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M2
Terms modified by M2 Selected AbstractsMolecular characterization of regenerated cardiomyocytes derived from adult mesenchymal stem cellsCONGENITAL ANOMALIES, Issue 1 2002Keiichi Fukuda ABSTRACT, We recently isolated a cardiomyogenic (CMG) cell line from murine bone marrow stroma, and in this paper characterize regenerated cardiomyocytes derived from adult mesenchymal stem cells at the molecular level. Stromal cells were immortalized, exposed to 5-azacytidine, and repeatedly screened for spontaneously beating cells. CMG cells began to beat spontaneously after 2 weeks, and beat synchronously after 3 weeks. They exhibited sinus-node-like or ventricular-cell-like action potentials. Analysis of the isoforms of contractile protein genes, such as of myosin and ,-actin, indicated that their phenotype was similar to that of fetal ventricular cardiomyocytes. The cells expressed Nkx2.5, GATA4, TEF-1, and MEF2-C mRNA before 5-azacytidine exposure, and MEF2-A and MEF2-D after exposure. CMG cells expressed ,1A, ,1B, and ,1D -adrenergic receptor mRNA prior to differentiation, and ,1, ,2 -adrenergic and M1, M2 -muscarinic receptors after acquiring the cardiomyocyte phenotype. Phenylephrine induced phosphorylation of ERK1/ 2, and the phosphorylation was inhibited by prazosin. Isoproterenol increased the cAMP level 38-fold and beating rate, cell motion, % shortening, and contractile velocity by 48%, 38%, 27%, and 51%, respectively, and the increases were blocked by CGP20712A (,1 -selective blocker). Car-bachol increased IP3 32-fold, and the increase was inhibited by AFDX116 (M2 -selective blocker). These findings demonstrated that the regenerated cardiomyocytes were capable of responding to adrenergic and muscarinic stimulation. This new cell line provides a model for the study of cardiomyocyte transplantation. [source] Targeted cell-ablation in Xenopus embryos using the conditional, toxic viral protein M2(H37A)DEVELOPMENTAL DYNAMICS, Issue 8 2007Stuart J. Smith Abstract Harnessing toxic proteins to destroy selective cells in an embryo is an attractive method for exploring details of cell fate and cell,cell interdependency. However, no existing "suicide gene" system has proved suitable for aquatic vertebrates. We use the M2(H37A) toxic ion channel of the influenza-A virus to induce cell-ablations in Xenopus laevis. M2(H37A) RNA injected into blastomeres of early stage embryos causes death of their progeny by late-blastula stages. Moreover, M2(H37A) toxicity can be controlled using the M2 inhibitor rimantadine. We have tested the ablation system using transgenesis to target M2(H37A) expression to selected cells in the embryo. Using the myocardial MLC2 promoter, M2(H37A)-mediated cell death causes dramatic loss of cardiac structure and function by stage 39. With the LURP1 promoter, we induce cell-ablations of macrophages. These experiments demonstrate the effectiveness of M2(H37A)-ablation in Xenopus and its utility in monitoring the progression of developmental abnormalities during targeted cell death experiments. Developmental Dynamics 236:2159,2171, 2007. © 2007 Wiley-Liss, Inc. [source] Effects and serum levels of glibenclamide and its active metabolites in patients with type 2 diabetesDIABETES OBESITY & METABOLISM, Issue 6 2001A. Jönsson SUMMARY Objective To study the effects and serum levels of glibenclamide (Gb) and its active metabolites in patients on chronic Gb medication on different daily doses. Material and methods Fifty patients with type 2 diabetes on regular Gb therapy (1.75,14.0 mg daily). Blood samples were taken immediately before and 90 min after regular Gb intake. A standardized breakfast was served 30 min after drug intake. Serum insulin and proinsulin levels were determined by ELISA methods without cross-reactivities. Serum drug levels were determined by HPLC. Fischer's R to Z -test (correlation coefficients) and paired Student t -tests were used when comparing values within the entire group and unpaired non-parametric Mann,Whitney tests were used when comparing high and low dose levels. A p-value <,0.05 was considered significant. Results There were significant correlations between daily Gb dose, on the one hand, and, on the other, HbAlc (r = 0.55), ,-insulin (r = , 0.59) and ,-proinsulin (r = , 0.52) levels. Significant correlations between Gb therapy duration and insulin (r = , 0.40) and proinsulin (r = , 0.34) secretion and between Gb dose and ratio proinsulin/insulin (RPI) at both time points (r = 0.32 and 0.30) were also found. The RPI was lower after Gb intake. In patients on , 10.5 mg steady state serum metabolite levels (Ml and Ml + M2) were higher (29(0,120) and 33 (0,120) ng/ml) than those of Gb itself (18(0,64) ng/ml). A great inter-subject variability in Gb levels at both time points was seen. Conclusions Our results indicate that, in patients on chronic medication, Gb is capable of stimulating both insulin and proinsulin secretion; the effect on insulin release is relatively greater. The effect was more pronounced in patients on a low Gb dose, either because of less impaired ,-cells in those receiving low doses, or due to reduced sulphonylurea sensitivity in those on high dosage (down-regulation). In patients on a daily dose of 10.5 mg or more, serum metabolite levels of clinical relevance were demonstrated; the metabolites may contribute to hypoglycaemic events. [source] Upregulation of glycolytic enzymes in proteins secreted from human colon cancer cells with 5-fluorouracil resistanceELECTROPHORESIS, Issue 12 2009Young-Kyoung Shin Abstract 5-Fluorouracil (5-FU) is the most commonly used chemotherapeutic agent for colorectal cancer (CRC). However, resistance to this drug is a major obstacle in CRC chemotherapy. Accurate prediction of response to 5-FU would avoid unnecessary chemotherapy and allow the selection of other effective drugs. To identify a candidate predictor of 5-FU resistance, we isolated secreted proteins that were up- or downregulated in a 5-FU-resistant cancer cell line, compared with the parent cell line (SNU-C4), using a stable