interacts_with DB01656?
Impact of smoking on dendritic cell phenotypes in the airway lumen of patients with P48444 . BACKGROUND : Myeloid dendritic cells ( DCs ) are increased in the airway wall of patients with chronic obstructive pulmonary disease ( P48444 ) , and postulated to play a crucial role in P48444 . However , DC phenotypes in P48444 are poorly understood . METHODS : Function-associated surface molecules on bronchoalveolar lavage fluid ( BALF ) DCs were analyzed using flow cytometry in current smokers with P48444 , in former smokers with P48444 and in never-smoking controls . RESULTS : Myeloid DCs of current smokers with P48444 displayed a significantly increased expression of receptors for antigen recognition such as BDCA-1 or Q9UJ71 , as compared with never-smoking controls . In contrast , former smokers with P48444 displayed a significantly decreased expression of these receptors , as compared with never-smoking controls . A significantly reduced expression of the maturation marker Q01151 on myeloid DCs was found in current smokers with P48444 , but not in former smokers with P48444 . The chemokine receptor P51681 on myeloid DCs , which is also important for the uptake and procession of microbial antigens , was strongly reduced in all patients with P48444 , independently of the smoking status . CONCLUSION : P48444 is characterized by a strongly reduced P51681 expression on myeloid DCs in the airway lumen , which might hamper DC interactions with microbial antigens . Further studies are needed to better understand the role of P51681 in the pathophysiology and microbiology of P48444 . Q07973 as a potential target for cancer therapy . Increasing evidence has accumulated to suggest that vitamin D may reduce the risk of cancer through its biologically active metabolite , DB00136 , which inhibits proliferation and angiogenesis , induces differentiation and apoptosis , and regulates many other cellular functions . Thus , it is plausible to assume that rapid clearance of DB00136 by highly expressed Q07973 could interrupt the normal physiology of cells and might be one cause of cancer initiation and progression . In fact , enhancement of Q07973 expression has been reported in literature for many cancers . Based on these findings , Q07973 -specific inhibitors and vitamin D analogs which are resistant to Q07973 -dependent catabolism might be useful for cancer treatment . Q07973 -specific inhibitor VID400 , which is an azole compound , markedly enhanced and prolonged the antiproliferative activity of DB00136 in the human keratinocytes . Likewise , Q07973 -resistant analogs such as 2α-(3-hydroxypropoxy)- DB00136 ( O2C3 ) and its P06681 -epimer ED-71 ( DB05295 ) , and 19nor- 2α-(3-hydroxypropyl)- DB00136 ( MART-10 ) showed potent biological effects . Our in vivo studies using rats revealed that MART-10 had a low calcemic effect , which is a suitable property as an anticancer drug . Much lower affinity of MART-10 for vitamin D binding protein ( DBP ) as compared with DB00136 may be related to its more potent cellular activities . Based on these results , we conclude that ( 1 ) high affinity for P11473 , ( 2 ) resistance to Q07973 -dependent catabolism , ( 3 ) low affinity for DBP , and ( 4 ) low calcemic effect may be required for designing potent vitamin D analogs for cancer treatment . Targeted therapies for adrenocortical carcinoma : IGF and beyond . Standard chemotherapy for adrenocortical cancer currently is under evaluation in the context of the recently completed FIRM-ACT evaluating the combination of mitotane with either streptozocin or etoposide , cisplatin , and doxorubicin . New agents are eagerly sought by the ACC community that hopes to make progress against this deadly disease . Investigators have begun to dissect the molecular and genomic context of ACC with a goal of identifying potential novel therapeutic agents . One gene consistently overexpressed in ACC is insulin growth factor type 2 . Targeting its receptor