Dual roles of CagA protein in Helicobacterpylori-induced chronic gastritis in mice

Cytotoxin-associated gene A (CagA) acts directly on gastric epithelial cells. However, the roles of CagA in host adaptive immunity against Helicobacter pylori (H. pylori) infection are not fully understood. In this study, to investigate the roles of CagA in the development of H. pylori-induced chronic gastritis, we used an adoptive-transfer model in which spleen cells from C57BL/6 mice with or without H. pylori infection were transferred into RAG2^−/− mice, with gastric colonization of either CagA⁺H. pylori or CagA⁻H. pylori. Colonization of CagA⁺H. pylori but not CagA⁻H. pylori in the host gastric mucosa induced severe chronic gastritis in RAG2^−/− mice transferred with spleen cells from H. pylori-uninfected mice. In addition, when CagA⁺H. pylori-primed spleen cells were transferred into RAG2^−/− mice, CD4⁺ T cell infiltration in the host gastric mucosa were observed only in RAG2^−/− mice infected with CagA⁺H. pylori but not CagA⁻H. pylori, suggesting that colonization of CagA⁺H. pylori in the host gastric mucosa is essential for the migration of H. pylori-primed CD4⁺ T cells. On the other hand, transfer of CagA⁻H. pylori-primed spleen cells into CagA⁺H. pylori-infected RAG2^−/− mice induced more severe chronic gastritis with less Foxp3⁺ regulatory T-cell infiltration as compared to transfer of CagA⁺H. pylori-primed spleen cells. In conclusion, CagA in the stomach plays an important role in the migration of H. pylori-primed CD4⁺ T cells in the gastric mucosa, whereas CagA-dependent T-cell priming induces regulatory T-cell differentiation, suggesting dual roles for CagA in the pathophysiology of H. pylori-induced chronic gastritis.     

Isolation of two murine H1 histone genes and chromosomal mapping of the H1 gene complement

The mammalian H1 histone gene complement consists of at least seven H1 protein isoforms. These include five S-phase-dependent H1 protein subtypes and two more distantly related proteins, which are expressed upon terminal differentiation (H1⁰) or during the pachytene stage of spermatogenesis (H1t). In the past, three replication-dependent murine H1 genes plus the H1 ⁰ and H1t genes have been isolated and characterized. In this report, we describe the sequences of two more H1 genes, and we show that all five murine replication-dependent H1 genes and the H1t gene map to the region A2-3 on Chromosome (Chr) 13. This is in agreement with our previous finding that the human H1 histone gene complement maps to 6p21.3, which corresponds to the A2-3 region on the murine Chr 13. Previous reports have shown that the replication-independent H1 ⁰ genes map to syntenic regions on Chrs 22 (human H1⁰) and 15 (murine H1 ⁰).     

Roles for Nitric Oxide as an Intra- and Interneuronal Messenger at NMDA Release-Regulating Receptors: Evidence from Studies of the NMDA-Evoked Release of [3H]Noradrenaline and d-[3H]Aspartate from Rat Hippocampal Slices

Abstract: N-Methyl-d -aspartate (NMDA) receptors regulating the release of [³H]noradrenaline ([³H]NA) and d -[³H]aspartate (d -[³H]Asp) were investigated in superfused slices of rat hippocampus in the presence and absence of nitrergic drugs to examine a possible role for nitric oxide (NO) in the release process. In Mg²⁺-free Krebs-Henseleit buffer, the NMDA-evoked release of [³H]NA and d -[³H]Asp was Ca²⁺ dependent and inhibited by the NMDA antagonist (±)-3-(2-carboxypiperazin-4-yl)propenyl-1-phosphonic acid. NMDA-stimulated release of [³H]NA was tetrodotoxin (TTX; 0.1–2 µM) sensitive, whereas that for d -[³H]Asp was TTX insensitive, indicating that the NMDA receptors involved are differentially localized; those for d -[³H]Asp appear to be presynaptic, whereas those for [³H]NA are extrasynaptic in location. l -Arginine (100 µM), the natural precursor of NO synthesis, enhanced NMDA-evoked release of [³H]NA (100%) and d -[³H]Asp (700%). Exogenous NO donors—sodium nitroprusside, 3-morpholinosyndnomine, and S-nitroso-N-acetylpenicillamine (all 100 µM)—stimulated the NMDA-evoked release. An exception was the inhibition by nitroprusside of NMDA-evoked release of [³H]NA, where the presence of antioxidants may influence channel activity. Inhibitors of NO synthase (N^G-nitro-, N^G-methyl-, and N^G-amino-l -arginine, all 100 µM) attenuated (50–80%) the NMDA-stimulated release of [³H]NA and d -[³H]Asp, as did KN-62 (10 µM), a specific inhibitor of calmodulin kinase II. Our data support roles for the NO transducing system subsequent to the activation of NMDA release-regulating receptors as both an intraneuronal (presynaptically) and an extraneuronal messenger.     

