interacts_with DB06271?
DB00092 ( anti- P06729 ) causes a selective reduction in circulating effector memory T cells ( Tem ) and relative preservation of central memory T cells ( Tcm ) in psoriasis . BACKGROUND : DB00092 ( anti- P06729 ) biological therapy selectively targets effector memory T cells ( Tem ) in psoriasis vulgaris , a model Type 1 autoimmune disease . METHODS : Circulating leukocytes were phenotyped in patients receiving alefacept for moderate to severe psoriasis . RESULTS : In all patients , this treatment caused a preferential decrease in effector memory T cells ( P32248 - CD45RA- ) ( mean 63 % reduction ) for both P01730 + and CD8+ Tem , while central memory T cells ( Tcm ) ( P32248 +CD45RA- ) were less affected , and naïve T cells ( P32248 +CD45RA+ ) were relatively spared . Circulating CD8+ effector T cells and Type 1 T cells ( P01579 -producing ) were also significantly reduced . CONCLUSION : DB00092 causes a selective reduction in circulating effector memory T cells ( Tem ) and relative preservation of central memory T cells ( Tcm ) in psoriasis . Matrix proteinase inhibition by DB05387 , a multifunctional antiangiogenic compound . BACKGROUND : Matrix metalloproteinases ( MMPs ) play an important role in tissue remodeling under normal physiological and pathological conditions and are thus attractive targets for both diagnostic and therapeutic purposes . Here , we examined the effect of DB05387 , an orally bioavailable standardized extract made of cartilage that shows significant antiangiogenic and antimetastatic properties in vivo , on the activity of various members of the MMP family . MATERIALS AND METHODS : The effect of DB05387 on the activity of MMPs was assessed by fluorimetric assays and by substrate gel zymography . RESULTS : DB05387 markedly inhibits the gelatinolytic activity of P08253 and to a lesser extent those of P03956 , P09237 , P14780 and P45452 . DB05387 also inhibited the elastinolytic activities of P08253 and P14780 as well as P39900 ( metalloelastase ) , porcine pancreatic elastase ( PPE ) , and human leukocyte elastase ( P08246 ) . Western blot analysis revealed the presence within DB05387 of immunoreactive P01033 -like proteins , suggesting that these proteins may be at least partly responsible for the observed MMP inhibition . CONCLUSIONS : Taken together , these results demonstrate that DB05387 contains P01033 -like proteins that could be responsible for the specific inhibition of MMPs . Given the recent studies suggesting the presence within this compound of specific inhibitor(s) of endothelial cell proliferation , DB05387 appears as a pleotropic agent able to interfere with several biochemical steps leading to angiogenesis and to other physiopathological conditions . Since DB05387 is currently under Phase III clinical investigations , these findings are also of considerable importance for our understanding of its anticancer properties . Oxidative stress-responsive transcription factor P18847 potentially mediates diabetic angiopathy . Previous results of our cDNA microarray analysis to look for genes whose expression level correlates well with in vitro tubulogenesis by NP31 endothelial cells revealed the transcription factor P18847 known to be responsive to stress such as reactive oxygen species ( ROS ) . Anti- P18847 small interfering RNA gave an inhibitory influence on tube formation by NP31 cells expressing an activated form of the vascular endothelial growth factor receptor 1 ( P17948 ) kinase . When expression of P18847 was regulated under the control of tetracycline system in NP31 cells , they acquired the tubulogenic ability upon P18847 induction . While P18847 failed to induce expressions of P15692 and VEGFR , it regulated those of P24941 , P11802 , p8 , plasminogen activator inhibitor 1 , integrin alpha1 , subunit and matrix metalloprotease P45452 . In H2O2-stimulated NP31 cells as well as endothelial cells of glomerulus and aorta of Otsuka-Long-Evans-Tokushima-Fatty diabetic model rats , concomitantly enhanced expressions of P18847 , P05121 , and p8 were observed . Given the proposed hypothesis of the close linkage between diabetic angiopathy