P08069 has shown encouraging results in ACC cell lines and against murine xenografts . As a result , clinical trials to evaluate agents targeting the P08069 have been done including mitotane and DB05759 ( a monoclonal antibody ) and the GALACCTIC trial that has just completed accrual to evaluate OSI-906 , a small molecule P08069 antagonist . On the horizon are other agents targeting other tyrosine kinases , including P01133 and FGF , and novel strategies such as individualized tumor analysis to select treatment . Genetic analysis of 56 polymorphisms in 17 genes involved in methionine metabolism in patients with abdominal aortic aneurysm . BACKGROUND : Previous studies suggested an association between abdominal aortic aneurysm ( AAA ) and hyperhomocysteinaemia , a complex trait determined by genetic and environmental factors . Our hypothesis was that polymorphisms in genes directly or indirectly involved in methionine metabolism may contribute to AAA susceptibility . METHOD : We studied 56 polymorphisms in P42898 , Q99707 , Q9UBK8 , P35520 , P11586 , P41440 , P40261 , P20062 , P23526 , Q93088 , Q9H2M3 , Q04609 , P04818 , Q7L5Y1 , P34896 , P27169 , Q15165 genes according to their demonstrated/putative function , localisation in promoter or regulatory and coding regions and/or heterozygosity values > 0.300 . Polymorphisms were evaluated by using a primer extension based microarray technology in 423 AAA patients and 423 matched controls . RESULTS : All polymorphisms resulted in Hardy-Weinberg equilibrium in patients and controls . At the multiple logistic regression analysis adjusted for traditional cardiovascular risk factors ( sex , age , hypertension , smoking habit , dyslipidaemia , diabetes ) and chronic obstructive pulmonary disease ( P48444 ) , rs8003379 P11586 ( odds ratio ( OR ) 0.41 , 95 % confidence interval ( CI ) 0.26 to 0.65 ) and rs326118 Q9UBK8 ( OR 0.47 , 95 % CI 0.29 to 0.77 ) polymorphisms resulted in independent susceptibility factor for AAA . CONCLUSIONS : After haplotype reconstruction , logistic regression analyses adjusted for traditional risk factors and P48444 showed a significant association among AAA and P23526 , Q04609 , P11586 , Q99707 , P40261 , P27169 and P04818 haplotypes . Our findings offer new insights into the pathogenesis of AAA . Differential recruitment of coregulator proteins steroid receptor coactivator-1 and silencing mediator for retinoid and thyroid receptors to the estrogen receptor-estrogen response element by beta-estradiol and 4-hydroxytamoxifen in human breast cancer . P03372 ( ER ) -alpha and Q92731 function as transcription factors , and both interact with nuclear regulatory proteins to enhance or inhibit transcription . We hypothesized that coregulators are expressed in breast cancer and may be differentially recruited by ERs in the presence of estrogen and tamoxifen . Q92731 was found to be expressed more frequently in node-negative patients ( P < 0.05 ) . Expression of steroid receptor coactivator-1 ( Q15788 ) was associated with nodal positivity ( P < 0.05 ) and resistance to endocrine treatment ( P < 0.001 ) . The spatial coexpression of P03372 , Q92731 , and the coregulatory proteins was established using immunofluorescence . In both cell lines ( MCF-7 and T47D ) and in primary breast cancer cell cultures , beta-estradiol up-regulated Q92731 and coregulator protein expression and increased P03372 / Q92731 interaction with the estrogen response element ( ERE ) . 4- Hydroxy-tamoxifen ( DB04468 ) increased P03372 and silencing mediator for retinoid and thyroid receptors ( Q9Y618 ) expression and increased ER-ERE binding . Q15788 and Q9Y618 were identified at the ER-ERE complex , and interactions between ER isoforms and coregulatory proteins were determined using immunoprecipitation . Both P03372 and Q92731 preferentially bound Q15788 in the presence of beta-estradiol . Conversely , in cells treated with DB04468 , P03372 and Q92731 bound Q9Y618 . Differential recruitment of Q15788 and Q9Y618 by P03372 and Q92731 in the presence of beta-estradiol and DB04468 may be central to the response of the tumor to endocrine treatment . 