铁筷子花瓣中花青素苷成分及含量对其呈色的影响北大核心CSCD

该研究以7个品种铁筷子(Helleborus thibetanus Franch.)为试验材料,借助目视测色、RHSCC比色卡、色差仪进行花色表型的测定,采用高效液相色谱法-光电二极管阵列检测方法(HPLC-DAD)及高效液相色谱-电喷雾离子化-质谱联用技术(HPLC-ESI-MS)测定分析铁筷子花瓣中花青素苷成分及含量,以探究不同品种铁筷子的花色与花青素苷成分及含量之间的关系。结果显示:(1)紫色系品种花瓣的a*值最高b*值最低,黄色系品种花瓣的b*值最高a*值最低,不同品种的铁筷子花色越深L*值越低。(2)从5个有花青素苷积累的铁筷子品种中检测出11种花青素苷成分,分别为6种矢车菊素苷,4种飞燕草素苷,1种矮牵牛素苷;供试的铁筷子材料中红色系2个品种的花青素苷含量最高,紫色系品种次之;矢车菊素苷与飞燕草素苷为影响铁筷子花瓣呈色的主要色素物质。(3)不同种类的花青素和修饰基团的差异,导致铁筷子花瓣呈现不同的色彩,含有多种酰基化修饰的飞燕草素苷使铁筷子花色蓝移进而使花色加深。(4)相关分析表明,铁筷子花瓣的L*值与a*值呈显著负相关关系,与b*值呈显著的正相关关系;L*值与总花青素苷含量呈显著负相关关系,且随着花青素苷含量的累积a*值增加,花色红移。研究表明,花青素苷的成分及含量是导致铁筷子花瓣呈现不同颜色的主要原因,矢车菊素苷和飞燕草素苷的互作以及酰基化的修饰使铁筷子呈现不同程度的紫色,花青素苷的不同累积量影响了花瓣颜色的明暗变化,从而使铁筷子花瓣颜色丰富。     

Effects of leaf water evaporative 2H‐enrichment and biosynthetic fractionation on leaf wax n‐alkane δ2H values in C3 and C4 grasses

Leaf wax n‐alkane δ²H values carry important information about environmental and ecophysiological processes in plants. However, the physiological and biochemical drivers that shape leaf wax n‐alkane δ²H values are not completely understood. It is particularly unclear why n‐alkanes in grasses are typically ²H‐depleted compared with plants from other taxonomic groups such as dicotyledonous plants and why C3 grasses are ²H‐depleted compared with C4 grasses. To resolve these uncertainties, we quantified the effects of leaf water evaporative ²H‐enrichment and biosynthetic hydrogen isotope fractionation on n‐alkane δ²H values for a range of C3 and C4 grasses grown in climate‐controlled chambers. We found that only a fraction of leaf water evaporative ²H‐enrichment is imprinted on the leaf wax n‐alkane δ²H values in grasses. This is interesting, as previous studies have shown in dicotyledonous plants a nearly complete transfer of this ²H‐enrichment to the n‐alkane δ²H values. We thus infer that the typically observed ²H‐depletion of n‐alkanes in grasses (as opposed to dicots) is because only a fraction of the leaf water evaporative ²H‐enrichment is imprinted on the δ²H values. Our experiments also show that differences in n‐alkane δ²H values between C3 and C4 grasses are largely the result of systematic differences in biosynthetic fractionation between these two plant groups, which was on average ?198‰ and?159‰ for C3 and C4 grasses, respectively.     

