHE) and cell volume were unaffected by intracellular cyclic adenosine monophosphate stimulation in P55011 (-) jejuna . CONCLUSIONS : These findings show a previously unrecognized role of functional P13569 expressed in villus epithelium : regulation of P48764 -mediated Na(+) absorption by alteration of epithelial cell volume . P13569 modulates neurosecretory function in pulmonary neuroendocrine cell-related tumor cell line models . The pulmonary neuroendocrine cell ( PNEC ) system consists of solitary cells and distinctive cell clusters termed neuroepithelial bodies ( P20929 ) localized in the airway epithelium . PNEC/ P20929 express a variety of bioactive substances , including amine ( serotonin , 5HT ) and neuropeptides . We have previously shown that P20929 cells are O(2) sensors expressing nicotinamide adenine diphosphate oxidase complex and O(2) sensitive K(+) channel . Recently , we demonstrated expression of functional cystic fibrosis transmembrane conductance regulator ( P13569 ) and Cl(-) conductances in P20929 cells of rabbit neonatal lung . Because PNEC/ P20929 are sparsely distributed and difficult to study in native lung , we investigated small-cell lung carcinoma ( SCLC ) and carcinoid tumor cell lines ( tumor counterparts of normal PNEC/ P20929 ) as models for PNEC/ P20929 . SCLC ( H146 , H345 ) and carcinoid ( H727 ) cell lines express neuroendocrine cell markers , including chromogranin A , neural cell adhesion molecule ( N- P62158 ) , 5HT , and tryptophan hydroxylase . We report that H146 , H345 , and H727 express P13569 messenger RNA ( reverse transcription polymerase chain reaction ) and protein ( immunoblotting ) and possess functional P13569 Cl(-) conductance , demonstrated by an iodide efflux assay inhibitable by transfection with antisense P13569 . Using an immunoassay to quantitate 5HT secretion , we also show that downregulation of P13569 abolishes hypoxia-induced 5HT release , and reduces secretory response to high potassium . Our findings suggest that P13569 may modulate neurosecretory activity of PNEC/ P20929 possessing O(2) sensor function . We propose that these tumor cell lines may be useful models for investigating the role of P13569 in PNEC/ P20929 functions in health and disease . DCEBIO stimulates Cl- secretion in the mouse jejunum . We investigated the effects of 5,6-dichloro-1-ethyl-1,3-dihydro-2H-benzimidazol-2-one(DCEBIO) on the Cl- secretory response of the mouse jejunum using the Ussing short-circuit current ( Isc ) technique . DCEBIO stimulated a concentration-dependent , sustained increase in Isc ( EC50 41 +/- 1 microM ) . Pretreating tissues with 0.25 microM forskolin reduced the concentration-dependent increase in Isc by DCEBIO and increased the EC50 ( 53 +/- 5 microM ) . DB00887 blocked ( 82 +/- 5 % ) the DCEBIO-stimulated Isc consistent with Cl- secretion . DCEBIO was a more potent stimulator of Cl- secretion than its parent molecule , 1-ethyl-2-benzimidazolinone . DB01016 or P16860 reduced the DCEBIO-stimulated Isc by > 80 % indicating the participation of P13569 in the DCEBIO-stimulated Isc response . Clotrimazole reduced DCEBIO-stimulated Isc by 67 +/- 15 % , suggesting the participation of the intermediate conductance Ca2+-activated K+ channel ( IKCa ) in the DCEBIO-activated Isc response . In the presence of maximum forskolin ( 10 microM ) , the DCEBIO response was reduced and biphasic , reaching a peak response of the change in Isc of 43 +/- 5 microA/cm2 and then falling to a steady-state response of 17 +/- 10 microA/cm2 compared with DCEBIO control tissues ( 61 +/- 6 microA/cm2 ) . The forskolin-stimulated Isc in the presence of DCEBIO was reduced compared with forskolin control tissues . Similar results were observed with DCEBIO and 8-BrcAMP where adenylate cyclase was bypassed . H89 , a PKA inhibitor , reduced the DCEBIO-activated Isc , providing evidence that DCEBIO increased Cl- secretion via a DB02527 /PKA-dependent manner . These data suggest that DCEBIO stimulates Cl- secretion of the mouse jejunum and that DCEBIO targets components of the Cl- secretory mechanism . P13569 mediated chloride secretion in the avian renal proximal tubule . In primary cell cultures of the avian ( Gallus gallus ) renal proximal tubule parathyroid hormone and DB02527 activation generate a Cl(-)-dependent short circuit current ( I(SC) ) response , consistent with net transepithelial Cl(-) secretion . In this study we investigated the expression and physiological function of the Na-K-2Cl ( NKCC ) transporter and P13569 chloride channel , both associated with Cl(-) secretion in a variety of tissues , in these proximal tubule cells . Using both RT-PCR and immunoblotting approaches , we showed that NKCC and P13569 are expressed , both in proximal tubule primary cultures and in a proximal tubule fraction of non-cultured ( native tissue ) fragments . We also used electrophysiological methods to assess the functional contribution of NKCC and P13569 to forskolin-activated I(SC) responses in filter grown cultured monolayers . DB00887 ( 10 μM ) , a specific blocker of NKCC , inhibited forskolin activated I(SC) by about 40 % , suggesting that basolateral uptake of Cl(-) is partially mediated by NKCC transport . In monolayers permeabilized on the basolateral side with nystatin , forskolin activated an apical Cl(-) conductance , manifested as bidirectional diffusion currents in the presence of oppositely directed Cl(-) gradients . Under these conditions the apical conductance appeared to show some bias towards apical-to-basolateral Cl(-) current . Two selective P13569 blockers , P13569 Inhibitor 172 and GlyH-101 ( both at 20 μM ) inhibited the forskolin activated diffusion currents by 38-68 % , with GlyH-101 having a greater effect . These data support the conclusion that avian renal proximal tubules utilize an apical P13569 Cl(-) channel to mediate DB02527 -activated Cl(-) secretion . Cross-talk between PKA-Cβ and p65 mediates synergistic induction of Q07343 by roflumilast and NTHi . Phosphodiesterase 4B ( Q07343 ) plays a key role in regulating inflammation . DB01656 , a phosphodiesterase (PDE)4-selective inhibitor , has recently been approved for treating severe chronic obstructive pulmonary disease ( P48444 ) patients with exacerbation . However , there is also clinical evidence suggesting the development of tachyphylaxis or tolerance on repeated dosing of roflumilast and the possible contribution of Q07343 up-regulation , which could be counterproductive for suppressing inflammation . Thus , understanding how Q07343 is up-regulated in the context of the complex pathogenesis and medications of P48444 may help improve the efficacy and possibly ameliorate the tolerance of roflumilast . Here we show that roflumilast synergizes with nontypeable Haemophilus influenzae ( NTHi ) , a major bacterial cause of P48444 exacerbation , to up-regulate PDE4B2 expression in human airway epithelial cells in vitro and in vivo . Up-regulated PDE4B2 contributes to the induction of certain important chemokines in both enzymatic activity-dependent and activity-independent manners . We also found that protein kinase A catalytic subunit β ( PKA-Cβ ) and nuclear factor-κB ( NF-κB ) p65 subunit were required for the synergistic induction of PDE4B2 . PKA-Cβ phosphorylates p65 in a DB02527 -dependent manner . Moreover , Ser276 of p65 is critical for mediating the PKA-Cβ-induced p65 phosphorylation and the synergistic induction of PDE4B2 . Collectively , our data unveil a previously unidentified mechanism underlying synergistic up-regulation of PDE4B2 via a cross-talk between PKA-Cβ and p65 and may help develop new therapeutic strategies to improve the efficacy of DB05876 inhibitor . Stimulation of Xenopus P47900 receptor activates P13569 in A6 cells . Nucleotide binding to purinergic P2Y receptors contributes to the regulation of a variety of physiological functions in renal epithelial cells . Here , we investigate the regulatory mechanism of the P47900 receptor agonist 2-methylthioadenosine diphosphate ( 2-MeSADP ) on Cl- transport in A6 cells , a commonly used model of the distal section of the Xenopus laevis nephron . Protein and mRNA expression analysis together with functional measurements demonstrated the basolateral location of the Xenopus P47900 receptor . 