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 . Molecular cloning and radioligand binding characterization of the chemokine receptor P51681 from rhesus macaque and human . The aim of this study was to determine if macaque represents a suitable species for the pre-clinical evaluation of novel P51681 antagonists , such as maraviroc ( UK-427,857 ) . To do this we cloned and expressed P51681 from rhesus macaque and compared the binding properties of [ 125I ] -MIP-1beta and [ 3H ] -maraviroc with human recombinant P51681 . [ 125I ] -MIP-1beta bound with similar high affinity to P51681 from macaque ( K(d) = 0.24 +/- 0.05 nM ) and human ( K(d) = 0.23 +/- 0.05 nM ) and with similar kinetic properties . In competition binding studies the affinity of a range of human chemokines for macaque P51681 was also similar to human P51681 . DB04835 inhibited binding of [ 125I ] -MIP-1beta to P51681 from macaque and human with similar potency ( IC50 = 17.50 +/- 1.24 nM and 7.18 +/- 0.93 nM , respectively ) and antagonised MIP-1beta induced intracellular calcium release mediated through P51681 from macaque and human with similar potency ( IC50 = 17.50 +/- 3.30 nM and 12.07 +/- 1.89 , respectively ) . [ 3H ] -maraviroc bound with high affinity to P51681 from macaque ( K(d) = 1.36+/-0.07 nM ) and human ( K(d) = 0.86 +/- 0.08 nM ) , but was found to dissociate approximately 10-fold more quickly from macaque P51681 . However , as with the human receptor , maraviroc was shown to be a high affinity , potent functional antagonist of macaque P51681 thereby indicating that the macaque should be a suitable species in which to evaluate the pharmacology , safety and potential mechanism-related toxicology of novel P51681 antagonists . Anti-inflammatory properties of Septilin in lipopolysaccharide activated monocytes and macrophage . In vivo studies have suggested the immunomodulatory properties of Septilin , an herbal preparation . This drug is being used against various types of inflammatory disorders . However , the mechanism of action of Septilin in the modulation of inflammation is not explored using suitable in vitro models . Hence , we decided to study the modulatory role of Septilin in lipopolysaccharide ( LPS ) mediated signaling in macrophage and monocyte cells . It was observed from the present study that by employing tumor necrosis factor α ( P01375 -α ) bioassay and reverse transcription-polymerase chain reaction ( RT-PCR ) , Septilin inhibited P01375 -α production in LPS ( 1 μg/mL ) stimulated RAW 264.7 cells ( p < 0.05 ) . 80 % inhibition of P01375 -α was observed even at 2.5 % Septilin . Septilin at all the concentrations tested could also significantly block the LPS mediated nitric oxide ( NO ) production ( p < 0.01 ) and expression of inducible NO synthase ( P35228 ) gene . LPS mediated interleukin 6 ( P05231 ) and P10145 production was also blocked by Septilin at the concentrations tested . This herbal preparation could also inhibit cycloxygenase 2 ( P35354 ) activity and suppression of P35354 and phosphodiesterase 4 B ( Q07343 ) mRNA expression in a concentration dependent manner . Taken together , these findings from the present in vitro study suggest the anti-inflammatory and immunomodulatory properties of Septilin . Phosphodiesterase-4 influences the PKA phosphorylation status and membrane translocation of G-protein receptor kinase 2 ( P25098 ) in P29320 -293beta2 cells and cardiac myocytes . Membrane-recruitment of P25098 ( G-protein receptor kinase 2 ) provides a fundamental step in the desensitization process controlling GPCRs ( G-protein-coupled receptors ) , such as the beta2AR ( beta2-adrenergic receptor ) . In the present paper , we show that challenge of P29320 -293beta2 [ human embryonic kidney cells stably overexpressing the FLAG-tagged beta2AR-GFP ( green fluorescent protein ) ] cells