isotope-coded labeling protocol. For validating the clinical applicability of this method, levels of the identified proteins were determined in the sera of 46 patients treated with 5-FU. In total, 238 proteins with molecular weights ranging from 50 to 75,kDa were identified. Among these, 45 and 35 secreted proteins were up- and downregulated in the 5-FU-resistant cell line, respectively. We observed significant upregulation of glycolytic enzymes, including glyceraldehyde-3-phosphate dehydrogenase, pyruvate kinase M2 (PK-M2), transketolase, and NADP(+)-dependent malic enzyme 1. In particular, the level of PK-M2, a key enzyme in the glycolytic pathway, showed an increasing tendency in both sera and tissues from CRC patients displaying no response to 5-FU-based chemotherapy (progressive and stable disease cases), compared with that in complete or partial responders to 5-FU-based chemotherapy; however, it did not reach the statistical significance. In conclusion, increasing pattern of PK-M2 observed with 5-FU resistance induced in vitro and in sera and tissues from CRC patients displaying poor response to 5-FU-based chemotherapy suggest the relevance of dysregulated glycolysis and 5-FU-resistant CRC. [source] The importance of independent risk-factors for long-term mortality prediction after cardiac surgeryEUROPEAN JOURNAL OF CLINICAL INVESTIGATION, Issue 9 2006I. K. Toumpoulis Abstract Background, The purpose of the present study was to determine independent predictors for long-term mortality after cardiac surgery. The European System for Cardiac Operative Risk Evaluation (EuroSCORE) was developed to score in-hospital mortality and recent studies have shown its ability to predict long-term mortality as well. We compared forecasts based on EuroSCORE with other models based on independent predictors. Methods, Medical records of patients with cardiac surgery who were discharged alive (n = 4852) were retrospectively reviewed. Their operative surgical risks were calculated according to EuroSCORE. Patients were randomly divided into two groups: training dataset (n = 3233) and validation dataset (n = 1619). Long-term survival data (mean follow-up 5·1 years) were obtained from the National Death Index. We compared four models: standard EuroSCORE (M1); logistic EuroSCORE (M2); M2 and other preoperative, intra-operative and post-operative selected variables (M3); and selected variables only (M4). M3 and M4 were determined with multivariable Cox regression analysis using the training dataset. Results, The estimated five-year survival rates of the quartiles in compared models in the validation dataset were: 94·5%, 87·8%, 77·1%, 64·9% for M1; 95·1%, 88·0%, 80·5%, 64·4% for M2; 93·4%, 89·4%, 80·8%, 64·1% for M3; and 95·8%, 90·9%, 81·0%, 59·9% for M4. In the four models, the odds of death in the highest-risk quartile was 8·4-, 8·5-, 9·4- and 15·6-fold higher, respectively, than the odds of death in the lowest-risk quartile (P < 0·0001 for all). Conclusions, EuroSCORE is a good predictor of long-term mortality after cardiac surgery. We developed and validated a model using selected preoperative, intra-operative and post-operative variables that has better discriminatory ability. [source] Pregnancy-induced sympathetic overactivity: a precursor of preeclampsia,EUROPEAN JOURNAL OF CLINICAL INVESTIGATION, Issue 6 2004T. Fischer Abstract Background, Preeclampsia has been shown to constitute a state of sympathetic overactivity. However, it remains unclear if the sympathetic activity precedes preeclampsia or represents only a secondary phenomenon. To further investigate this issue, we performed a prospective study in pregnant women considered to be at increased risk for preeclampsia owing to preeclampsia during a preceding pregnancy. Materials and methods, Twenty-two women with a history of preeclampsia were longitudinally studied on three occasions: twice during pregnancy (M1: 22 ± 4, M2: 33 ± 5 weeks) and once postpartum (M3: 26 ± 6 weeks postpartum). We measured muscle sympathetic nerve activity (MSNA), forearm blood flow, and blood pressure at rest and during reactive hyperaemia after forearm occlusion. Results, At M1 and M2, none of the subjects was hypertensive, however, muscle sympathetic nerve activity levels were significantly augmented, compared with their postpartum values (M1: 21 ± 9, M2: 29 ± 14, M3: 9 ± 5 bursts min,1; P < 0·05). Forearm vascular resistance did not significantly change from M1 through M3 (M1: 16 ± 9, M2: 15 ± 7, M3: 16 ± 7 U; P = NS). Gestational muscle sympathetic nerve activity values did not differ significantly among the subjects with subsequent preeclampsia compared with those who remained normotensive [with preeclampsia (n = 6): M1: 21 ± 5, M2: 27 ± 6, M3: 7 ± 4 bursts min,1; without preeclampsia (n = 16): M1: 21 ± 11, M2: 30 ± 16, M3: 9 ± 6 bursts min,1; P = NS]. Conclusion, Invariably, all women at risk for preeclampisa showed a pregnancy-induced increase in MSNA (pregnancy-induced sympathetic overactivity, PISO), which normalized after delivery. Most importantly, PISO is not necessarily associated with peripheral vasoconstriction and hypertension. Furthermore, only a subset of patients developed preeclampsia later on. Therefore, we hypothesize that PISO constitutes a precursor of preeclampsia which is physiologically compensated for by vasodilating mechanisms, leading to preeclampsia only when they fail. [source] High epitope density in a single protein molecule significantly enhances antigenicity as well as immunogenicity: a novel strategy for modern vaccine development and a preliminary investigation about B,cell discrimination of monomeric proteinsEUROPEAN JOURNAL OF IMMUNOLOGY, Issue 2 2005Wanli Liu Abstract Although early studies have shown a close correlation between epitope density and epitope-specific humoral immune responses, few attempts have been made to quantitatively compare the antigenic and immunogenic differences between protein molecules bearing low or high degrees of epitope density, nor have studies quantitatively investigated the mechanism of B,cell discrimination of monomeric