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1.
The effects of synthetic rat adrenomedullin (rAM), a novel vasorelaxant peptide originally isolated from human pheochromocytoma, on receptor binding and cAMP generation were studied in cultured rat vascular smooth muscle cells (VSMC). A binding study using [ 125I]rAM revealed the presence of a single class of high-affinity ( Kd1.3 × 10 −8 M) binding sites for rAM in VSMC. The apparent Ki of rat calcitonin gene-related peptide (rCGRP) was 3 × 10 −7 M. Affinity labeling of VSMC membranes with [ 125I]rAM revealed two distinct labeled bands with apparent molecular weights of 120 and 70 kDa, both of which were abolished by excess unlabeled rAM or rCGRP. rAM stimulated cAMP formation with an approximate EC 50 of 10 −8 M, the effect of which was additive with isoproterenol, but not with rCGRP. The rAM-induced cAMP response was unaffected by propranalol, indomethacin, or quinaerine, but inhibited by a CGRP receptor antagonist, human CGRP[8–37]. These data suggest that VSMC possesses specific AM receptors functionally coupled to adenylate cyclase with which CGRP interacts. 相似文献
2.
Ethyl N-methyl-4-hydroxy-5-oxo-3-pyrroline-3-carboxylate forms a deep red chelate with iron salts. The color intensity is directly related to the iron concentration. The photosta-bility of the red color was determined at pH 1.2 and 5 by spectrophotometric assay at 484 nm at intervals during irradiation by tungsten light at 1020 μW/cm 2. After 528 hr of continuous irradiation in deionized water, 90.9% of the iron chelate had decomposed. The reaction followed zero order kinetics. Maximal stability was observed at pH 5 at both 10 -- 2 and 10 -- 2 molar concentrations of the iron chelate: no detectable decomposition occurred after 192 hr of continuous irradiation. The iron chelate in biological tissues is stable for 18 months. The staining technique is superior to other histological methods for estimating low concentrations of iron in tissue. 相似文献
3.
Reactive oxygen is an important regulator of vascular cell biology; however, the mechanisms involved in transducing signals from oxidants in endothelial cells are poorly defined. Because protein phosphorylation is a major mechanism for signal ransduction, cultured aortic endothelial cells were exposed to nonlethal concentrations of H 2O 2 to examine oxidant-sensitive changes in phosphorylation state. Addition of H 2O 2 increases the phosphorylation of the heat shock protein 27 (HSP27) within 2 min. This response is maximal by 20 min and remains constant for more than 45 min. Levels of intrcellular free Ca 2+ in endothelial cells did not change following addition of 100 μM H 2O 2, nor did the ability of the cells to respond to bradykinin. H 2O 2-induced phosphorylations were either not affected or were slightly increased in cells pretreated with PKC inhibitors (H-8, staurosporin, or calphostin c). Two-dimensional analysis of phosphoproteins from homogenates of 32P-labeled cells revealed that phorbol myristate acetate (PMA) did not cause the same degree of HSP27 phosphorylation as H 2O 2. Simultaneous addition of 10 ηM PMA and 50 μM H 2O 2 decreased the oxidant-stimulated phoshorylation of the most acidic HSP27 isoform. These data suggest that signal transduction for H 2O 2-sensitive endothelial cell responses are not only independent of PKC, but may also be suppressed by the action of the kinase. 相似文献
4.