and ROS , those data suggest that ROS-associated diabetic complication may involve P18847 -mediated pathological angiogenesis . P08069 targeted therapeutics : novel compounds and novel treatment strategies for cancer medicine . The insulin-like growth factor 1 receptor ( IGF-1R ) and its associated signalling system has provoked considerable interest over recent years as a novel therapeutic target in cancer . A brief outline of the IGF-1R signalling system and the rationale for its use in cancer medicine is given . This is followed by a discussion of the different possible targets within the IGF-1R system , and drugs developed to interact at each target . A systems-based approach is then used to review the in vitro and in vivo data in the published literature of the following compounds targeting IGF-1R components using specific examples : growth hormone releasing hormone antagonists ( e.g. JV-1-38 ) , growth hormone receptor antagonists ( e.g. pegvisomant ) , IGF-1R antibodies ( e.g. CP-751,871 , AVE1642/EM164 , DB05759 , P35240 -717454 , BIIB022 , Q99217 479 , MK-0646/h7C10 ) , and IGF-1R tyrosine kinase inhibitors ( e.g. BMS-536942 , BMS-554417 , DB00238 -AEW541 , DB00238 -ADW742 , AG1024 , potent quinolinyl-derived imidazo (1,5-a)pyrazine PQIP , picropodophyllin PPP , Nordihydroguaiaretic acid Insm-18/NDGA ) . The following tumour types are specifically discussed : lung , breast , colorectal , pancreatic , NETs , sarcoma , prostate , leukaemia , multiple myeloma . Other tumour types are mentioned briefly : squamous cell carcinoma of the head and neck , melanoma , glioblastoma , ovary , gastric and mesothelioma . Results of early stage clinical trials , involving recently patented drugs. are included where appropriate . We then outline the current understanding of toxicity related to IGF-1R targeted therapy , and finally outline areas for further research . Inhibition of cyclin-dependent kinases , GSK-3beta and CK1 by hymenialdisine , a marine sponge constituent . BACKGROUND : Over 2000 protein kinases regulate cellular functions . Screening for inhibitors of some of these kinases has already yielded some potent and selective compounds with promising potential for the treatment of human diseases . RESULTS : The marine sponge constituent hymenialdisine is a potent inhibitor of cyclin-dependent kinases , glycogen synthase kinase-3beta and casein kinase 1 . DB02950 competes with DB00171 for binding to these kinases . A P24941 -hymenialdisine complex crystal structure shows that three hydrogen bonds link hymenialdisine to the Glu81 and Leu83 residues of P24941 , as observed with other inhibitors . DB02950 inhibits Q00535 /p35 in vivo as demonstrated by the lack of phosphorylation/down-regulation of Pak1 kinase in E18 rat cortical neurons , and also inhibits GSK-3 in vivo as shown by the inhibition of P46821 phosphorylation . DB02950 also blocks the in vivo phosphorylation of the microtubule-binding protein tau at sites that are hyperphosphorylated by GSK-3 and Q00535 /p35 in Alzheimer 's disease ( cross-reacting with Alzheimer's-specific AT100 antibodies ) . CONCLUSIONS : The natural product hymenialdisine is a new kinase inhibitor with promising potential applications for treating neurodegenerative disorders . Expression of Th2-skewed pathology mediators in monocyte-derived type 2 of dendritic cells ( DC2 ) . The information conveyed from dendritic cells ( DCs ) to naïve P01730 (+) T cells has crucial influence on their differentiation toward effector T cells . In an effort to identify DC-derived molecules directly contributing to T cell differentiation , we searched for molecules distinctively expressed between two DC subtypes , which were differentiated from peripheral monocytes by cultivation with GM- P04141 ( for Q9NPG8 ) or P08700 ( for DC2 ) in the presence of P05112 and had the ability to induce naïve T cells to differentiate into Th1 or Th2 cells , respectively . As the first step to address this issue , we subtracted Q9NPG8 transcripts from those of DC2 and compiled the gene profile dominantly expressed in DC2 , whose products are known to reside in other than the nucleus . Intriguingly , many