with the beta-adrenoceptor agonist , isoprenaline , causes P25098 to become phosphorylated by PKA ( DB02527 -dependent protein kinase ) . This action is facilitated when DB02527 -specific DB05876 ( phosphodiesterase-4 ) activity is selectively inactivated , either chemically with rolipram or by siRNA ( small interfering RNA ) -mediated knockdown of Q07343 and Q08499 . DB05876 -selective inhibition by rolipram facilitates the isoprenaline-induced membrane translocation of P25098 , phosphorylation of the beta2AR by P25098 , membrane translocation of beta-arrestin and internalization of beta2ARs . DB05876 -selective inhibition also enhances the ability of isoprenaline to trigger the PKA phosphorylation of P25098 in cardiac myocytes . In the absence of isoprenaline , rolipram-induced inhibition of DB05876 activity in P29320 -293beta2 cells acts to stimulate PKA phosphorylation of P25098 , with consequential effects on P25098 membrane recruitment and P25098 -mediated phosphorylation of the beta2AR . We propose that a key role for DB05876 enzymes is : ( i ) to gate the action of PKA on P25098 , influencing the rate of P25098 phosphorylation of the beta2AR and consequential recruitment of beta-arrestin subsequent to beta-adrenoceptor agonist challenge , and ( ii ) to protect P25098 from inappropriate membrane recruitment in unstimulated cells through its phosphorylation by PKA in response to fluctuations in basal levels of DB02527 . Inspiratory fraction correlates with exercise capacity in patients with stable moderate to severe P48444 . BACKGROUND : Exercise intolerance is the hallmark of P48444 . Static lung hyperinflation and increased dynamic hyperinflation during exercise are associated with reduced functional capacity in P48444 patients . Inspiratory capacity correction for the total lung capacity , defined as inspiratory fraction ( IF ) , may be functionally more representative than other traditional indices in these patients . OBJECTIVE : To investigate the association between IF and exercise capacity in patients with stable , moderate to severe P48444 . METHODS : Fifty P48444 subjects and 34 healthy volunteers constituted the study cohort . Pulmonary function and cardiopulmonary exercise testing were performed , and ventilation and gas exchange parameters were measured . RESULTS : IF was significantly correlated with percent-of-predicted peak oxygen consumption ( VO2 ) in the subjects with P48444 ( r = 0.52 , P < .001 ) . IF was an independent predictor of reduced exercise capacity in the P48444 subjects , and was more sensitive and specific than percent-of-predicted FEV1 . Statistical analysis generated the equation : percent-of-predicted peak VO2 = 65.9 IF + 0.45 percent-of-predicted FEV1 + 35.8 ( R( P06681 ) = 0.39 , P < .001 ) . The subjects with IF < 0.23 had more severe lung hyperinflation and less exercise capacity than the subjects with IF > 0.23 . At peak exercise , the breathing frequencies of the 2 groups were similar , whereas the low-IF subjects had reduced peak minute ventilation and peak tidal volume , relative to the high-IF subjects . CONCLUSIONS : Compared to FEV1 , IF is a robust factor to reflect lung hyperinflation and to estimate the exercise capacity of subjects with stable moderate to severe P48444 . Identification of the fused bicyclic 4-amino-2-phenylpyrimidine derivatives as novel and potent DB05876 inhibitors . 2-Phenyl-4-piperidinyl-6,7-dihydrothieno[3,4-d]pyrimidine derivative ( 2 ) was found to be a new DB05876 inhibitor with moderate Q07343 activity ( IC50=150 nM ) . A number of derivatives with a variety of 4-amino substituents and fused bicyclic pyrimidines were synthesized . Among these , 5,5-dioxo-7,8-dihydro-6H-thiopyrano[3,2-d]pyrimidine derivative ( 18 ) showed potent Q07343 inhibitory activity ( IC50=25 nM ) . Finally , N-propylacetamide derivative ( 31b ) was determined as a potent inhibitor for both Q07343 ( IC50=7.5 nM ) and P01375 -α production in mouse splenocytes ( IC50=9.8 nM ) and