2-MeSADP increased intracellular [ Ca2+ ] and DB02527 and Cl- efflux , responses that were all inhibited by the specific P47900 receptor antagonist P59665 2179 . Cl- efflux was also inhibited by the cystic fibrosis transmembrane conductance regulator ( P13569 ) blocker glibenclamide . Inhibition of either protein kinase A ( PKA ) or the binding between A-kinase-anchoring proteins ( AKAPs ) and the regulatory PKA RII subunit blocked the 2-MeSADP-induced activation of P13569 , suggesting that PKA mediates P47900 receptor regulation of P13569 through one or more AKAPs . Further , the truncation of the PDZ1 domain of the scaffolding protein Na+/H+ exchanger regulatory factor-2 ( Q15599 ) inhibited 2-MeSADP-dependent stimulation of Cl- efflux , suggesting the involvement of this scaffolding protein . Activation or inhibition of PKC had no effect per se on basal Cl- efflux but potentiated or reduced the 2-MeSADP-dependent stimulation of Cl- efflux , respectively . These data suggest that the X laevis P47900 receptor in A6 cells can increase both DB02527 /PKA and Ca2+/PKC intracellular levels and that the PKC pathway is involved in P13569 activation via potentiation of the PKA pathway . Microelectrode measurements of the effects of basolateral adenosine in polarized human intestinal epithelial cells in culture . Activation of the basolateral receptor for adenosine in HT-29cl.19A cells , by 100 microM adenosine , increased the equivalent short-circuit current ( DeltaIsc= 24+/-2 microA/cm2 ) , depolarized the intracellular potential ( DeltaVa= 26+/-2 mV ) and decreased the fractional apical membrane resistance ( DeltafRa=-0.48 ) . The changes in all parameters reached their peak values simultaneously . This suggests that the primary action of the adenosine-activated pathway is on only one membrane . DB00887 inhibited the transepithelial response and repolarized the cell potential . After preincubation with 100 microM forskolin , application of 300 microM adenosine caused a significant further change in Va , Isc , the transepithelial potential ( Vt ) and fRa . Together with the results from ion-replacement studies , the observations indicate that adenosine activates channels other than the cystic fibrosis transmembrane conductance regulator ( P13569 ) . The rank order of potencies of adenosine and adenosine analogues implies that the receptor is of the A2 subtype . Preincubation with 4-bromophenacyl bromide ( 4-BPB ) inhibited the effect of an adenosine analogue by 50 % , indicating that activation of phospholipase A2 may be involved in the adenosine-induced response . [ Measurement of rifampicin and clarithromycin in serum by high-performance liquid chromatography with electrochemical detection ] . DB01045 ( RFP ) induces hepatic drug-metabolizing enzymes , making drug interactions a very important clinical problem . DB01211 ( P62158 ) metabolism is reportedly enhanced by induction of hepatic drug-metabolizing enzymes ( P08684 ) by RFP , so that the blood lend of P62158 decreases when RFP is administered concurrently . We connected an electrochemical detector to a high-performance liquid chromatograph ( HPLC ) for simple , rapid , easy measurement of blood concentrations of RFP and P62158 . Using samples of patient serum , normal serum , and reference standards , we compared HPLC by an external laboratory and the results of LC/MS/MS analysis with those of this new assay . A strong correlation was seen between our HPLC results and those of the external laboratory in RFP levels ( r=0.975 , p < 0.01 ) . A strong correlation was also seen between results we obtained for P62158 with the electrochemical detector in this assay and values measured by LC/MS/MS analysis ( r=0.995 , p < 0.01 ) . Our method enabled simple , rapid measurement of RFP and P62158 by connecting the HPLC and electrochemical detector in tandem . This system was used to modulate dosage during combined therapy with RFP and P62158 . The therapeutic effect for nontuberculous mycobacteriosis is expected to improve , and our HPLC is expected to be useful for simple , rapid , easy measurement of blood concentrations . DB00452 -arginine conjugate , a novel HIV-1 Tat antagonist : synthesis and anti-HIV activities . HIV-1 transactivating protein Tat is essential for virus replication and progression of HIV disease . HIV-1 Tat stimulates transactivation by binding to HIV-1 transactivator responsive element ( TAR ) RNA , and while secreted extracellularly , it acts as an immunosuppressor , an activator of quiescent T-cells for productive HIV-1 infection , and by binding to CXC chemokine receptor type 4 ( P61073 ) as a chemokine analogue . Here we present a novel HIV-1 Tat antagonist , a neomycin B-hexaarginine conjugate ( NeoR ) , which inhibits Tat transactivation and antagonizes Tat extracellular activities , such as increased viral production , induction of P61073 expression , suppression of CD3-activated proliferation of lymphocytes , and upregulation of the CD8 receptor . Moreover , Tat inhibits binding of fluoresceine isothiocyanate ( FITC ) -labeled NeoR to human peripheral blood mononuclear cells ( PBMC ) , indicating that Tat and NeoR bind to the same cellular target . This is further substantiated by the finding that NeoR competes with the binding of monoclonal Abs to P61073 . Furthermore , NeoR suppresses HIV-1 binding to cells . Importantly , NeoR accumulates in the cell nuclei and inhibits the replication of M- and T-tropic HIV-1 laboratory isolates ( EC(50) = 0.8-5.3 microM ) . A putative model structure for the TAR-NeoR complex , which complies with available experimental data , is presented . We conclude that NeoR is a multitarget HIV-1 inhibitor ; the structure , and molecular modeling and dynamics , suggest its binding to TAR RNA . NeoR inhibits HIV-1 binding to cells , partially by blocking the P61073 HIV-1 coreceptor , and it antagonizes Tat functions . NeoR is therefore an attractive lead compound , capable of interfering with different stages of HIV infection and AIDS pathogenesis . Multipotent stem/progenitor cells in human biliary tree give rise to hepatocytes , cholangiocytes , and pancreatic islets . Multipotent stem/progenitors are present in peribiliary glands of extrahepatic biliary trees from humans of all ages and in high numbers in hepato-pancreatic common duct , cystic duct , and hilum . They express endodermal transcription factors ( e.g. , Sox9 , Q9H6I2 , Q9Y261 , PDX1 , HES1 , Q9Y4Z2 , Q92786 ) intranuclearly , stem/progenitor surface markers ( EpCAM , P13591 , CD133 , P61073 ) , and sometimes weakly adult liver , bile duct , and pancreatic genes ( albumin , cystic fibrosis transmembrane conductance regulator [ P13569 ] , and insulin ) . They clonogenically expand on plastic and in serum-free medium , tailored for endodermal progenitors , remaining phenotypically stable as undifferentiated cells for months with a cell division initially every ≈36 hours and slowing to one every 2-3 days . Transfer into distinct culture conditions , each comprised of a specific mix of hormones and matrix components , yields either cords of hepatocytes ( express albumin , P08684 , and transferrin ) , branching ducts of cholangiocytes ( expressing anion exchanger-2- P04920 and P13569 ) , or regulatable C-peptide secreting neoislet-like clusters ( expressing glucagon , insulin ) and accompanied by changes in gene expression correlating with the adult fate . Transplantation into