of them were molecules involved in Th2-skewed disease pathologies , such as P02751 , P38570 , Q14956 , Q03405 , P25089 , Q8NHJ6 , P05121 , P16050 , P24557 , P19878 , P10147 , P18510 , P09486 , and Q9NY15 , suggesting that DCs function not only as antigen presenting cells but also as producers of Th2 pathology specific milieus leading to disease deteriorations . We also found that expressions of Q02318 , O14495 , Q8WXG1 , and O15438 were up-regulated in DC2 , implying their significant function in Th2-deviated states . The identification of differentially expressed genes between DC subtypes provides new insights into their functions and our comparative gene expression profile will be highly useful for the identification of DC-derived key molecules for T cell differentiation . Two different P01579 nonresponsive variants derived from the B-cell lymphoma 70Z/3 . The kappa immunoglobulin ( Igk ) light chain locus is transcriptionally silent in the mouse B-cell lymphoma 70Z/3 . However , exposure to lipopolysaccharide ( LPS ) or interferon-gamma ( IFN ) causes a marked increase in Igk transcription . By immunoselection , we isolated two variants that are nonresponsive to IFN . One variant , AT7.2 , has retained its response to LPS ( IFN-LPS+ ) , whereas the other , P01008 .3 , is also nonresponsive to LPS ( IFN-LPS- ) . Stable transfection of an intact Igk gene does not rescue the phenotype of either variant . Both variants have intact Igk genes and neither is deficient in the binding or uptake of IFN . Nuclear extracts from LPS-treated wild-type 70Z/3 cells show strong increases in three transcription factors : P09086 , NF-kappa B , and kBF-A . Remarkably , when the IFN-LPS- variant is treated with LPS , all three transcription factors are still observed in the nuclear extracts . Treatment of wild-type cells with either LPS or IFN also causes a decrease in nuclear complexes that bind to two other regions of the Igk intron enhancer , the octenh and the E kappa MHCIC regions . Both of these changes are also observed after LPS or IFN treatment of the IFN-LPS- variant . Thus , this variant transduces the IFN and LPS signals at least into the nuclear compartment , but still fails to activate Igk transcription . In contrast , the IFN-LPS+ variant decreases neither the octenh nor the E kappa MHCIC binding complexes in response to IFN . This variant may be defective in transducing the IFN signal to the nucleus . These variants will be useful in studying the activation of Igk transcription and the IFN signaling pathway in B cells . Growth-inhibitory effects of vitamin D analogues and retinoids on human pancreatic cancer cells . Retinoids and vitamin D are important factors that regulate cellular growth and differentiation . An additive growth-inhibitory effect of retinoids and vitamin D analogues has been demonstrated for human myeloma , leukaemic and breast cancer cells . We set out to study the effects of the vitamin D analogue EB1089 and the retinoids all-trans- and 9-cis-retinoic acid on the human pancreatic adenocarcinoma cell lines Capan 1 and Capan 2 and the undifferentiated pancreatic carcinoma cell line Hs766T . The cell lines investigated expressed vitamin D receptor , retinoic acid receptor ( RAR ) -alpha and gamma as determined by polymerase chain reaction after reverse transcription . P10826 was expressed only in Hs766T cells . Addition of all-trans-retinoic acid increased the amount of P10276 mRNA in the three cell lines and induced P10826 mRNA in Capan 1 and Capan 2 cells . All-trans-retinoic acid at a concentration of 10 nM inhibited the growth of Capan 1 and Capan 2 cells by 40 % relative to controls . DB00523 was less effective . Neither all-trans-retinoic acid nor 9-cis-retinoic acid affected the growth of Hs766T cells . EB1089 , if added alone to the cells , did not significantly inhibit growth . However , the combination of 1 nM EB1089 with 10 nM all-trans-retinoic acid exerted a growth-inhibitory effect of 90 % in Capan 1 cells and of 70 % in Capan 2 cells . Our data suggest that vitamin D analogues together with retinoids inhibit the growth of human pancreatic cancer cells . However , in