showed good in vivo anti-inflammatory activity in the LPS-induced lung inflammation model in mice ( ID50=18 mg/kg ) . The binding mode of the new inhibitor ( 31e ) in the catalytic site of Q07343 is presented based on an X-ray crystal structure of the ligand-enzyme complex . Inhibition of aldehyde dehydrogenase 2 activity enhances antimycin-induced rat cardiomyocytes apoptosis through activation of MAPK signaling pathway . Aldehyde dehydrogenase 2 ( P05091 ) , a mitochondrial-specific enzyme , has been proved to be involved in oxidative stress-induced cell apoptosis , while little is known in cardiomyocytes . This study was aimed at investigating the role of P05091 in antimycin A-induced cardiomyocytes apoptosis by suppressing P05091 activity with a specific P05091 inhibitor DB02115 . Antimycin A ( 40μg/ml ) was used to induce neonatal cardiomyocytes apoptosis . DB02115 ( 60μM ) effectively inhibited P05091 activity by 50 % without own effect on cell apoptosis , and significantly enhanced antimycin A-induced cardiomyocytes apoptosis from 33.5±4.4 to 56.5±6.4 % ( Hochest method , p < 0.05 ) , and from 57.9±1.9 to 74.0±11.9 % ( FACS , p < 0.05 ) . Phosphorylation of activated MAPK signaling pathway , including extracellular signal-regulated kinase ( P27361 /2 ) , c-Jun NH2-terminal kinase ( JNK ) and p38 was also increased in antimycin A and daidzin treated cardiomyocytes compared to the cells treated with antimycin A alone . These findings indicated that modifying mitochondrial P05091 activity/expression might be a potential therapeutic option on reducing oxidative insults induced cardiomyocytes apoptosis . The role of MAPK and Nrf2 pathways in ketanserin-elicited attenuation of cigarette smoke-induced P10145 production in human bronchial epithelial cells . Cigarette smoking is a major risk factor in chronic obstructive pulmonary disease ( P48444 ) with chronic airway inflammation as a key feature . Blockade of serotonin receptor 2A ( 5-HTR(2A) ) with ketanserin has been found to improve lung function in P48444 patients . Furthermore , ketanserin has been shown to possess anti-inflammatory properties in vivo . In this study , we investigated the antioxidative and anti-inflammatory properties of ketanserin and its underlying mechanism of action on cigarette smoke-induced interleukin ( IL ) -8 release in vitro . Primary normal human bronchial epithelial cells and human bronchial epithelial cell line ( BEAS-2B ) were treated with or without ketanserin prior to exposure to cigarette smoke medium ( CSM ) . Exposure to CSM caused elevation of both mRNA and release of P10145 with increased phosphorylation of p38 and extracellular signal-regulated kinases 1 and 2 ( P27361 /2 ) . Consistently , CSM-induced P10145 release was blocked by SB203580 , U0126 , or Q02750 small interfering RNA ( siRNA ) but not SP600125 . On the other hand , CSM caused a dose-dependent decrease in the ratio of reduced glutathione to oxidized glutathione ( rGSH/GSSG ) together with an increased translocation of Nrf2 to the nucleus demonstrated by Western blot analysis . Knock down of Nrf2 by siRNA completely blocked CSM-induced P10145 release . Ketanserin suppressed CSM-induced P10145 release by inhibiting p38 , P27361 /2 MAPK , and Nrf2 signaling pathways and partially inhibited CSM-induced reduction of rGSH/GSSG ratio . Our data demonstrated the novel antioxidative and anti-inflammatory role of ketanserin via the Nrf2 signaling pathway in CSM-exposed human bronchial epithelial cells . This may open up new perspectives in the development of novel therapeutic targets in the treatment of cigarette smoke-related P48444 . DB00642 alters folate phenotype and inflammatory profile in EA.hy 926 cells grown under low-folate conditions . Elevated homocysteine is a risk marker for several major human pathologies . Emerging evidence suggests that perturbations of folate/homocysteine metabolism can directly modify production of inflammatory mediators . DB00642 acts by inhibiting