quiescent livers of immunocompromised mice results in functional human hepatocytes and cholangiocytes , whereas if into fat pads of streptozocin-induced diabetic mice , results in functional islets secreting glucose-regulatable human C-peptide . CONCLUSION : The phenotypes and availability from all age donors suggest that these stem/progenitors have considerable potential for regenerative therapies of liver , bile duct , and pancreatic diseases including diabetes . DNA damage and related modifier genes in Italian cystic fibrosis patients . Cystic Fibrosis ( CF ) is an autosomal recessive multisystemic disorder showing a highly heterogeneous phenotype , even among siblings carrying identical P13569 mutations . Moreover , oxidative stress is of central importance in the pathogenesis of cystic fibrosis . The present study seeks to value the presence of oxidative damage in CF patients and the possible modifier effect of repair and glutathione-S-transferase genes . We analysed the presence of DNA damage in leukocytes of 63 CF patients at an Italian CF centre and 63 controls , through the alkaline Comet assay to detect DNA strand breaks . Furthermore , controls and 93 CF subjects were genotyped for 5 genes by RFLP-PCR ( P18887 ,0GG1, P09211 ) and PCR assay ( P09488 , P30711 ) . No difference in Comet assay values was observed comparing controls to CF patients , although CF subjects showed slightly higher mean values . The crude Odds-Ratio ( OR ) was higher than one for P18887 and P09211 genotypes and liver status and for P18887 and O15527 genotypes and pancreatic insufficiency , but in all cases the p-values were not significant . In this case-control study , neither DNA damage ñor gene polymorphisms seem to influence CF manifestation . Methoxsalen stimulates electrogenic Cl- secretion in the mouse jejunum . We used the short-circuit current ( I(sc) ) and patch-clamp techniques to investigate the effects of methoxsalen ( MTX ) on the electrogenic Cl- secretion of the mouse jejunum . MTX stimulated a sustained increase in Isc that was dose dependent . DB00887 inhibited MTX-stimulated Isc in a dose-dependent manner consistent with activation of Cl- secretion . MTX failed to stimulate I(sc) following maximal activation of the DB02527 pathway by forskolin , but did increase Isc after a submaximal dose of forskolin . DB01016 , a blocker of the cystic fibrosis transmembrane conductance regulator ( P13569 ) , reduced the MTX-stimulated increase of Isc by 59 +/- 6 % . The DB02527 -dependent K+ channel blocker 293B did not alter the MTX-activated I(sc) ; however , clotrimazole , an intermediate Ca2(+)-activated K+ channel ( IK(Ca) ) blocker , reduced the MTX-stimulated I(sc) . MTX did not alter Na(+)-glucose cotransport across the mouse jejunum . In cell-attached membrane patches , MTX increased the open probability of the basolateral IK(Ca) channel of isolated crypts . These data suggest that the P13569 and IK(Ca) channels participate in the MTX-activated , sustained Cl- secretory response of the mouse jejunum . DB00887 blocks P13569 GCl in the native sweat duct . DB00887 is well known for its ability to inhibit the nonconductive Na+-K+-2Cl- cotransporter . We were surprised in preliminary studies to find that bumetanide in the contraluminal bath also inhibited NaCl absorption in the human sweat duct , which is apparently poor in cotransporter activity . Inhibition was accompanied by a marked decrease in the transepithelial electrical conductance . Because the cystic fibrosis transmembrane conductance regulator ( P13569 ) Cl- channel is richly expressed in the sweat duct , we asked whether bumetanide acts by blocking this anion channel . We found that bumetanide 1 ) significantly increased whole cell input impedance , 2 ) hyperpolarized transepithelial and basolateral membrane potentials , 3 ) depolarized apical membrane potential , 4 ) increased the ratio of apical-to-basolateral membrane resistance , and 5 ) decreased transepithelial Cl- conductance ( GCl ) . These results indicate that bumetanide inhibits P13569 GCl in both cell membranes of this epithelium . We excluded bumetanide interference with the protein kinase A phosphorylation activation process by " irreversibly " phosphorylating P13569 [ by using adenosine 5'-O-(3-thiotriphosphate) in the presence of a phosphatase inhibition cocktail ] before bumetanide application . We then activated P13569 GCl by adding 5 mM DB00171 . DB00887 in the cytoplasmic bath ( 10(-3) M ) inhibited approximately 71 % of this DB00171 -activated P13569 GCl , indicating possible direct inhibition of P13569 GCl . We conclude that bumetanide inhibits P13569 GCl in apical and basolateral membranes independent of phosphorylation . The results also suggest that > 10(-5) M bumetanide can not be used to specifically block the Na+-K+-2Cl- cotransporter . Characterization of plant P18887 and its interaction with proliferating cell nuclear antigen . In plants , there are no P06746 ( Pol beta ) and P49916 ( Lig3 ) genes . Thus , the plant short-patch base excision repair ( short-patch BER ) pathway must differ considerably from that in mammals . We characterized the rice ( Oryza Sativa L. cv. Nipponbare ) homologue of the mammalian X-ray repair cross complementing 1 ( P18887 ) , a well-known BER protein . The plant P18887 lacks the N-terminal domain ( NTD ) which is required for Pol beta binding and is essential for mammalian cell survival . The recombinant rice P18887 ( OsXRCC1 ) protein binds single-stranded DNA ( ssDNA ) as well as double-stranded DNA ( dsDNA ) and also interacts with rice proliferating cell nuclear antigen ( OsPCNA ) in a pull-down assay . Through immunoprecipitation , we demonstrated that OsXRCC1 forms a complex with P12004 in vivo . OsXRCC1 mRNA was expressed in all rice organs and was induced by application of bleomycin , but not of MMS , H(2)O(2) or UV-B . DB00290 also increased the fraction of OsXRCC1 associated with chromatin . These results suggest that OsXRCC1 contributes to DNA repair pathways that differ from the mammalian BER system . Genetic factors influencing outcome from neurotrauma . PURPOSE OF REVIEW : Clinical outcome after neurotrauma is considerably variable and can only partly be explained by known prognostic factors . There is converging evidence from genetic research that a number of genetic variants may contribute to this variability . This review provides recent data from human studies , published in the previous year , on genetic factors influencing outcome after neurotrauma . The bibliographic databases MEDLINE , EMBASE and PsycINFO were searched to identify relevant studies . RECENT FINDINGS : Genetic susceptibility to various aspects of clinical outcome after neurotrauma was reported in recent clinical studies . Genetic loci investigated include polymorphisms in P02649 , P21397 , P23560 , NOS3 , P05231 , P12036 , P31645 , P21964 , P48454 and Q8IX03 genes . The importance of these findings and future directions are discussed . SUMMARY : Recent genetic studies have revealed emerging aspects and extended the existing knowledge regarding the pathogenesis of neurotrauma and the genetic influence on phenotypic diversity . A better understanding of the underlying biological pathways and molecular mechanisms of an individual 's response to neurotrauma may hold the promise of novel treatment strategies and improved clinical outcome . Role of chloride transport proteins in the vasorelaxant action of nitroprusside in isolated rat aorta . Chloride ions play a key role in smooth muscle contraction , but little is known concerning their role in smooth muscle relaxation . Here we investigated the effect of chloride transport inhibitors on the vasorelaxant responses to nitroprusside in isolated and endothelium-denuded rat aorta , precontracted with phenylephrine 1 muM . Incubation of aortic rings in NO(3)(-) media