antigens. In this study, we prepared glutathione S-transferase (GST) fusion proteins bearing various copies of the M2e epitope from the influenza virus M2,protein [GST-(M2e)8, GST-(M2e)4 and GST-(M2e)1], which were used to detect and compare the real-time kinetic binding with M2e-specific mAb by surface plasma resonance. Our data show clearly that fusion proteins bearing higher M2e epitope density resulted in higher average avidity for M2e-specific mAb. Furthermore, it was observed that fusion proteins bearing high M2e epitope density could induce polyclonal antibodies (pAb) with enhanced an average affinity constant (KA) for M2e epitope peptide compared to fusion proteins bearing low epitope density. The average KA of pAb induced by GST-(M2e)8 (3.08 × 108,M,1 or 9.96 × 108,M,1) was up to two orders of magnitude greater than the average KA of pAb induced by GST-(M2e)1 (2.00 × 106,M,1 or 3.43 × 106,M,1). Thus, the data presented here demonstrate that high epitope density in a single protein molecule significantly enhances antigenicity and immunogenicity. These findings enrich our knowledge of how epitope density might relate to the recognition, activation and antibody production processes of epitope-specific immature B,cells. [source] Comparison of Isoelectronic Heterometallic and Homometallic Binuclear Cyclopentadienylmetal Carbonyls: The Iron,Nickel vs. the Dicobalt SystemsEUROPEAN JOURNAL OF INORGANIC CHEMISTRY, Issue 8 2008Jun D. Zhang Abstract The heterometallic binuclear cyclopentadienylironnickel carbonyl compounds Cp2FeNi(CO)n (n = 3, 2, 1; Cp = ,5 -C5H5) have been studied by density functional theory (BP86) for comparison with the isoelectronic homometallic dicobalt derivatives Cp2Co2(CO)n. The FeNi tricarbonyl is shown to be the doubly bridged isomer Cp2Fe(CO)Ni(,-CO)2 with an Fe,Ni distance of 2.455 Å (BP86), in accord with experiment and in contrast to Cp2Co2(CO)3 where singly and triply bridged but not doubly bridged isomers are found. The dicarbonyl compounds Cp2FeNi(,-CO)2 and Cp2Co2(,-CO)2 both have analogous doubly bridged structures with M=M distances around 2.35 Å, suggesting formal M=M double bonds. The monocarbonyl compounds have analogous singly bridged axial structures Cp2FeNi(,-CO) and Cp2Co2(,-CO) with metal,metal distances in the range 2.05 Å (M2 = Co2) to 2.12 Å (M2 = FeNi) consistent with the formal M,M triple bonds required for the favored 18-electron configuration. Open-shell states of Cp2FeNi(,-CO) are found to have even lower energies than the closed-shell structure, which indicates that the ground state of Cp2FeNi(,-CO) might be a high spin structure. However, the global minimum for the monocarbonyl is found to be a singlet "hot dog" perpendicular Cp2NiFe(CO) structure with a terminal CO group bonded to the iron atom. Other higher energy perpendicular structures are also found for Cp2FeNi(CO)n (n = 3, 2, 1) with terminal CO groups and bridging Cp rings. (© Wiley-VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2008) [source] Using Chiral Ligand Substituents To Promote the Formation of Dinuclear, Double-Stranded Iron, Manganese, and Zinc MesocatesEUROPEAN JOURNAL OF INORGANIC CHEMISTRY, Issue 33 2007Stuart D. Reid Abstract The synthesis and structures of dinuclear manganese, iron, and zinc complexes of chiral di-iminodipyrromethane ligands (L) are reported. Schiff base condensation reactions between 5,5,-diformyl-2,2,-dipyrromethane and the chiral amines (,)-(R)-CH(Me)tBu and (+)-(R)-CH(Me)Ph result in the straightforward synthesis of the new, chiral ligands H2L2 and H2L3, respectively. Salt elimination reactions between K2L and divalent Mn and Fe halides, and protonolysis reactions between ZnMe2 and H2L result in the formation of the new dinuclear complexes [M2(L)2]. Investigation of the structures of these compounds in solution and in the solid state reveal that chiral mesocates are formed for L = L2, whereas for L = L3, a racemic mixture of helicates is present.(© Wiley-VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2007) [source] Theoretical Studies on Metal,Metal Interaction and Intrinsic 1,3[,*(d),(s/p)] Excited States of Dinuclear d10 Complexes with Bridging Phosphane LigandsEUROPEAN JOURNAL OF INORGANIC CHEMISTRY, Issue 5 2006Qing-Jiang Pan Abstract To explore the metal,metal interaction and spectroscopic properties, the ground- and excited-state structures of [M2(dpm)2]2+ [M = Ag (2), Cu (3), dpm = bis(diphosphanyl)methane] and their solvated species [M2(dpm)2]2+·(MeCN)2 were optimized by the MP2 and CIS methods, respectively. In the ground states, the calculated M,M distances and their corresponding M,M stretching frequencies for 2 and 3 indicate the presence of metallophilic attraction; there is strong N,Cu/Ag coordination in acetonitrile, which is different from the case in previous studies of [Au2(dpm)2]2+ (1). CIS calculations show that 2 and 3 have 1,3[,*(d),(s/p)] as their lowest-energy excited state, as is also the case for 1, confirmed by unrestricted MP2 calculations. On the basis of the CIS-optimized structures, the TD-DFT (B3LYP) method was employed to calculate the emission spectra of such complexes. For 3, the phosphorescent emissions were calculated at 424 and 514 nm in the solid state and acetonitrile, which is comparable to the experimental data of 475 and 480 nm, respectively. The comparison between the gas-phase and solution emissions for 1,3 reveals that the N,M coordination results in a large red-shift of the emission wavelength. Taking previous studies into account , we found that the M,M distances are linearly correlated with the M,M stretching frequencies for the dinuclear d10 complexes. (© Wiley-VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2006) [source] Non-Covalent Aggregation of Discrete Metallo-Supramolecular Helicates into Higher Assemblies by Aromatic Pathways: Structural and Chemical Studies of New Aniline-Based Neutral Metal(II) DihelicatesEUROPEAN JOURNAL OF INORGANIC CHEMISTRY, Issue 17 2005Miguel Vázquez Abstract Neutral manganese(II), iron(II), cobalt(II), nickel(II), zinc(II) and cadmium(II) complexes with an N -tosyl-substituted N4 -donor Schiff base containing a 4,4'-methylenedianiline residue as spacer [H2La: N,N' -bis(2-tosylaminobenzylidene)-4,4'-methylenedianiline], and the zinc(II) complex with an analogous ligand [H2Lb: N,N' -bis(2-tosylaminobenzylidene)-4,4'-oxodianiline] have been prepared by an electrochemical procedure. FAB and ESI mass spectra of the complexes show peaks due to species corresponding to a general formula [M2(La,b)2 + H]+, thereby suggesting their dinuclear nature. A detailed study of the crystal packing in the unit cell of the zinc(II) complex with H2La shows that the helicates aggregate to form discrete prismatic moieties containing three molecules held together by ,,, and ,,, interactions. Moreover, the ZnII neutral dihelicate with H2Lb forms a 3D network in the solid state due to intermolecular ,-stacking interactions. 