vivo studies are necessary to examine the potential use of retinoids and vitamin D analogues on pancreatic cancer . DB02342 in the human corpus luteum throughout the luteal phase and its influence on lutein cell steroidogenesis and angiogenic activity . OBJECTIVE : To quantitate 2-methoxyestradiol ( 2-ME ) in human corpus luteum ( CL ) of different ages and to determine the expression of cytochrome-P450-1A1 ( P04798 ) and catechol-O-methyl transferase ( P21964 ) in CL and the action of 2-ME on P , vascular endothelial growth factor ( P15692 ) secretion , and luteal angiogenesis . DESIGN : Experimental study . SETTING : University division of reproductive endocrinology . PATIENT(S) : Twenty-four women of reproductive age . INTERVENTION(S) : CL was collected from 15 women during the minilaparotomy for tubal sterilization . Granulosa lutein cells were harvested 36 hours after hCG administration in patients undergoing IVF . MAIN OUTCOMES MEASURE(S) : Levels of 2-ME were determined by high-performance liquid chromatography in CL . P04798 and P21964 were assessed by immunohistochemistry and Western blot . P and P15692 were measured by radioimmunoassay and ELISA . The angiogenic potential was analyzed using EA.hy926 cells . RESULT(S) : Plasma levels of E₂ decreased in the late luteal phase in association with an increase in luteal tissue of 2-ME concentrations . Concomitantly , there was a significant reduction of angiogenic activity in late CL . There was no significant variation in P04798 and P21964 expression in all CL . In physiological doses , 2-ME inhibited basal P15692 by granulosa lutein cells and diminished the angiogenic activity in conditioned media but did not prevent P and P15692 production stimulated by hCG . CONCLUSION(S) : These data suggest the participation of 2-ME in physiological luteolysis by reducing angiogenesis . However , 2-ME did not prevent in vitro hCG stimulation of P biosynthesis , providing a mechanism for CL rescue in the cycle of conception . Prevention of thrombus formation and growth by antithrombin III and heparin cofactor II-dependent thrombin inhibitors : importance of heparin cofactor II . DB01109 ( HEP ) prevents thrombus formation ( TF ) and thrombus growth ( TG ) , by accelerating thrombin ( THR ) inhibition by antithrombin III ( P01008 ) . Recent studies suggest that dermatan sulphate which catalyzes thrombin inhibition by heparin cofactor II ( HCII ) , can inhibit TF and TG as effectively as HEP . This study compared the antithrombotic effects of HEP and another agent , DB06271 ( SLX ) which catalyzes thrombin inhibition by P01008 and HCII simultaneously . TF was induced in rabbit jugular veins , using the stasis/hypercoagulation model . TG was measured as the accretion of 125I-fibrin onto existing thrombi in rabbit jugular veins . HEP and SLX inhibited TF when given in doses of 10 and 5 anti-thrombin U/kg , respectively . SLX ( 16 anti-thrombin U/kg or 260 micrograms/kg ) was more effective than HEP ( 120 anti-thrombin U/kg or 800 micrograms/kg ) in preventing TG when administered either as a bolus or by continuous infusion . These data suggest that agents which accelerate THR inhibition by both P01008 and HCII simultaneously , can inhibit TF and TG with less systemic anticoagulation than comparable antithrombotic doses of HEP . Identification of antithrombin-modulating genes . Role of O95461 , a gene encoding a bifunctional glycosyltransferase , in the secretion of proteins ? The haemostatic relevance of antithrombin together with the low genetic variability of P01008 , and the high heritability of plasma levels encourage the search for modulating genes . We used a hypothesis-free approach to identify these genes , evaluating associations between plasma antithrombin and 307,984 polymorphisms in the GAIT study ( 352 individuals from 21 Spanish families ) . Despite no SNP reaching the genome wide significance threshold , we verified milder positive associations in 307 blood donors from a different cohort . This validation study suggested O95461 , a gene encoding a protein with xylosyltransferase and glucuronyltransferase activities that forms heparin-like linear