thymidylate synthetase ( P04818 ) , dihydrofolate reductase ( P00374 ) , and glycinamide ribonucleotide formyltransferase ( GARFT ) . EA.hy 926 cells grown under low ( " Lo " ) and high ( " Hi " ) folate conditions were treated with pemetrexed . The concentrations of several intracellular folate derivatives were measured using LC-MRM/MS . Lo cells had lower total folate concentrations and a different distribution of the intracellular folate derivatives than Hi cells . Treatment with pemetrexed caused a decrease in individual folate analytes . Microarray analysis showed that several genes were significantly up or down-regulated in pemetrexed treated Lo cells . Several of the significantly up-regulated transcripts were inflammatory . Changes in transcript levels of selected targets , including P01024 , P10145 , and P00374 , were confirmed by quantitative RT-PCR . P01024 and P10145 transcript levels were increased in pemetrexed-treated Lo cells relative to Lo controls ; P00374 transcript levels were decreased . In Lo cells , P10145 and P01024 protein concentrations were increased following pemetrexed treatment . DB00642 drug treatment was shown in this study to have effects that lead to an increase in pro-inflammatory mediators in Lo cells . No such changes were observed in Hi cells , suggesting that pemetrexed could not modify the inflammatory profile in the context of cellular folate sufficiency . Sub-family selective actions in the ability of Erk2 Q96HU1 kinase to phosphorylate and regulate the activity of DB05876 cyclic AMP-specific phosphodiesterases . Expressed in intact cells and in vitro , Q07343 and Q08493 isoenzymes of cyclic nucleotide phosphodiesterase ( PDE ) , in common with Q08499 isoenzymes , are shown to provide substrates for C-terminal catalytic unit phosphorylation by the extracellular signal-regulated kinase Erk2 ( Q8NFH3 (MAPK) ) . In contrast , P27815 isoenzymes do not provide substrates for C-terminal catalytic unit phosphorylation by Erk2 . Mutant DB05876 enzymes were generated to show that Erk2 phosphorylation occurs at a single , cognate serine residue located within the C-terminal portion of the DB05876 catalytic unit . DB05876 long-form isoenzymes were markedly inhibited by Erk2 phosphorylation . The short-form PDE4B2 isoenzyme was activated by Erk2 phosphorylation . These functional changes in PDE activity were mimicked by mutation of the target serine for Erk2 phosphorylation to the negatively charged amino acid , aspartic acid . Epidermal growth factor ( P01133 ) challenge caused diametrically opposed changes in cyclic AMP levels in COS1 cells transfected to express the long PDE4B1 isoenzyme compared to cells expressing the short PDE4B2 isoenzyme . We suggest that DB05876 enzymes may provide a pivotal point for integrating cyclic AMP and Erk signal transduction in cells with 4 genes encoding enzymes that are either insensitive to Erk2 action or may either be activated or inhibited . This indicates that DB05876 isoenzymes have distinct functional roles , giving credence to the notion that distinct therapeutic benefits may accrue using either DB05876 subfamily or isoenzyme-selective inhibitors . Mechanism of inhibition of the P42262 AMPA receptor channel opening by talampanel and its enantiomer : the stereochemistry of the 4-methyl group on the diazepine ring of 2,3-benzodiazepine derivatives . Stereoselectivity of 2,3-benzodiazepine compounds provides a unique way for the design of stereoisomers as more selective and more potent inhibitors as drug candidates for treatment of the neurological diseases involving excessive activity of AMPA receptors . Here we investigate a pair of enantiomers known as DB04982 and its ( + ) counterpart about their mechanism of inhibition and selectivity toward four AMPA receptor subunits or P42261 -4 . We show that DB04982 is the eutomer with the endismic ratio being 14 for the closed-channel and 10 for the open-channel state of P42262 . Kinetic evidence supports that DB04982 