1H NMR studies of the diamagnetic compounds reported herein have been performed. Finally, the ligand H2La and its ZnII and CdII complexes have been studied by spectrophotometric and spectrofluorimetric techniques in order to get a better understanding of the formation mechanisms of the complexes and of the nature of their fluorescence emission. Emission studies show that the ZnII and CdII dihelicates with H2La display a green fluorescence in acetonitrile solution (, = 473 nm, , = 0.03 and , = 476 nm, , = 0.01, respectively). (© Wiley-VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2005) [source] Synthesis, Structure and Spectroscopic Properties of Novel9,10-Bis{[6-(diphenylphosphanyl)-2-pyridylmethyl]propylaminomethyl}anthracene-Bridged Group 11 Metal (M = CuI and AgI) Dinuclear ComplexesEUROPEAN JOURNAL OF INORGANIC CHEMISTRY, Issue 14 2005Hu Xu Abstract Dinuclear complexes [M2(,-BPNNAn)]X2 (M = CuI, X = ClO4,1, X = BF4,2; M = AgI, X = BF4,3, X = ClO4,4) have been prepared by treating 9,10-bis{[6-(diphenylphosphanyl)-2-pyridylmethyl]propylaminomethyl}anthracene (BPNNAn) with M(CH3CN)4X in CH2Cl2. The structures of complexes 1, 2 and 3 have been determined by single-crystal X-ray diffraction studies. In the complexes, BPNNAn acts as a chelating ligand and two metal ions riding on two bridgehead carbon atoms in the anthracene group leads to the deformation of the anthracenyl ring. The Cu,Arene charge transfer interaction is observed from the nonfluorescent emission of Cu complex 1, while the Ag complex 3 exhibits dual emission at high concentrations in a CH2Cl2 solution. (© Wiley-VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2005) [source] Synthesis and Structural Characterization of Three-Coordinate MnII, FeII, and ZnII Complexes Containing a Bulky Diamide Ligand [DippN(CH2)3NDipp]2, (Dipp = 2,6- iPr2C6H3)EUROPEAN JOURNAL OF INORGANIC CHEMISTRY, Issue 24 2004Jianfang Chai Abstract The reaction of DippNH(CH2)3NHDipp (1) (Dipp = 2,6- iPr2C6H3) with 2 equiv. of MeLi in diethyl ether resulted in the formation of the monomeric dilithium salt [(Dipp)N(CH2)3N(Dipp)][Li(OEt2)]2 (2) in high yield. Further reactions of 2 with MnCl2, FeCl2, and ZnCl2, respectively, afforded the complexes [M2{N(Dipp)(CH2)3N(Dipp)}2] [M = Mn (3), Fe (4), Zn (5)] with three-coordinate metal centers. Complexes 2,5 were characterized by X-ray structural analysis. The complexes contain a nonplanar MNMN central core. The diamide ligand in complexes 3,5 displays a boat conformation and is both chelating and bridging, so that one of the nitrogen atoms is three-coordinate and the other four-coordinate. Complexes 3,5 are the first examples with diamide ligands in such a bonding mode. The magnetic investigations of compounds 3 and 4 reveal an antiferromagnetic exchange between the metal atoms. (© Wiley-VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2004) [source] Synthesis and Characterization of Mixed Phthalocyaninato and meso -Tetrakis(4-chlorophenyl)porphyrinato Triple-Decker Complexes , Revealing the Origin of Their Electronic AbsorptionsEUROPEAN JOURNAL OF INORGANIC CHEMISTRY, Issue 19 2004Xuan Sun Abstract Two series of mixed phthalocyaninato and porphyrinato rare earth(III) triple-decker complexes [M2(Pc)(TClPP)2] (1a,10a) and [M2(Pc)2(TClPP)] (1b,11b) [M = Y, La,Er except Ce and Pm; Pc = phthalocyaninate; TClPP = tetrakis(4-chlorophenyl)porphyrinate] have been prepared by treating the half-sandwich complexes [M(TClPP)(acac)] (acac = acetylacetonate), generated in situ from [M(acac)3]·nH2O and H2(TClPP), with Li2(Pc). All the triple-decker complexes have been characterized by a wide range of spectroscopic and electrochemical methods. The molecular structures of [M2(Pc)(TClPP)2] (M = Y, Ho) have also been determined, and show a symmetrical disposition of ligands, with two outer domed TClPP and one inner Pc rings. A systematic investigation of the optical and electrochemical data of these complexes has revealed the nature of the HOMO and LUMO, as well as the origin of the electronic absorptions of these triple-decker complexes. (© Wiley-VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2004) [source] Chain-Like Tetra-, Penta- and Heptanuclear Cyanide-Bridged Complexes by Attachment of Organometallic Cyanides to M2, M3 and M5 UnitsEUROPEAN JOURNAL OF INORGANIC CHEMISTRY, Issue 20 2003Tianlu Sheng Abstract Attachment of the "ligands" [Cp(dppe)Fe-CN], [Cp(PPh3)2Ru-CN], [Cp(CO)2Fe-CN] and [(CO)5M-CN],, (where M = Cr, Mo, W) to the polymetallic centres [Ru2(OAc)4]+, [Co3(dpa)4]2+ and [Ni5(tdpa)4]2+ yielded two tetra-, three penta- and five heptanuclear complexes with chain-like linear arrangements of the metal ions and the bridging cyanide ligands, as proved by the structure determination of [Ni5(tdpa)4{Cp(dppe)2Fe-CN}2](PF6)2. IR and CV data show that the Ru2, Co3 and Ni5 centres of these complexes withdraw electron density from the external organometallic units. This prevents electronic interactions between the two terminal metal ions, yet allows multistep one- and two-electron redox processes. (© Wiley-VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2003) [source] Acute action of rotenone on nigral dopaminergic neurons , involvement of reactive oxygen species and disruption of Ca2+ homeostasisEUROPEAN