Characterization of the Ryanodine Receptor-Ca2+ Release Channel from the Thoracic Tissues of the Lepidopteran Insect Heliothis virescens

The existence of invertebrate forms of the RyR has recently been confirmed (Takeshima et al., 1994, Puente et al., 2000). However, information on the functional properties of this insect RyR is still limited. We report the functional characterization of a RyR from the thoracic muscle of H. virescens (Scott-Ward et al., 1997). A simple purification protocol produced membranes from homogenized prefrozen H. virescens thoracic muscle with a [³H]-ryanodine binding activity of 1.19 ± 0.21 pmol/mg protein (mean ±se; n= 4). [³H]-Ryanodine binding to the H. virescens receptor was dependent on the ryanodine concentration in a hyperbolic fashion with a K _D of 3.82 nm (n= 4). [³H]-ryanodine binding was dependent on [Ca²⁺] in a biphasic manner and was stimulated by 1 mm ATP. Millimolar caffeine did not stimulate [³H]-ryanodine binding to H. virescens membranes in the presence of either nanomolar or micromolar Ca²⁺. A protein of at least 400 KDa was recognized in H. virescens membrane proteins by a specific anti-H. virescens RyR antibody. Discontinuous density sucrose gradient fractionation of microsomal membranes produced vesicles suitable for single-channel studies. Ca²⁺-sensitive, Ca²⁺-permeable channels were successfully inserted into artificial lipid bilayers from H. virescens membrane vesicles. The H. virescens RyR-channel displayed a Ca²⁺ conductance of ∼110 pS and underwent a persistent and characteristic modification of ion handling and gating following addition of 100 nm ryanodine. The gating of H. virescens channels was sensitive to ATP and ruthenium red in a manner similar to mammalian RyR. This is the first report to describe the single channel and [³H]-ryanodine binding properties of a native insect RyR. Received: 3 July 2000/Revised: 17 October 2000     

Support for Radiolabeling of a Glycine Recognition Domain on the N-Methyl-d-Aspartate Receptor Ionophore Complex by 5,7-[3H]Dichlorokynurenate in Rat Brain

Abstract: Pretreatment with Triton X-100 more than doubled the binding of radiolabeled 5,7-dichlorokynurenic acid (DCKA), a proposed antagonist at a glycine (Gly) recognition domain on the N-methyl-d -aspartate (NMDA) receptor ionophore complex, in rat brain synaptic membranes. The binding exhibited an inverse temperature dependency, reversibility, and saturability, the binding sites consisting of a single component with a high affinity (27.5 nM) and a relatively low density (2.87 pmol/mg of protein). The binding of both [³H]DCKA and [³H]Gly was similarly displaced by numerous putative agonists and antagonists at the Gly domain in a concentration-dependent manner at a concentration range of 100 nM to 0.1 mM. Among the 24 putative ligands tested, DCKA was the second most potent displacer of the binding of both radioligands with no intrinsic affinity for the binding of [³H]kainic acid and α-amino-3-hydroxy-5-[³H]methylisoxazole-4-propionic acid (AMPA) to the non-NMDA receptors. In contrast, the other proposed potent Gly antagonist, 5,7-dinitroquinoxaline-2,3-dione, was active in displacing the binding of [³H]glutamic ([³H]Glu) and D,L-(E)-2-amino-4-[³H]propyl-5-phosphono-3-pentenoic acids to the NMDA recognition domain with a relatively high affinity for the non-NMDA receptors. In addition, the proposed antagonist at the AMPA-sensitive receptor, 2,3-dihydroxy-6-nitro-7-sulfamoyl-benzo(F)quinoxaline, not only displaced weakly the binding of both [³H]- Gly and [³H]DCKA, but also inhibited the binding of (+)-5-[³H]methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine ([³H]MK-801) to an ion channel associated with the NMDA-sensitive receptor in the presence of added Glu alone in a manner sensitive to antagonism by further added Gly. Clear correlations were seen between potencies of the displacers to displace [³H]DCKA binding and [³H]Gly binding, in addition to between the potencies to displace [³H]-DCKA or [³H]Gly binding and to potentiate or inhibit [³H]MK-801 binding. All quinoxalines tested were invariably more potent displacers of [³H]DCKA binding than [³H]Gly binding, whereas kynurenines were similarly effective in displacing the binding of both [³H]Gly and [³H]-DCKA. These results undoubtedly give support to the proposal that [³H]DCKA is one useful radioligand available in terms of its high selectivity and affinity for the Gly domain in the brain. Possible multiplicity of the Gly domain is suggested by the differential pharmacological profiles between the binding of [³H]Gly and [³H]DCKA.     