polysaccharides , as a potential modulator of antithrombin based on the significant association of one SNPs , rs762057 , with anti-FXa activity , particularly after adjustment for age , sex and P01008 rs2227589 genotype , all factors influencing antithrombin levels ( p = 0.02 ) . Additional results sustained this association . O95461 silencing inHepG2 and P29320 -EBNA cells did not affect P01008 mRNA levels but significantly reduced the secretion of antithrombin with moderate intracellular retention . Milder effects were observed on α1-antitrypsin , prothrombin and transferrin . Our study suggests O95461 as the first known modifier of plasma antithrombin , and proposes a new role for O95461 in modulating extracellular secretion of certain glycoproteins . Normal and perturbed endothelial cells from canine femoral arteries and femoral veins exhibit heterogeneity in hemostatic properties and growth characteristics . BACKGROUND : We sought to examine the heterogeneity of endothelial cells from the same anatomic site but different vascular systems and described P04275 ( P04275 ) release and morphological change in response to injury-associated factor in femoral vessels from canine in vitro . METHODS : Levels of hemostatic factors ( P04275 , plasminogen activator inhibitor type 1( P05121 ) , antithrombin III ( P01008 ) , in tissue sections and cultured endothelial cells of canine femoral arteries and canine femoral veins were compared by the immunohistochemistry technique . In addition to comparing cell growth density and cell protein contents , cultured femoral arterial endothelial cells ( FAECs ) and cultured femoral venous endothelial cells ( FVECs ) were incubated with a series concentration of basic fibroblast factor ( P09038 ) ( 1 , 10 , 100 ng/ml ) for up to 48 hours to test the amount of P04275 secretion and morphological change . RESULTS : Both in tissue sections and cultured cells , the levels of P04275 are higher in FVECs than in FAECs . We were unable to differentiate the level of P05121 and P01008 difference between FAECs and FVECs. P09038 ( 10 ng/ml ) significantly increased P04275 secretion from cultured FAECs but not from FVECs . The size of cultured FAECs is smaller than of FVECs ; however , FAECs have higher amounts of protein contents than FVECs . CONCLUSIONS : These comparative studies provide evidence indicating that the characteristics of FVECs differ from those of FAECs . These differences may be indicated heterogeneity with either inherited or acquired thrombotic disease . P54252 represses transcription via chromatin binding , interaction with histone deacetylase 3 , and histone deacetylation . P54252 ( P01008 ) , the disease protein in spinocerebellar ataxia type 3 ( P54252 ) , has been associated with the ubiquitin-proteasome system and transcriptional regulation . Here we report that normal P01008 binds to target DNA sequences in specific chromatin regions of the matrix metalloproteinase-2 ( P08253 ) gene promoter and represses transcription by recruitment of the histone deacetylase 3 ( O15379 ) , the nuclear receptor corepressor ( NCoR ) , and deacetylation of histones bound to the promoter . Both normal and expanded P01008 physiologically interacted with O15379 and NCoR in a P54252 cell model and human pons tissue ; however , normal P01008 -containing protein complexes showed increased histone deacetylase activity , whereas expanded P01008 -containing complexes had reduced deacetylase activity . Consistently , histone analyses revealed an increased acetylation of total histone H3 in expanded P01008 -expressing cells and human P54252 pons . Expanded P01008 lost the repressor function and displayed altered DNA/chromatin binding that was not associated with recruitment of O15379 , NCoR , and deacetylation of the promoter , allowing aberrant P08253 transcription via the transcription factor GATA-2 . For transcriptional repression normal P01008 cooperates with O15379 and requires its intact ubiquitin-interacting motifs ( UIMs ) , whereas aberrant transcriptional activation by expanded P01008 is independent of the UIMs but requires the catalytic cysteine of the ubiquitin protease domain . These findings demonstrate that normal P01008 binds target promoter regions and represses transcription of a GATA-2-dependent target gene via formation of histone-deacetylating repressor complexes requiring its UIM-associated function . Expanded P01008 aberrantly activates transcription via its catalytic site and loses the ability to form deacetylating repressor complexes on target chromatin regions . Effects of the enterokinetic prucalopride ( R093877 ) on colonic motility in fasted dogs . The novel enterokinetic drug prucalopride was tested at various intravenous and oral doses in fasted dogs to assess : ( i ) the effects on colonic contractile motility patterns ; and ( ii ) the mediation of these effects by 5-hydroxytryptamine ( Q13639 ) receptors . Colonic motility patterns were assessed in conscious dogs with four chronically implanted strain-gauge force transducers that were sutured on the serosal side of the colon . DB06480 altered colonic contractile motility patterns in a dose-dependent fashion by stimulating high-amplitude clustered contractions in the proximal colon and by inhibiting contractile activity in the distal colon . DB06480 was equipotent after oral and intravenous administration , as reflected by the values for the effective dose that induced 50 % of maximum effect ( 95 % confidence limits ) : 0.04 mg kg(-1) p.o . ( 0.01-0.1 mg kg(-1) ) and 0.01 mg kg(-1) i.v. ( 0.006-0.04 mg kg(-1) ) . DB06480 also caused a dose-dependent decrease in the time to the first giant migrating contraction ( GMC ) ; at higher doses of prucalopride , the first GMC generally occurred within the first half-hour after treatment . Subcutaneous pretreatment with the Q13639 receptor antagonist GR125487 ( 40 microg kg(-1) bodyweight ) completely prevented the effects of orally administered prucalopride ( 0.31 mg kg(-1) bodyweight ) . DB06480 , given orally or intravenously , alters colonic motility in the fasted conscious dog in a dose-dependent fashion . It induces GMCs and causes proximal colon stimulation and distal colon inhibition of contractile motility patterns by stimulating Q13639 receptors . P01308 -like growth factor-I:vitronectin complex-induced changes in gene expression effect breast cell survival and migration . Recent studies have demonstrated that P05019 associates with vitronectin ( VN ) through IGF-binding proteins ( IGFBP ) , which in turn modulate IGF-stimulated biological functions such as cell proliferation , attachment , and migration . Because IGFs play important roles in transformation and progression of breast tumors , we aimed to describe the effects of P05019 :IGFBP:VN complexes on breast cell function and to dissect mechanisms underlying these responses . In this study we demonstrate that substrate-bound P05019 :IGFBP:VN complexes are potent stimulators of MCF-7 breast cell survival , which is mediated by a transient activation of P29323 /MAPK and sustained activation of phosphoinositide 3-kinase/AKT pathways . Furthermore , use of pharmacological inhibitors of the MAPK and phosphoinositide 3-kinase pathways confirms that both pathways are involved in P05019 :IGFBP:VN complex-mediated increased cell survival . Microarray analysis of cells stimulated to migrate in response to P05019 :IGFBP:VN complexes identified differential expression of genes with previously reported roles in migration , invasion , and survival ( P52799 , Sharp-2 , Tissue-factor , P31947 , P05121 , P35568 ) . These changes were not detected when the P05019 analogue ( [L(24)][A(31)]- P05019 ) , which fails to bind to the P08069 , was substituted ; confirming the P05019 -dependent differential expression of genes associated with enhanced cell migration . Taken together , these studies have established that P05019 :IGFBP:VN complexes enhance breast cell migration and survival , processes central to facilitating metastasis . This study highlights the interdependence of extracellular matrix and growth factor interactions in biological functions critical for metastasis and identifies potential novel therapeutic targets directed at preventing breast cancer progression . All-trans retinoic acid inhibits the increases