JOURNAL OF NEUROSCIENCE, Issue 10 2009Peter S. Freestone Abstract Rotenone is a toxin used to generate animal models of Parkinson's disease; however, the mechanisms of toxicity in substantia nigra pars compacta (SNc) neurons have not been well characterized. We have investigated rotenone (0.05,1 ,m) effects on SNc neurons in acute rat midbrain slices, using whole-cell patch-clamp recording combined with microfluorometry. Rotenone evoked a tolbutamide-sensitive outward current (94 ± 15 pA) associated with increases in intracellular [Ca2+] ([Ca2+]i) (73.8 ± 7.7 nm) and intracellular [Na+] (3.1 ± 0.6 mm) (all with 1 ,m). The outward current was not affected by a high ATP level (10 mm) in the patch pipette but was decreased by Trolox. The [Ca2+]i rise was abolished by removing extracellular Ca2+, and attenuated by Trolox and a transient receptor potential M2 (TRPM2) channel blocker, N -(p -amylcinnamoyl) anthranilic acid. Other effects included mitochondrial depolarization (rhodamine-123) and increased mitochondrial reactive oxygen species (ROS) production (MitoSox), which was also abolished by Trolox. A low concentration of rotenone (5 nm) that, by itself, did not evoke a [Ca2+]i rise resulted in a large (46.6 ± 25.3 nm) Ca2+ response when baseline [Ca2+]i was increased by a ,priming' protocol that activated voltage-gated Ca2+ channels. There was also a positive correlation between ,naturally' occurring variations in baseline [Ca2+]i and the rotenone-induced [Ca2+]i rise. This correlation was not seen in non-dopaminergic neurons of the substantia nigra pars reticulata (SNr). Our results show that mitochondrial ROS production is a key element in the effect of rotenone on ATP-gated K+ channels and TRPM2-like channels in SNc neurons, and demonstrate, in these neurons (but not in the SNr), a large potentiation of rotenone-induced [Ca2+]i rise by a small increase in baseline [Ca2+]i. [source] Negative cross-talk between presynaptic adenosine and acetylcholine receptorsEUROPEAN JOURNAL OF NEUROSCIENCE, Issue 1 2006A. V. Shakirzyanova Abstract Functional interactions between presynaptic adenosine and acetylcholine (ACh) autoreceptors were studied at the frog neuromuscular junction by recording miniature end-plate potentials (MEPPs) during bath or local application of agonists. The frequency of MEPPs was reduced by adenosine acting on presynaptic adenosine A1 receptors (EC50 = 1.1 µm) or by carbachol acting on muscarinic M2 receptors (EC50 = 1.8 µm). However, carbachol did not produce the depressant effect when it was applied after the action of adenosine had reached its maximum. This phenomenon implied that the negative cross-talk (occlusion) had occurred between A1 and M2 receptors. Moreover, the occlusion was receptor-specific as ATP applied in the presence of adenosine continued to depress MEPP frequency. Muscarinic antagonists [atropine or 1-[[2-[(diethylamino)methyl)-1-piperidinyl]acetyl]-5,11-dihydro-6H-pyrido [2,3-b][1,4]benzodiazepine-6-one) (AFDX-116)] had no effect on the inhibitory action of adenosine and adenosine antagonists [8-(p -sulfophenyl)theophylline (8-SPT) or 1,3-dipropyl-8-cyclopentylxanthine (DPCPX)] had no effect on the action of carbachol. These data suggested that membrane,delimited interactions did not occur between A1 and M2 receptors. Both carbachol and adenosine similarly inhibited quantal release triggered by high potassium, ionomycin or sucrose. These results indicated a convergence of intracellular pathways activated by M2 and A1 receptors to a common presynaptic effector located downstream of Ca2+ influx. We propose that the negative cross-talk between two major autoreceptors could take place during intense synaptic activity and thereby attenuate the presynaptic inhibitory effects of ACh and adenosine. [source] Modulation by adenosine of both muscarinic M1 -facilitation and M2 -inhibition of [3H]-acetylcholine release from the rat motor nerve terminalsEUROPEAN JOURNAL OF NEUROSCIENCE, Issue 11 2002Laura Oliveira Abstract The crosstalk between adenosine and muscarinic autoreceptors regulating evoked [3H]-acetylcholine ([3H]-ACh) release was investigated on rat phrenic nerve-hemidiaphragm preparations. Motor nerve terminals possess facilitatory M1 and inhibitory M2 autoreceptors that can be activated by McN-A-343 (1,30 µm) and oxotremorine (0.3,100 µm), respectively. The muscarinic receptor antagonist, dicyclomine (3 nm,10 µm), caused a biphasic (inhibitory/facilitatory) effect, indicating that M1 -facilitation prevails during 5 Hz stimulation trains. Concomitant activation of AF,DX 116-sensitive M2 receptors was partially attenuated, as pretreatment with M1 antagonists, muscarinic toxin 7 (MT-7, 0.1 nm) and pirenzepine (1 nm), significantly enhanced inhibition by oxotremorine. Activation of A2A -adenosine receptors with CGS 21680C (2 nm) (i) potentiated oxotremorine inhibition, and (ii) shifted McN-A-343-induced facilitation into a small inhibitory effect. Conversely, the A1 -receptor agonist, R- N6 -phenylisopropyl adenosine (R-PIA, 100 nm), attenuated the inhibitory effect of oxotremorine, without changing facilitation by McN-A-343. Synergism between A2A and M2 receptors is regulated by a reciprocal interaction with facilitatory M1 receptors, which may be prevented by pirenzepine (1 nm). During 50 Hz-bursts, facilitation (M1) of [3H]-ACh release by McN-A-343 disappeared, while the inhibitory (M2) effect of oxotremorine became predominant. This muscarinic shift results from the interplay with A2A receptors, as it was precluded by the selective A2A receptor antagonist, ZM 241385 (10 nm). In conclusion, when the muscarinic M1 positive feedback loop is fully operative, negative regulation of ACh release is mediated by adenosine A1 receptors. During high frequency bursts, tonic activation of A2A receptors promotes M2 autoinhibition by braking the M1 receptor operated counteraction. [source] New metabolic and pharmacokinetic characteristics of thiocolchicoside and its active metabolite in healthy humansFUNDAMENTAL & CLINICAL PHARMACOLOGY, Issue 4 2004M. Trellu Abstract Thiocolchicoside (TCC) has