Role of mouse and human class II transgenes in susceptibility to and protection against mouse autoimmune nhyroiditis

 Mouse experimental autoimmune thyroiditis (EAT), a model for Hashimoto’s thyroiditis, is induced by immunizing with mouse thyroglobulin (MTg). To study the extent of H2A involvement in EAT, we introduced AaAb genes from susceptible k mice into resistant or intermediately susceptible strains which do not express H2E molecules. Thyroiditis was severe in resistant B10.M (H2 ^f ) mice carrying the double transgene Aa ^k Ab ^k . Likewise, thyroid infiltration was significantly extended in intermediate B10.Q (H2 ^q ) mice with the same transgene. To examine the effect of H2E molecules in the presence of H2A-mediated susceptibility, we introduced an Ea ^k transgene into E^– B10.S mice to express the Eβ^s molecule and observed significant reduction in EAT severity in B10.S(E⁺) mice. On the other hand, the presence of an Eb ^d transgene in B10.RQB3 (H2A ^q ) mice resulting in the expression of H2Eβ^d molecules did not alter EAT susceptibility, suggesting a role for Eb gene polymorphism in protection against EAT. We have shown recently that the HLA-DRB1 ^* 0301 (DR3) transgene conferred EAT susceptibility to B10.M as well as class II-negative B10.Ab⁰ mice. However, we report here that the HLA-DQB1 ^* 0601 (DQ6b) transgene in B10.M or HLA-DQA1 ^* 0301/DQB1 ^* 0302 (DQ8) transgene in class II-negative Ab⁰ mice did not. These studies show the differential effects of class II molecules on EAT induction. Susceptibility can be determined when class II molecules from a single locus, H2A or HLA-DQ, are examined in transgenic mice, but the overall effect may depend upon the presence of both class II molecules H2A and H2E in mice and HLA-DQ and HLA-DR in humans. Received: 28 January 1997 / 24 March 1997     

Convergent pathways of gibberellin A1 biosynthesis in Brassica

[²H, ³H]Gibberellin A₄ (GA₄) or [²H, ³H] GA₉ were applied to the shoot tips of seedlings of elongated internode (ein), a tall mutant of rapid cycling Brassica rapa. Following [²H]GA₉ application, [²H]GA₅₁, [²H]GA₂₀ and [²H]GA₄ were identified as products by GC-MS, while [²H]GA₃₄ and [²H]GA₁ were formed from [²H]GA₄. Other isotopically labelled products, including abundant putative conjugates, were also produced, but were not identified. Thus, in B. rapa, GA₁ biosynthesis involves the convergence of at least two metabolic pathways; it can be formed via GA₄ or GA₂₀, the latter of which can originate from GA₉ or from GA₁₉.     

Interleukin-2 Modulates Evoked Release of [3H]Dopamine in Rat Cultured Mesencephalic Cells

Abstract: Mesencephalic cell cultures were used as a model to investigate the effects of interleukin-2 (IL-2) on evoked release of [³H]dopamine ([³H]DA) and γ-[³H]-aminobutyric acid ([³H]GABA). At low concentrations (10^?13-10^?12M), IL-2 potentiated [³H]DA release evoked by the excitatory amino acids N-methyl-D-aspartate (NMDA) and kainate, whereas higher IL-2 concentrations (10^?9-10^?8M) had no effect. IL-2 (10^?14-10^?8M) modulated K⁺-evoked [³H]DA release in a biphasic manner, with low concentrations (10^?12-10^?11M) of IL-2 potentiating and higher concentrations (10^?9-10^?8M) inhibiting K⁺-induced [³H]DA release. IL-2 (10^?14-10^?8M) by itself failed to alter spontaneous [³H]DA release. The inhibition by IL-2 of K⁺-evoked [³H]DA release was reversible and not due to neurotoxicity, as preexposure to IL-2 (10^?8M) had no significant effect on the subsequent ability of dopaminergic cells to take up and to release [³H]DA. Under our experimental conditions, IL-2 (10^?8 M) did not alter Ca²⁺-independent [³H]GABA release evoked by either K⁺ or NMDA. The results of this study indicate that IL-2 is able to potentiate [³H]DA release evoked by a number of different stimuli, including K⁺ depolarization and activation of both NMDA and non-NMDA receptor subtypes in mesencephalic cell cultures. IL-2 is active at very low concentrations, a finding that indicates a potent effect of IL-2 on dopaminergic neurons and implicates a physiological role for this cytokine in the modulation of DA release.     