in fibronectin and P05121 induced by TGF-beta1 and Ang II in rat mesangial cells . AIM : To investigate the effect of all-trans RA ( atRA ) on the increases in plasminogen activator inhibitor-1 ( P05121 ) and fibronectin that are induced by transforming growth factor-beta1 ( TGF-beta1 ) and angiotensin II ( Ang II ) in cultured rat glomerular mesangial cells . METHODS : Subconfluent glomerular mesangial cells were serum-starved for 48 h and pretreated with atRA with subsequent stimulation of TGF-beta1 and Ang II . Protein expressions of cell-associated fibronectin and P05121 in glomerular mesangial cells were evaluated by Western blot analysis . mRNA expression of RA receptors in glomerular mesangial cells was examined by RT-PCR . RESULTS : Retinoic acid receptor-alpha , -gamma ( P10276 , -gamma ) and retinoid X receptor-alpha , -beta , -gamma ( RXR-alpha , -beta , -gamma ) mRNA were expressed in rat glomerular mesangial cells . atRA pretreatment effectively reduced fibronectin expression in glomerular mesangial cells stimulated with TGF-beta 1 or Ang II for 48 h . TGF-beta 1 stimulated P05121 expression reached a maximum at 5 h . atRA did n't affect the early ( 5 h ) P05121 induction by TGF-beta 1 , but markedly attenuated the sustained ( 48 h ) P05121 induction . atRA also decreased the prolonged effect of Ang II on P05121 expression . CONCLUSION : These results indicate that atRA inhibits the increases in fibronectin that are induced by TGF-beta1 and Ang II in cultured glomerular mesangial cells . The data also suggest that this effect of atRA is associated with a change in P05121 levels . DB01645 potentiates the P01160 effect on a K(+)-conductance in P29320 -293 cells . P29320 -293 cells are known to reflect many features of the late distal tubule . Furthermore , they have the ability to release urodilatin , the structural analog to P01160 . RT-PCR was performed to test for the expression of natriuretic peptide receptors . While the mRNA for the human P01160 receptor ( P16066 , P16066 ) could be amplified , the P09543 -specific receptor P20594 ( P20594 ) and the receptor specific for guanylins , P25092 , could not be detected . In patch clamp experiments the effects of P01160 ( 10 nM ) on membrane voltage ( V(m) ) were monitored and P29320 -293 cells depolarized by 2.3 +/- 0.5 mV ( n=14 ) . In the presence of the P01133 receptor blocker genistein ( 10 microM ) the effect of P01160 was increased by 65 % to 3.9 +/- 0.8 mV ( n=14 ) . After removal of genistein the P01160 -mediated depolarization further increased by 147 % to 5.7 +/- 1.0 mV ( n=14 ) . P01160 given repetitively without genistein had no increasing depolarizing effect in P29320 -293 cells with time . The P01160 effect could be fully blocked by 1 mM Ba(2+) and by 1 microM of the specific PKG inhibitor KT5823 indicating that P01160 inhibits a K(+)-conductance via a cGMP-dependent protein kinase . DB01645 itself hyperpolarized the membrane voltage of P29320 -293 cells by -3.9 +/- 0.6 mV ( n=11 ) and this effect could also be fully blocked by Ba(2+) ( -0.3 +/- 0.1 mV , n=5 ) , indicating that genistein activates a K(+)-conductance which contributes significantly to the membrane potential of P29320 -293 cells . Estrogenic chemicals at puberty change ERalpha in the hypothalamus of male and female rats . The effects of two environmental endocrine disruptors , the synthetic pharmaceutical estrogen DB00977 ( EE ) and bisphenol-A ( Q03001 ) , were analysed in male and female rats in a very sensitive developmental period , puberty . Immunohistochemistry was used to evaluate changes in the number of cells expressing estrogen receptors ( P03372 ) in the arcuate nucleus ( ARC ) , ventromedial nucleus ( VMH ) and medial preoptic area ( DB00603 ) of the hypothalamus . Animals were treated during early puberty , from P01160 23 to P01160 30 , with EE and Q03001 given orally every day . They were then sacrificed and perfused on P01160 37 or P01160 90 , and blood and brains were collected for hormonal determination ( testosterone and estradiol ) and immunohistochemistry ( estrogen receptors , ER ) . At P01160 37 , ER-labelled neurons were higher in males