been prescribed for several years as a muscle relaxant drug, but its pharmacokinetic (PK) profile and metabolism still remain largely unknown. Therefore, we re-investigated its metabolism and PK, and we assessed the muscle relaxant properties of its metabolites. After oral administration of 8 mg (a therapeutic dose) of 14C-labelled TCC to healthy volunteers, we found no detectable TCC in plasma, urine or faeces. On the other hand, the aglycone derivative obtained after de-glycosylation of TCC (M2) was observed and, in addition, we identified, as the major circulating metabolic entity, 3-O-glucuronidated aglycone (M1) obtained after glucuro-conjugation of M2. One hour after oral administration, M1 plus M2 accounted for more than 75% of the circulating total radioactivity. The pharmacological activity of these metabolites was assessed using a rat model, the muscle relaxant activity of M1 was similar to that of TCC whereas M2 was devoid of any activity. Subsequently, to investigate the PK profile of TCC in human PK studies, we developed and validated a specific bioanalytical method that combines liquid chromatography and ultraviolet detection to assay both active entities. After oral administration, TCC was not quantifiable with an lower limit of quantification set at 1 ng/mL, whereas its active metabolite M1 was detected. M1 appeared rapidly in plasma (tmax = 1 h) and was eliminated with an apparent terminal half-life of 7.3 h. In contrast, after intramuscular administration both active entities (TCC and M1) were present; TCC was rapidly absorbed (tmax = 0.4 h) and eliminated with an apparent terminal half-life of 1.5 h. M1 concentration peaked at 5 h and this metabolite was eliminated with an apparent terminal half-life of 8.6 h. As TCC and M1 present an equipotent pharmacological activity, the relative oral pharmacological bioavailability of TCC vs. intramuscular administration was calculated and represented 25%. Therefore, to correctly investigate the PK and bioequivalence of TCC, the biological samples obtained must be assayed with a bioanalytical method able to specifically analyse TCC and its active metabolite M1. [source] PRDX4, a member of the peroxiredoxin family, is fused to AML1 (RUNX1) in an acute myeloid leukemia patient with a t(X;21)(p22;q22)GENES, CHROMOSOMES AND CANCER, Issue 4 2004Yanming Zhang The AML1 gene (also known as RUNX1) at 21q22 codes for core binding factor (CBF) ,, which forms a heterodimer with CBF , that acts as a transcriptional activating factor. CBF is a critical regulator in the generation and differentiation of definitive hematopoietic stem cells and is frequently disrupted in leukemia through chromosome translocations. We cloned a novel AML1 partner gene, PRDX4, in an X;21 translocation in a 74-year-old male patient diagnosed with acute myeloid leukemia,M2. Chromosome analysis detected a t(X;21)(p22;q22) as the sole abnormality in bone marrow samples. The involvement of AML1 was confirmed by fluorescence in situ hybridization studies. Using 3, RACE-PCR, we cloned a fusion between exon 5 of AML1 and exon 2 of PRDX4. RT-PCR confirmed the fusion and detected another fusion between exon 6 of AML1 and exon 2 of PRDX4, indicating alternative splicing of exon 6 of AML1 in the fusion transcripts. PRDX4 is one of six peroxiredoxin-family genes that are highly conserved in eukaryotes and prokaryotes and are ubiquitously expressed. Peroxiredoxin genes exhibit thioredoxin-dependent peroxidase activity and have been implicated in a number of other cellular functions such as cell proliferation and differentiation. PRDX4 plays a regulatory role in the activation of the transcription factor NF-,B and is significantly down-regulated in acute promyelocytic leukemia. This is the first example of antioxidant enzyme involvement in a chromosome translocation in leukemia. © 2004 Wiley-Liss, Inc. [source] Isolation and Structure Elucidation of Enniatins L, M1, M2, and N: Novel Hydroxy AnalogsHELVETICA CHIMICA ACTA, Issue 8 2004Pornrapee Vongvilai Four new cyclohexadepsipeptides, enniatins L (1), M1 (2), M2 (3), and N (4), have been isolated from an unidentified fungus (BCC 2629), together with the known enniatins B (5), H (6), and I (7), MK1688 (8), and enniatin B4 (9). Compounds 1,4 are the first enniatin analogs with an OH group at the side chain of one of the 2-hydroxycarboxylic acid residues. The structures of 1,4 were elucidated by spectroscopic means and by X-ray crystallography. [source] Genotype-dependent sensitivity of hepatitis C virus to inhibitors of the p7 ion channel,HEPATOLOGY, Issue 6 2008Stephen Griffin The hepatitis C virus (HCV) p7 protein plays a critical role during particle formation in cell culture and is required for virus replication in chimpanzees. The discovery that it displayed cation channel activity in vitro led to its classification within the "viroporin" family of virus-coded ion channel proteins, which includes the influenza A virus (IAV) M2 protein. Like M2, p7 was proposed as a potential target for much needed new HCV therapies, and this was supported by our finding that the M2 inhibitor, amantadine, blocked its activity in vitro. Since then, further compounds have been shown to inhibit p7 function but the relationship between inhibitory effects in vitro and efficacy against infectious virus is controversial. Here, we have sought to validate multiple p7 inhibitor compounds using a parallel approach combining the HCV infectious culture system and a rapid throughput in vitro assay for p7 function. We identify a genotype-dependent and subtype-dependent sensitivity of HCV to p7 inhibitors, in which results in cell culture largely mirror the sensitivity of recombinant protein in vitro; thus building separate sensitivity profiles for different p7 sequences. Inhibition of virus entry also occurred, suggesting that p7 may be a virion component. Second site effects on both cellular and viral processes were identified for several compounds in addition to their efficacy against p7 in vitro. Nevertheless, for some compounds antiviral effects were specific to a block of ion channel