Opiate Receptor-Mediated Inhibition of Catecholamine Release in Primary Cultures of Bovine Adrenal Chromaffin Cells

Abstract: Primary cultures of chromaffin cells from bovine adrenal medulla were used to evaluate the ability of several opiates to reduce the release of catecholamines induced by stimulation of nicotinic receptors. Etorphine, β-endorphin, Met-enkephalin[Arg⁶,Phe⁷], and the synthetic peptide [d -Ala²,Me-Phe⁴,Met(O)^s-ol]enkephalin inhibited the acetylcholine-induced release of catecholamines with an IC₃₀ varying from 10^?7 to 1 × 10^?6M. The effect was stereospecific because levorphanol (IC₃₀= 7.5 × 10^?7M) was approximately two orders of magnitude more potent than dextrorphan. Morphine (μ-receptor agonist), [d -Ala², d -Leu⁵]enkephalin (δ-receptor agonist), ethylketazocine (k -receptor agonist), and N-allylnormetazocine (σ-receptor agonist) were at least 100–1000 times less potent than etorphine. Diprenorphine (IC₅₀= 5 × 10^?7M) and naloxone (IC₅₀= 10^?6M) antagonized the effect of etorphine. High-affinity, saturable, and stereospecific binding sites for [³H]etorphine, [³H]dihydromorphine, [³H-d -Ala²,d -Leu⁵]enkephalin, [³H]ethylketazocine, and for [³H]N-allylnormetazocine, [³H]diprenorphine, and [³H]naloxone were detected in chromaffin cell membranes and in membranes obtained from adrenal medulla homogenates. However, the number of binding sites for [³H]etorphine and [³H]diprenorphine was 10–70 times higher than the number of sites measured with the other ³H ligands. The rank order of potency of these compounds for the displacement of [³H]etorphine binding correlates (r = 0.90) with the rank order of potency of the same compounds for the inhibition of acetylcholine-induced catecholamine release. These data suggest that a stereoselective opiate receptor (different from the classic μ-, δ-, k -, or σ-receptor) with high affinity for etorphine, diprenorphine, β-endorphin, and Met-enkephalin[Arg⁶,Phe⁷] modulates the function of the nicotinic receptor in adrenal chromaffin cells.     

Characterisation of the Binding of [3H]WAY-100635, a Novel 5-Hydroxytryptamine1A Receptor Antagonist, to Rat Brain

Abstract: The specific binding of [³H]WAY-100635 {N-[2-[4-(2-[O-methyl-³H]methoxyphenyl)-1-piperazinyl]ethyl]-N-(2-pyridinyl)cyclohexane carboxamide trihydrochloride} to rat hippocampal membrane preparations was time, temperature, and tissue concentration dependent. The rates of [³H]WAY-100635 association (k₊₁ = 0.069 ± 0.015 nM^?1 min^?1) and dissociation (k_?1 = 0.023 ± 0.001 min^?1) followed monoexponential kinetics. Saturation binding isotherms of [³H]WAY-100635 exhibited a single class of recognition site with an affinity of 0.37 ± 0.051 nM and a maximal binding capacity (B_max) of 312 ± 12 fmol/mg of protein. The maximal number of binding sites labelled by [³H]WAY-100635 was ～36% higher compared with that of 8-hydroxy-2-(di-n-[³H]-propylamino)tetralin ([³H]8-OH-DPAT). The binding affinity of [³H]WAY-100635 was significantly lowered by the divalent cations CaCl₂ (2.5-fold; p < 0.02) and MnCl₂ (3.6-fold; p < 0.05), with no effect on B_max. Guanyl nucleotides failed to influence the K_D and B_max parameters of [³H]WAY-100635 binding to 5-HT_1A receptors. The pharmacological binding profile of [³H]WAY-100635 was closely correlated with that of [³H]8-OH-DPAT, which is consistent with the labelling of 5-hydroxytryptamine_1A (5-HT_1A) sites in rat hippocampus. [³H]WAY-100635 competition curves with 5-HT_1A agonists and partial agonists were best resolved into high- and low-affinity binding components, whereas antagonists were best described by a one-site binding model. In the presence of 50 µM guanosine 5′-O-(3-thiotriphosphate) (GTPγS), competition curves for the antagonists remained unaltered, whereas the agonist and partial agonist curves were shifted to the right, reflecting an influence of G protein coupling on agonist versus antagonist binding to the 5-HT_1A receptor. However, a residual (16 ± 2%) high-affinity agonist binding component was still apparent in the presence of GTPγS, indicating the existence of GTP-insensitive sites.     