than in females in the ARC and DB00603 . EE and Q03001 increased ER-labelled neurons in the ARC and DB00603 . At P01160 90 , females showed higher ER-labelled neurons in the VMH . EE and Q03001 increased ER-labelled neurons in the DB00603 in females . EE increased testosterone in males at P01160 37 and estradiol in females at P01160 90 . These results indicate the ability of estrogenic chemicals to change the reproductive neural circuits during puberty in male and female rats . Purification and characterization of mouse O15528 overproduced by an Escherichia coli system coexpressing molecular chaperonins GroEL/ES . The expression of mouse O15528 in Escherichia coli has been dramatically enhanced by coexpression of GroEL/ES . To reveal the enzymatic properties of O15528 , we measured its hydroxylation activity toward vitamin D3 and DB01436 ( 1alpha(OH)D3 ) in addition to the physiological substrate DB00146 . Surprisingly , O15528 converted vitamin D3 to 1alpha, DB00146 . Both 1alpha-hydroxylation activity toward vitamin D3 , and 25-hydroxylation activity toward 1alpha(OH)D3 were observed . The Km and Vmax values for 25-hydroxylation activity toward 1alpha(OH)D3 were estimated to be 1.7 microM and 0.51 mol/min/mol P450 , respectively , while those for 1alpha-hydroxylation activity toward DB00146 were 0.050 microM and 2.73 mol/min/mol P450 , respectively . Note that the substrate must be fixed in the opposite direction in the substrate-binding pocket of O15528 between 1alpha-hydroxylation and 25-hydroxylation . Based on these results and the fact that human Q02318 and Streptomyces CYP105A1 also convert vitamin D3 to 1alpha, DB00146 , 1alpha-hydroxylation , and 25-hydroxylation of vitamin D3 appear to be closely linked together . A surface plasmon resonance-based solution affinity assay for heparan sulfate-binding proteins . A surface plasmon resonance-based solution affinity assay is described for measuring the K(d) of binding of heparin/heparan sulfate-binding proteins with a variety of ligands . The assay involves the passage of a pre-equilibrated solution of protein and ligand over a sensor chip onto which heparin has been immobilised . DB01109 sensor chips prepared by four different methods , including biotin-streptavidin affinity capture and direct covalent attachment to the chip surface , were successfully used in the assay and gave similar K(d) values . The assay is applicable to a wide variety of heparin/HS-binding proteins of diverse structure and function ( e.g. , P05230 , P09038 , P15692 , P10145 , P80075 , P01008 , P02776 ) and to ligands of varying molecular weight and degree of sulfation ( e.g. , heparin , DB05808 , sucrose octasulfate , naphthalene trisulfonate ) and is thus well suited for the rapid screening of ligands in drug discovery applications . Brominated cyclodipeptides from the marine sponge Geodia barretti as selective 5-HT ligands . The brominated cyclodipeptides barettin ( cyclo[(6-bromo-8-entryptophan)arginine] ) and 8,9-dihydrobarettin ( cyclo[(6-bromotryptophan)arginine] ) isolated from the marine sponge Geodia barretti have previously been shown to inhibit settlement of barnacle larvae in a dose-dependent manner in concentrations ranging from 0.5 to 25 microM . To further establish the molecular target and mode of action of these compounds , we investigated their affinity to human serotonin receptors . The tryptophan residue in the barettins resembles that of endogenous serotonin [ 5-hydroxytryptamine ] . A selection of human serotonin receptors , including representatives from all subfamilies ( 1-7 ) , were transfected into P29320 -293 cells . Barettin selectively interacted with the serotonin receptors 5- Q13049 , P28335 , and Q13639 at concentrations close to that of endogenous serotonin , with the corresponding Ki values being 1.93 , 0.34 , and 1.91 microM , respectively . 8,9-Dihydrobarettin interacted exclusively with the P28335 receptor with a Ki value of 4.63 microM ; it failed to show affinity to 5- Q13049 and Q13639 , indicating that the double bond between the tryptophan and arginine residue plays an important role in the interaction with the receptor proteins .