strongly potentiated the vasorelaxant responses to nitroprusside . DB00887 , DIDS ( 4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid ) and acetazolamide strongly potentiated the vasorelaxant responses to nitroprusside ( by 70-100 % ) . EC(50) were 2.3+/-0.5 microM for bumetanide , 26+/-15 microM for DIDS and 510+/-118 microM for acetazolamide ( n=6 for condition ) . Niflumic acid , a selective inhibitor of ClCa ( calcium-activated chloride channels ) , potentiated nitroprusside relaxation to a similar extent as chloride transport inhibitors , in a non-additive manner . Zinc and nickel ions , both modestly potentiated nitroprusside vasorelaxation ( by 20-30 % ) . Cobaltum had negligible effect on nitroprusside vasorelaxation . P15085 ( p-chlorophenoxy-acetic acid ) , an inhibitor of volume-sensitive chloride channels ( ClC ) , slightly potentiated nitroprusside vasorelaxation ( by 15 % ) , and the cystic fibrosis transmembrane conductance regulator ( P13569 ) chloride channel inhibitors P13569 (inh)172 ( 5-[(4-Carboxyphenyl)methylene]-2-thioxo-3- [ (3-trifluoromethyl)phenyl-4-thiazolidinone ) , DPC ( diphenylamine-2,2'-dicarboxylic acid ) and glibenclamide were without significant effect . In conclusion , inhibition of chloride transport proteins strongly potentiates the vasorelaxant responses to nitroprusside in isolated rat aorta . This effect seems mediated by chloride depletion and inhibition of a chloride channel activated by both , calcium and cyclic GMP ( cGMP ) . DB01645 stimulates electrogenic Cl(-) secretion in mouse jejunum . We used the short-circuit current ( I(sc) ) technique to investigate the effects of the isoflavone genistein on the electrogenic Cl(-) secretion of the mouse jejunum . DB01645 stimulated a sustained increase in I(sc) that was dose dependent . DB00887 inhibited 76 +/- 5 % of the genistein-stimulated I(sc) consistent with activation of Cl(-) secretion . DB01645 failed to stimulate I(sc) following maximal activation of the DB02527 pathway by forskolin . In addition , forskolin had a reduced effect on I(sc) of the mouse jejunum in the presence of genistein . DB01016 , a blocker of P13569 , eliminated the genistein-stimulated increase of I(sc) and reduced the forskolin-activated I(sc) . Clotrimazole , a Ca(2+)-activated K(+) channel blocker , failed to reduce the genistein-stimulated I(sc) . Vanadate , a blocker of tyrosine-dependent phosphatases , reduced the genistein-activated I(sc) . Tyrphostin A23 , a tyrosine kinase inhibitor , reduced basal I(sc) , after which genistein failed to stimulate I(sc) . These data suggest that genistein activated a sustained Cl(-) secretory response of the mouse jejunum and that the effect of genistein was via a tyrosine-dependent phosphorylation pathway . Action of neltenexine on anion secretion in human airway epithelia . Neltenexine has been applied to human lung diseases such as chronic obstructive pulmonary disease ( P48444 ) as a mucolytic agent . However , we have no information on the neltenexine action in bronchial epithelial cells . We studied the neltenexine action on the ion transport in human submucosal serous Calu-3 cells . Under a hyper-secreting condition caused by terbutaline ( a beta2-adrenergic agonist ) , neltenexine diminished anion secretion by inhibiting the Cl- and HCO3- uptake via Na+/K+/2Cl- cotransporter and Na+/HCO3- cotransporter without blockade of the cystic fibrosis transmembrane conductance regulator ( P13569 ) channel , and also diminished anion secretion via stimulation of Cl-/HCO3- exchanger , which facilitates the extrusion of more P13569 -permeant anion , Cl- , with the uptake of less P13569 -permeant anion , HCO3- . Thus , neltenexine reduced the hyper-secretion to keep an appropriate fluid level in the airway , providing a possibility that neltenexine can be an effective drug in airway obstructive diseases by decreasing the airway resistance under a hyper-secreting