Transport, degradation and cell wall-integration of XXFGol, a growth-regulating nonasaccharide of xyloglucan, in pea stems

When [glucitol-³H]XXFGol (a NaB³H₄-reduced xyloglucan nonasaccharide) was applied to excised shoots of pea (Pisum sativum L. cv. Progress) at the base of the epicotyl, it inhibited growth in the elongation zone, 4–5 cm distal. Experiments were conducted to discover whether such ³H-oligosaccharides are translocated intact over this distance, or whether an intercellular second messenger would have to be postulated. After 24 h, ³H from [glucitol-³H]XXFGol and [glucitol-³H]XXXGol showed U-shaped distributions, with most ³H at the base and apex of the stem. Radioactivity from [fucosyl-³H]XXFG and [xylosyl-³H]XXFG also moved acropetally, but did not concentrate at the apex, possibly owing to removal from the transpiration stream of fucose and xylose formed by partial hydrolysis of XXFG en route. When 10⁻⁷ M [glucitol-³H]XXFGol was supplied, about 14 fmol ·  seedling^–1 of apparently intact [³H]XXFGol was extractable from the elongation zone after 24 h. Larger amounts of degradation products were extractable (including free [³H]glucitol) and some wall-bound ³H-hemicellulose was present (presumably formed by the oligosaccharides acting as acceptor substrates for transglycosylation). We conclude that biologically active oligosaccharides of xyloglucan can move through the stem acropetally and that they are maintained at low steady-state concentrations by both hydrolysis and transglycosylation. Received: 1 April 1997 / Accepted: 28 May 1997     

Lack of influence of photoperiod on the metabolism of gibberellin A20 in Salix pentandra

The influence of photoperiod on the metabolism of GA₂₀ in Salix pentandra was studied by feeding [³H]-GA₂₀ to seedlings which had been grown previously under long day (LD) or short day (SD) conditions. After 48 h in LD or SD, metabolites were separated on sequential, silica gel partition columns and reversed-phase C₁₈ HPLC. The principal metabolite co-chromatographed with [³H]-GA₁ and this conversion was confirmed by feeding [²H]-GA₂₀, which was converted to [²H]-GA₁ as identified by gas chromatography-selected ion monitoring. Chromatographic evidence also indicated the minor conversion of [³H]-GA₂₀ to [³H]-GA₈ (via [³H]-GA₁) and trace conversion to [³H]-GA₂₉ (GAs A_1.8,20.29 are native in Salix). Ethyl acetate-insoluble [³H] metabolites were formed and could be cleaved by cellulase to release putative [³H]-GA₂₀ and [³H]-GA₁ suggesting the conversion to glucosyl conjugates of these GAs. Metabolism of [³H]-GA₂₀ was slightly more rapid in plants previously grown under LD than SD, an effect which reflected the generally increased shoot growth under LD. However, altering the photoperiod after [³H]-GA₂₀ addition had only a slight effect on the metabolism of [³H]-GA₂₀ in Salix seedlings. This indicates that the conversion of GA₂₀ to GA₁ is not a controlling step in the photoperiodic regulation of growth cessation in Salix.     