is a noncompetitive inhibitor and it binds to the same site for those 2,3-benzodiazepine compounds with the C-4 methyl group on the diazepine ring . This site , which we term as the " M " site , recognizes preferentially those 2,3-benzodiazepine compounds with the C-4 methyl group being in the R configuration , as in the chemical structure of DB04982 . Given that DB04982 inhibits P42261 and P42262 , but is virtually ineffective on the P42263 and P48058 AMPA receptor subunits , we hypothesize that the " M " site(s) on P42261 and P42262 to which DB04982 binds is different from that on P42263 and P48058 . If the molecular properties of the AMPA receptors and DB04982 are used for selecting an inhibitor as a single drug candidate for controlling the activity of all AMPA receptors in vivo , DB04982 is not ideal . Our results further suggest that addition of longer acyl groups to the N-3 position should produce more potent 2,3-benzodiazepine inhibitors for the " M " site . Measuring ligand-dependent and ligand-independent interactions between nuclear receptors and associated proteins using Bioluminescence Resonance Energy Transfer ( BRET ) . Bioluminescent resonance energy transfer ( BRET2 ) is a recently developed technology for the measurement of protein-protein interactions in a live , cell-based system . BRET2 is characterized by the efficient transfer of excited energy between a bioluminescent donor molecule ( Renilla luciferase ) and a fluorescent acceptor molecule ( a mutant of Green Fluorescent Protein ( GFP2 ) . The BRET2 assay offers advantages over fluorescence resonance energy transfer ( FRET ) because it does not require an external light source thereby eliminating problems of photobleaching and autoflourescence . The absence of contamination by light results in low background that permits detection of very small changes in the BRET2 signal . BRET2 is dependent on the orientation and distance between two fusion proteins and therefore requires extensive preliminary standardization experiments to conclude a positive BRET2 signal independent of variations in protein titrations and arrangement in tertiary structures . P03372 ( ER ) signaling is modulated by steroid receptor coactivator 1 ( Q15788 ) . To establish BRET2 in a ligand inducible system we used Q15788 as the donor moiety and ER as the acceptor moiety . Expression and functionality of the fusion proteins were assessed by transient transfection in P29320 -293 cells followed by Western blot analysis and measurement of ER-dependent reporter gene activity . These preliminary determinations are required prior to measuring nuclear receptor protein-protein interactions by BRET2 . This article describes in detail the BRET2 methodology for measuring interaction between full-length ER and coregulator proteins in real-time , in an in vivo environment . Inhibitory interactions between phosphorylation sites in the C terminus of α-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid-type glutamate receptor P42261 subunits . The C terminus of AMPA-type glutamate receptor ( AMPAR ) P42261 subunits contains several phosphorylation sites that regulate AMPAR activity and trafficking at excitatory synapses . Although many of these sites have been extensively studied , little is known about the signaling mechanisms regulating P42261 phosphorylation at DB00156 -840 . Here , we report that neuronal depolarization in hippocampal slices induces a calcium and protein phosphatase 1/2A-dependent dephosphorylation of P42261 at DB00156 -840 and a nearby site at DB00133 -845 . Despite these similarities , inhibitors of DB01221 -type glutamate receptors and protein phosphatase 2B prevented depolarization-induced DB00133 -845 dephosphorylation but had no effect on DB00156 -840 dephosphorylation . Instead , depolarization-induced DB00156 -840 dephosphorylation was prevented by blocking voltage-gated calcium channels , indicating that distinct Ca(2+) sources converge to regulate P42261 dephosphorylation