function. Conclusion: These data validate p7 inhibitors as prototype therapies for chronic HCV disease. (HEPATOLOGY 2008;48:1779-1790.) [source] DESCRIPTION OF A NEW SPECIES OF THE GENUS CULICOIDES (DIPTERA: CERATOPOGONIDAE) FROM XINGJUNG UYGUR AUT.INSECT SCIENCE, Issue 3 2001Abstract A new species of Culicoides, C. tahemanensis sp. nov. is described from Xingjiang Uygur Aut. Reg. in China. The new species is closely allied to Culicoides grisescens Edwards, but both of them are distinctly different in pale spots of cell R5 and cell A of wing of female, and shape of distal portion of aedeagus of male. The ninth tergum of new species is somewhat allied to C. nipponensis Tokunaga, but distinctly different in pale spots of cell R5, cell M2 and cell A of wing, and shape of aedeagus of male. The type speciemens are deposited in the Institute of Military Medical Sciences, Shenyang Military District, Shengyang 110034, China. [source] The effect of mineral trioxide aggregate on phagocytic activity and production of reactive oxygen, nitrogen species and arginase activity by M1 and M2 macrophagesINTERNATIONAL ENDODONTIC JOURNAL, Issue 8 2007T. M. B. Rezende Abstract Aim, To assess the influence of co-culture with mineral trioxide aggregate (MTA) on phagocytosis and the production of reactive oxygen intermediates (ROI) and nitrogen (NO) species and the arginase activity by M1 and M2 peritoneal macrophages. Methodology, Cellular viability, adherence and phagocytosis of Saccharomyces boulardii were assayed in the presence of MTA. Macrophages were stimulated with zymosan for ROI assays and with Fusobacterium nucleatum and Peptostreptococcus anaerobius and IFN- , for NO production and arginase activity, when in contact with capillaries containing MTA. Data were analysed by T, anova, Kruskall,Wallis and Mann,Whitney tests. Results, M2 macrophages displayed greater cellular viability in polypropylene tubes, greater ability to ingest yeast and smaller production of ROI and higher arginase activity when compared with M1 macrophages. Both macrophages, M1 and M2, presented similar cell adherence and NO production. The addition of bacterial preparations to macrophages interfered with NO and arginase productions. MTA did not interfere with any of the parameters measured. Conclusions, Phagocytosis and the ability of the two macrophage subtypes to eliminate microbes were not affected by MTA. [source] High-Temperature Ferromagnetism and Tunable Semiconductivity of (Ba, Sr)M2,±,xRu4,,,xO11 (M,=,Fe, Co): A New Paradigm for Spintronics,ADVANCED MATERIALS, Issue 7 2008Larysa Shlyk Ternary ruthenium ferrites (Ba,Sr)Fe2,±,xRu4,,,xO11 exhibit long-range ferromagnetic order well above room temperature along with favorable narrow- gap semiconducting properties. Electronic structure and physical properties can be tuned by simple chemical substitution of two elements, Fe and Co, or by varying the relative concentration of 3d and 4d elements within the homogeneity range. These promising properties within a single structural family open up a host of potential device applications. [source] CYP1A1 variants and smoking-related lung cancer in San Francisco bay area Latinos and African AmericansINTERNATIONAL JOURNAL OF CANCER, Issue 1 2005Margaret R. Wrensch Abstract We examined CYP1A1 T6235C (M1) and A4889G (M2) polymorphisms in San Francisco Bay Area African Americans and Latinos who were newly diagnosed with primary lung cancer from September 1998 to November 2002 and in age-gender-ethnicity frequency-matched controls. Owing mainly to rapid mortality of cases, overall percentages of cases genotyped were 26% and 32% for Latinos and African Americans, respectively. CYP1A1 variants were genotyped for Latinos (104 cases, 278 controls) and African Americans (226 cases, 551 controls). M1 and M2 frequencies in controls were 0.23 and 0.02 for African Americans and 0.38 and 0.29 for Latinos. In Latinos, the overall inverse odds ratio (OR) of 0.51 (95% CI = 0.32,0.81) for M1 variant genotype resulted from an inverse interaction with smoking. Nonsmokers with M1 genotype had a slight elevated OR (1.5; 0.59,3.7), but those with less than 30 or 30 or more pack-year history had 0.20 (0.06,0.70) and 0.21 (0.06,0.81) times (about 1/5) the odds expected if smoking and genotype were independent lung cancer risk factors. African Americans had interactions of similar magnitude that were not statistically significant. Results for M2 were very similar. Inverse interactions of CYP1A1 variants and smoking-associated lung cancer risk in Latinos might be causal, due to undetected bias or confounding, or represent a unique linkage disequilibrium between a new lung cancer locus and CYP1A1 in this highly admixed population. [source] Decreased pyruvate kinase M2 activity linked to cisplatin resistance in human gastric carcinoma cell linesINTERNATIONAL JOURNAL OF CANCER, Issue 4 2004Byong Chul Yoo Abstract Resistance to anticancer drugs is a major obstacle preventing effective treatment of disseminated cancers. Understanding the molecular basis to chemoresistance is likely to provide better treatment. Cell lines resistant to cisplatin or 5-fluorouracil (5-FU) were established from human gastric carcinoma cell lines SNU-638 and SNU-620. Comparative proteomics involving 2-dimensional gel electrophoresis (2-DE) and matrix-associated laser desorption ionization-mass spectroscopy (MALDI-MS) was performed on protein extracts from these parental and drug-resistant derivative lines to screen drug resistance-related proteins. Pyruvate kinase M2 (PK-M2) was identified as a protein showing lower expression in cisplatin-resistant cells compared to parental cells. Consistent with this finding, PK-M2 activity was also lower in cisplatin-resistant cells. Suppression of PK-M2 expression by antisense oligonucleotide resulted in acquired cisplatin resistance in SNU-638 cells. Furthermore, PK-M2 activity in 11 individual human gastric carcinoma cell lines positively correlated with cisplatin sensitivity. Taken together, PK-M2 protein and activity levels were lower in cisplatin-resistant human gastric carcinoma