Proton/hydroxide conductance through lipid bilayer membranes

Summary A simple method of measuring proton/hydroxide conductance (G _H/OH) through planar lipid bilayer membranes is described. First the total conductance (G _m ) is measured electrically. Then the H⁺/OH^– transference number (T _H/OH) is estimated from the diffusion potential (V _m ) produced by a transmembrane pH gradient. The pH gradient is produced by a pair of buffered solutions which have identical concentrations of all ions except H⁺ and OH^–. Thus,V _m is due entirely to H⁺/OH^– diffusion andG _H/OH can be calculated from the relations,V _m =T _H/OH E _H/OH andG _H/OH=T _H/OH G _m , whereE _H/OH is the equilibrium potential for H⁺ and OH^–. In bilayers made from bacterial phosphatidylethanolamine (PE) inn-decane,G _H/OH is nearly independent of pH, ranging from about 10^–9 S cm^–2 at pH 1.6 to 10^–8 S cm^–2 at pH 10.5. BecauseG _H/OH is nearly independent of pH, the calculated permeability coefficients to H⁺ and/or OH^– are extremely pH dependent, which partly explains the wide range of values reported for phospholipid vesicles and biological membranes.G _H/OH appears to be independent of the membrane surface charge, because titrating either the phosphate or the amino group of PE has little effect onG _H/OH.G _H/OH is reduced about 10-fold when the water activity is reduced 33% by replacement with glycerol. Although the mechanism of H⁺/OH^– conductance is not known, the relation betweenG _H/OH and water activity suggests that several water molecules are involved in the H⁺/OH^– transport process.     

Microcalorimetric studies of the action of Er3+ on Halobacterium halobium R1 growth

A microcalorimetric technique was used to evaluate the influence of Er³⁺ on Halobacterium halobium R1 growth. By means of a LKB-2277 Bioactivity Monitor ampoule method, we obtained the thermogenic curves of H. halobium R1 growth at 37°C. In order to analyze the results, the relationship between k and C was obtained. The addition of Er³⁺ in low concentration cause a decrease of the maximum heat production P _max and growth rate constants k; however, Er³⁺ in a high concentration might promote growth of H. halobium R1. When Er³⁺ is in a much higher concentration, the growth of H. halobium R1 is inhibited completely. For comparison, the shapes of H. halobium R1 cells were observed by means of transmission electron microscope (TEM). According to the thermogenic curves and TEM photos of H. halobium R1 under different conditions, it is clear that the metabolic mechanism of H. halobium R1 growth has been changed with the addition of Er³⁺.     

An NMR investigation of putative interresidue H-bonding in methyl α-cellobioside in solution

Methyl α-cellobioside (methyl β-d-glucopyranosyl-(1→4)-α-d-glucopyranoside) was labeled with ¹³C at C4′ for use in NMR studies in DMSO-d₆ solvent to attempt the detection of a trans-H-bond J-coupling (^3hJ_CCOH) between C4′ and OH3. Analysis of the OH3 signal at 600 MHz revealed only the presence of two homonuclear J-couplings: ³J_H3,OH3 and a smaller, longer range J_HH. No evidence for ^3hJ_C4′,OH3 was found. The longer range J_HH was traced to ⁴J_H4,OH3 based on 2D ¹H–¹H COSY data and inspection of the H2 and H4 signal lineshapes. A limited set of DFT calculations was performed on a methyl cellobioside mimic to evaluate the structural dependencies of ⁴J_H2,O3H and ⁴J_H4,O3H on the H3–C3–O3–H torsion angle. Computed couplings range from about −0.7 to about +1.1 Hz, with maximal values observed when the C–H and O–H bonds are roughly diaxial.     

Helicobacter pylori infection can change the intensity of gastric Lewis antigen expressions differently between adults and children

This study tested whether there were different expressions of gastric Lewis antigens between children and adults with Helicobacter pylori infection, and whether the difference was related to the infection outcome. About 68 dyspeptic children and 110 dyspeptic adults were enrolled to check H. pylori infection, its colonization density, and the related histology. Gastric Lewis antigens b (Le^b), x (Le^x), and sialyl-Lewis x (sialyl-Le^x) were immunohistochemically stained and scored for the intensity. The H. pylori-infected adults, but not the children, had a lower Le^b intensity over the antrum (p = 0.019) but higher Le^b intensity over the corpus (p = 0.001) than the non-infected ones. Over the antrum, both the H. pylori-infected children and adults had a lower Le^x and higher sialyl-Le^x intensity than those non-infected ones (p < 0.05). The H. pylori-infected adults had a higher bacterial density (p = 0.004) and Le^b intensity (p = 0.016) over the corpus than the H. pylori-infected children. For the H. pylori-infected adults, but not children, the corpus had a higher Le^b (p = 0.038) and lower Le^x (p = 0.005) intensity than the antrum. Furthermore, the H. pylori-infected adults expressed a higher Le^b and had a higher bacterial density than those with weak Le^b (antrum, p < 0.001; corpus, p = 0.001). In conclusion, H. pylori infection is associated with the intensity change of Lewis antigen expressions in the stomach. The changes of gastric Lewis antigen expressions are different between adults and children with H. pylori infection, which may exert different H. pylori colonization over the corpus between adults and children.     