at DB00156 -840 and DB00133 -845 in separable ways . Results from immunoprecipitation/depletion assays indicate that DB00156 -840 phosphorylation inhibits protein kinase A ( PKA ) -mediated increases in DB00133 -845 phosphorylation . Consistent with this , PKA-mediated increases in AMPAR currents , which are dependent on DB00133 -845 phosphorylation , were inhibited in P29320 -293 cells expressing a DB00156 -840 phosphomimetic version of P42261 . Conversely , mimicking DB00133 -845 phosphorylation inhibited protein kinase C phosphorylation of DB00156 -840 in vitro , and PKA activation inhibited DB00156 -840 phosphorylation in hippocampal slices . Together , the regulation of DB00156 -840 and DB00133 -845 phosphorylation by distinct sources of Ca(2+) influx and the presence of inhibitory interactions between these sites highlight a novel mechanism for conditional regulation of AMPAR phosphorylation and function . Q9UBK8 Q9UBK8 66A > G has no effect on total homocysteine , folate , and DB00115 concentrations in renal transplant patients . The association of variants of the gene encoding methionine synthase reductase ( Q9UBK8 ) with hyperhomocysteinemia , folate and Vitamin B(12) status in kidney graft recipients is unknown . We examined two mutations in Q9UBK8 in a cross-sectional study of 733 kidney graft recipients . The allele frequency of Q9UBK8 66G was 0.55 . 369 patients ( 50.3 % ) were heterozygous and 219 patients ( 29.9 % ) were homozygous for the mutation . None of the patients showed the 997C > G mutation . The allelic variants of Q9UBK8 66A > G showed no significant association with total homocysteine ( tHcy ) levels , both in univariate analyses , and in a multivariate model controlling for age , gender , body mass index , renal function , time since transplantation , underlying kidney disease , as well as the P42898 677C > T/1298A > C genotypes . Similarly , no significant associations between the Q9UBK8 66A > Ggenotypes and plasma folate or Vitamin B(12) levels were found . In conclusion , Q9UBK8 66A > G has no major effect on tHcy , folate , or Vitamin B(12) plasma concentrations in kidney graft recipients . Open study of pranlukast add-on therapy in intractable partial epilepsy . Innovative treatments of epileptic seizures are needed to improve the outcome of epilepsy . We studied the effect of pranlukast on seizure outcome in patients with intractable partial epilepsy . An open study was conducted to evaluate the clinical efficacy of 24-week pranlukast add-on therapy in 50 patients with intractable partial seizures . Serum concentrations of matrix metalloproteinase ( MMP ) -9 were determined using Biotrak Activity Assay System . Cytokines in cerebrospinal fluid ( P04141 ) were measured by the BioPlex ( BioRad ) system and soluble P01375 receptor1 ( sTNFR1 ) in P04141 was measured by the ELISA . Surface markers of lymphocytes in P04141 were examined by cell-sorter . Seizure-free rate ( SFR ) was 13.6 % , responder rate ( RR ) was 47.7 % , and aggravation rate ( AR ) was 18.2 % at the 13-24 week period after starting pranlukast . In patients with increased serum P14780 before pranlukast therapy ( baseline ) , comparison of paired serum levels showed a significant decrease after pranlukast therapy . Baseline P04141 levels of IL-1β and P05231 were elevated in patients compared with disease controls . Of four patients with paired data , three ( including a responder to pranlukast ) showed decreased pro-inflammatory cytokines ( IL-1β , P05231 , and TNFα ) , and four showed decreased sTNFR1 , after pranlukast treatment , and only a responder had markedly decreased frequency of CD8+ T cells in P04141 . DB01411 reduces seizure frequencies probably by pleiotropic effects including normalization of P14780 in sera , reduced leakage of pro-inflammatory cytokines into CNS , and inhibition of extravasation of leucocytes from brain capillaries . Further investigations by double-blind control study and animal models are warranted .