cell lines compared to their parental cell lines. Furthermore, suppression of PK-M2 expression using antisense oligonucleotides increased cisplatin resistance. These data clearly link PK-M2 and cisplatin resistance mechanisms. © 2003 Wiley-Liss, Inc. [source] Peroxidase activity, chlorophylls and antioxidant profile of two leaf vegetables (Solanum nigrum L. and Amaranthus cruentus L.) under six pretreatment methods before cookingINTERNATIONAL JOURNAL OF FOOD SCIENCE & TECHNOLOGY, Issue 1 2008Odunayo Clement Adebooye Summary The study evaluated the effects of six pretreatment methods before cooking on the peroxidase activity, chlorophyll and antioxidant profile of Amaranthus cruentus L. and Solanum nigrum L. The six pretreatments methods used were chopped only (raw sample) (coded M1); chopped and dried at 50 °C for 5 h (coded M2); chopped and squeezed in water (at room temperature) (coded M3); chopped and soaked in warm water (approximately 60 °C), then cooled and squeezed (coded M4); chopped and soaked in salt-treated water (approximately 20 g NaCl per litre of water) for 15 min, then squeezed (coded M5) and chopped and soaked in boiling water (100 °C), then cooled and squeezed (coded M6). The main effect of vegetable type and the main effect of pretreatment methods have significant effects (P 0.05) on the parameters measured, while the interaction of vegetable type and pretreatment methods have no significant effect on the parameters measured. Statistical analyses (P 0.05) showed that chlorophyll a and b occur in ratio 3:1 in the two vegetables, irrespective of the pretreatment imposed. Peroxidase activity test showed that A. cruentus, irrespective of the pretreatment imposed showed, no peroxidase activity, while S. nigrum showed high peroxidase activity for all the pretreatments except for M6. Results showed that there was a significantly (P 0.05) higher content of carotenoids in A. cruentus when compared with S. nigrum, while the total phenolics, total flavonoids and total tannins contents were higher in S. nigrum when compared with A. cruentus, irrespective of the pretreatment method used. For the two vegetables, the percentage losses in total carotenoids, phenolics, flavonoids and total tannins at M6 when compared with M1 were 53.3,60.5%, 55.6,57.1%, 62.4,63.6% and 66.1,73.5%, respectively. There was a sharp drop in the carotenoids, phenolics, flavonoids and tannins contents of the two vegetables at M4 and M6, with both treatments having closely similar values for each parameter. [source] FIAT represses bone matrix mineralization by interacting with ATF4 through its second leucine zipperJOURNAL OF CELLULAR BIOCHEMISTRY, Issue 3 2008Vionnie W.C. Yu Abstract We have characterized FIAT, a 66 kDa leucine zipper (LZ) protein that dimerizes with activating transcription factor 4 (ATF4) to form inactive dimers that cannot bind DNA. Computer analysis identifies three putative LZ motifs within the FIAT amino acid sequence. We have used deletion- and/or site-specific mutagenesis to individually inactivate these motifs in order to identify the functional LZ that mediates the FIAT,ATF4 interaction. Amino acids 194,222 that encode the FIAT LZ2 were deleted (mutant FIAT ZIP2 DEL). We inactivated each zipper individually by replacing two or three leucine residues within each zipper by alanine residues. The engineered mutations were L142A/L149A (mutant M1, first zipper), L208A/L215A/L222A (mutant M2, second zipper), and L441A/L448A (mutant M3, third zipper). MC3T3-E1 osteoblastic cells with an integrated 1.3 kb mouse osteocalcin gene promoter fragment driving expression of luciferase were transfected with expression vectors for ATF4 and the various FIAT deletion- or site-specific mutants. Inhibition of ATF4-mediated transcription was compared between wild-type (WT) and LZ FIAT mutants. The deletion mutant FIAT ZIP2 DEL and the sequence-specific M2 mutant did not interact with ATF4 and were unable to inhibit ATF4-mediated transcription. The M1 or M3 mutations did not affect the ability of FIAT to contact ATF4 or to inhibit its transcriptional activity. Stable expression of WT FIAT in osteoblastic cells inhibited mineralization, but not expression of the FIAT ZIP2 DEL and M2 mutants. This structure,function analysis reveals that FIAT interacts with ATF4 and modulates its activity through its second leucine zipper motif. J. Cell. Biochem. 105: 859,865, 2008. © 2008 Wiley-Liss, Inc. [source] Novel parameter for the diagnosis of distal middle cerebral artery stenosis with transcranial Doppler sonographyJOURNAL OF CLINICAL ULTRASOUND, Issue 8 2010Suk-Won Ahn MD Abstract Purpose Transcranial Doppler sonography (TCD) is commonly used for the diagnosis of middle cerebral artery (MCA) stenosis. However, TCD indices to predict distal MCA (M2) stenosis have not yet been established. We compared TCD and magnetic resonance angiography (MRA) to validate a new index for the diagnosis of M2 stenosis. Methods Consecutive patients who underwent TCD and MRA were included. Based on MRA, M2 stenosis was defined as >50% narrowing beyond the bifurcation area. TCD index of the M2/M1 ratio was defined as the ratio between the mean flow velocity (MFV) obtained at a depth of 30,44 mm (M2) and a depth of 45,65 mm (M1). Sensitivity and specificity of the M2/M1 ratio were calculated from the receiver operating characteristic curve. The diagnostic yield of elevated MFV (>80 cm/s) and asymmetry index of >30% for M2 stenosis were also investigated. Results Among the consecutive patients, 105 with M2 stenosis were compared with 123 without MCA stenosis. The M2/M1 ratio was significantly higher in the M2 stenosis group (1.10 versus 0.86, p < 0.001). Sensitivity and specificity for M2 stenosis were most satisfying when the M2/M1 ratio of 0.97 was adopted as the cutoff value. Diagnostic yield of the M2/M1 ratio was better than MFV or asymmetry index. Conclusions The M2/M1 ratio may be a highly specific parameter for assessing M2 stenosis with TCD. © 2010 Wiley Periodicals, Inc. J Clin Ultrasound 38:420,425, 2010 [source] | ||||||||||||||||||||||||||||||||||||||||