Radioactive contamination of the marine environment: Uptake and distribution of3H inDunaliella bioculata

The marine flagellateDunaliella bioculata, which is easily cultivated under laboratory conditions, is a suitable organism for assessing the importance of the radioactive contamination by³H bound to organic molecules. We have studied the uptake of the following tritiated precursors: thymidine-methyl-³H, adenine-2-³H, uridine-5-³H, l-leucine-4-³H, glycine-2-³H, l-arginine-3.4-³H, 1-aspartic acid-2. 3-³H, 1-phenylalanine-2.3-³H, D-glucose-2-³H and D-glucose-6-³H. Under the experimental conditions (2000 lux; incubation time 30 min), all tritiated molecules are taken up byD. bioculata. Their intracellular concentration may reach that of the external medium. However, leucine and adenine accumulate in the algae: their respective concentrations are 10 and 100 times higher than in the culture medium. The molecular distribution of³H has been studied by various biochemical techniques and by sieve chromatography on sepharose 4B. It has been found that more l-leucine-4-³H is incorporated into acid and acetone soluble substances than into proteins. Adenine-2-³H is mainly incorporated into macromolecules of biological significance (RNA, DNA). CsCl gradient centrifugation has shown that the total DNA ofDunaliella is constituted by a major (ϖ=1.707 g/cm³) and by a minor (ϖ=1.693 g/cm³) component. Fellow of the Commission of the European Communities     

Binding of [3H] γ-Aminobutyric Acid and [3H]Muscimol in Purified Rat Brain Synaptic Plasma Membranes and the Effects of Bicuculline

Abstract: The binding of [³H] γ-aminobutyric acid ([³H]GABA) and [³H]muscimol has been studied in purified synaptic plasma membrane (SPM) preparations from rat brain. Scatchard analysis of specific binding (defined as that displaced by 100 μMγ-aminobutyrate) indicated that the binding of both radiolabelled ligands was best described by a two component Langmuir adsorption isotherm. The apparent K_D and B_max values for [³H]GABA at 4°C were K_D1, 20 nM; K_D2,165 nM; B_max1, 0.48 pmol;B_max2, 6.0 pmol. mg^?1; for [³H]muscimol at 4°C they were: K_D1, 1.75 nM; K_D2, 17.5 nM; B_maxl, 0.84 pmol. mg^?1; B_max2, 4.8 pmol.mg^?1; and for [³H]muscimol at 37°C they were: K_D1, 7.0 nM; K_m, 60 nM; B_max], 0.5 pmol-mg^?1; B_max2, 7.2 pmol-mg¹. Under the experimental conditions used, the similar B_milx values for [³H]GABA and [³H]muscimol binding to the SPM preparations suggests that the high- and low-affinity components for the two radiolabeled ligands are identical. The effects of the GAB A antagonist bicuculline on the binding of [³H]muscimol at 4^CC and 37°C were studied. At 4°C, antagonism of muscimol binding appeared to be competitive at the high-affinity site but noncompetitive at the low-affinity site. At 37°C, antagonism was again competitive at the high-affinity site but was of a mixed competitive/noncompetitive nature at the low-affinity site. Assuming that binding to the high-affinity site is associated with the pharmacological actions of bicuculline, the apparent K_D values obtained suggest a pA₂ value of 5.3 against [³H]muscimol at 4°C and 37°C. This figure is in good agreement with several estimates of the potency of bicuculline based on pharmacological measurements. Results from displacement studies using [³H]GABA and [³H]muscimol suggest that [³H]GABA might be a more satisfactory ligand than [³H]muscimol in GABA radioreceptor assays.