condition . P05231 and P10145 increase the expression of glycosyltransferases and sulfotransferases involved in the biosynthesis of sialylated and/or sulfated Lewisx epitopes in the human bronchial mucosa . Bronchial mucins from patients suffering from CF ( cystic fibrosis ) exhibit glycosylation alterations , especially increased amounts of the sialyl-Lewis(x) ( NeuAcalpha2-3Galbeta1-4[Fucalpha1-3]GlcNAc-R ) and 6-sulfo-sialyl-Lewis(x) ( NeuAcalpha2-3Galbeta1-4[Fucalpha1-3][SO(3)H-6]GlcNAc-R ) terminal structures . These epitopes are preferential receptors for Pseudomonas aeruginosa , the bacteria responsible for the chronicity of airway infection and involved in the morbidity and early death of CF patients . However , these glycosylation changes can not be directly linked to defects in P13569 ( CF transmembrane conductance regulator ) gene expression since cells that secrete airway mucins express no or very low amounts of the protein . Several studies have shown that inflammation may affect glycosylation and sulfation of various glycoproteins , including mucins . In the present study , we show that incubation of macroscopically healthy fragments of human bronchial mucosa with P05231 ( interleukin-6 ) or P10145 results in a significant increase in the expression of alpha1,3/4-fucosyltransferases [ Q495W5 ( fucosyltransferase 11 gene ) and P21217 ] , alpha2-6- and alpha2,3-sialyltransferases [ Q9Y274 ( alpha2,3-sialyltransferase 6 gene ) and Q96JF0 ( alpha2,6-sialyltransferase 2 gene ) ] and GlcNAc-6-O-sulfotransferases [ Q8NCG5 ( carbohydrate sulfotransferase 4 gene ) and Q9GZX3 ] mRNA . In parallel , the amounts of sialyl-Lewis(x) and 6-sulfo-sialyl-Lewis(x) epitopes at the periphery of high-molecular-mass proteins , including Q99102 , were also increased . In conclusion , our results indicate that P05231 and -8 may contribute to the increased levels of sialyl-Lewis(x) and 6-sulfo-sialyl-Lewis(x) epitopes on human airway mucins from patients with CF . Structure of the human Q9H244 receptor in complex with an antithrombotic drug . P2Y receptors ( P2YRs ) , a family of purinergic G-protein-coupled receptors ( GPCRs ) , are activated by extracellular nucleotides . There are a total of eight distinct functional P2YRs expressed in human , which are subdivided into P47900 -like receptors and Q9H244 -like receptors . Their ligands are generally charged molecules with relatively low bioavailability and stability in vivo , which limits our understanding of this receptor family . P2Y12R regulates platelet activation and thrombus formation , and several antithrombotic drugs targeting P2Y12R -- including the prodrugs clopidogrel ( Plavix ) and prasugrel ( DB06209 ) that are metabolized and bind covalently , and the nucleoside analogue ticagrelor ( DB08816 ) that acts directly on the receptor -- have been approved for the prevention of stroke and myocardial infarction . However , limitations of these drugs ( for example , a very long half-life of clopidogrel action and a characteristic adverse effect profile of ticagrelor ) suggest that there is an unfulfilled medical need for developing a new generation of P2Y12R inhibitors . Here we report the 2.6 Å resolution crystal structure of human P2Y12R in complex with a non-nucleotide reversible antagonist , AZD1283 . The structure reveals a distinct straight conformation of helix V , which sets P2Y12R apart from all other known class A GPCR structures . With AZD1283 bound , the highly conserved disulphide bridge in GPCRs between helix III and extracellular loop 2 is not observed and appears to be dynamic . Along with the details of the AZD1283-binding site , analysis of the extracellular interface reveals an adjacent ligand-binding region and suggests that both pockets could be required for dinucleotide binding . The structure provides essential insights for the development of improved P2Y12R ligands and allosteric modulators as drug candidates .