The toxicity of H 2O 2 in Escherichia coli wild type and superoxide dismutase mutants was investigated under different experimental conditions. Cells were either grown aerobically, and then treated in M9 salts or K medium, or grown anoxically, and then treated in K medium. Results have demonstrated that the wild type and superoxide dismutase mutants display a markedly different sensitivity to both modes of lethality produced by H 2O 2 (i.e. mode one killing, which is produced by concentrations of H 2O 2 lower than 5 mM, and mode two killing which results from the insult generated by concentrations of H 2O 2 higher than 10 mM). Although the data obtained do not clarify the molecular basis of H 2O 2 toxicity and/or do not explain the specific function of superoxide ions in H 2O 2-induced bacterial inactivation, they certainly demonstrate that the latter species plays a key role in both modes of H 2O 2 lethality. A mechanism of H 2O 2 toxicity in E. coli is proposed, involving the action of a hypothetical enzyme which should work as an O 2-• generating system. This enzyme should be active at low concentrations of H 2O 2 (<5 mM) and high concentrations of the oxidant (>5 mM) should inactivate the same enzyme. Superoxide ions would then be produced and result in mode one lethality. The resistance at intermediate H 2O 2 concentrations may be dependent on the inactivation of such enzyme with no superoxide ions being produced at levels of H 2O 2 in the range 5–10 mM. Mode two killing could be produced by the hydroxyl radical in concert with superoxide ions, chemically produced via the reaction of high concentrations of H 2O 2 (>10 mM) with hydroxyl radicals. The rate of hydroxyl radical production may be increased by the higher availability of Fe 2+ since superoxide ions may also reduce trivalent iron to the divalent form. 相似文献
5.
1. Single reduced methyl viologen (MV .+) acts as an electron donor in a number of enzyme systems. The large changes in extinction coefficient upon oxidation (λ max 600 nm; MV .+, = 1.3 · 10 4 M −1 · cm −1; oxidised form of methyl viologen (MV 2+), = 0.0) make it ideally suited to kinetic studies of electron transfer reactions using stopped-flow and standard spectrophotometric techniques. 2. A convenient electrochemical preparation of large amounts of MV.+ has been developed. 3. A commercial stopped-flow apparatus was modified in order to obtain a high degree of anaerobicity. 4. The reaction of MV.+ with O2 produced H2O2 (k > 5 · 106 M−1 · s−1, pH 7.5, 25 °C). H2O2 subsequently reacted with excess MV.+ (k = 2.3 · 103 M−1 · s−1, pH 7.5, 25 °C) to produce water. The kinetics of this reaction were complex and have only been interpreted over a limited range of concentrations. 5. The results support the theory that the herbicidal action of methyl viologen (Paraquat, Gramoxone) is due to H2O2 (or radicals derived from H2O2) induced damage of plant cell membrane. 相似文献
6.
Amylin binding sites in a human hepatoblastoma cell line (HepG2) have been characterized in detail. 125I-Amylin (rat) bound to HepG2 cells with high affinity. Binding was reversible and selective, and dependent on time and temperature. Scatchard analysis revealed the presence of high ( Kd = 0.11 ± 0.04 n M) and low ( Kd = 1.3 ± 0.4 μ M) affinity binding sites for 125I-amylin in HepG2 cells. The dissociation experiments also showed that 125I-amylin dissociated from high- and low-affinity sites. The association data, however, indicated the presence of only one binding site. Rat amylin was more potent than human amylin and rat calcitonin gene-related peptide (CGRP) in displacing 125I-amylin bound to HepG2 cells. Nonhomologous peptides did not displace 125I-amylin. Rat amylin was, however, less potent than rat CGRP in displacing 125I[Tyr 0]CGRP from HepG2 cells. Pretreatment of HepG2 cells with rat amylin (10 n M) reduced the specific binding of 125I-amylin by 75%, whereas rat CGRP (10 n M) pretreatment had no effect on amylin binding. Calcitonin gene-related peptide, as well as rat and human amylin, stimulated the adenylate cyclase activity of HepG2 cell membrane preparation in a dose-dependent manner, with an order of potency of CGRP > rat amylin > human amylin. A CGRP antagonist, CGRP(8–37), significantly attenuated the stimulatory effect of both amylin and CGRP on adenylate cyclase activity. These investigations show that distinct receptors of amylin and CGRP are present in HepG2 cells and that amylin stimulates adenylate cyclase activity through CGRP receptors. This system could now be exploited for studying amylin receptors and amylin-mediated signal transduction. 相似文献
7.
Carbohydrate-bearing polymers of biologically inert design are versatile tools to delineate functional aspects of oligosaccharides. Binding of synthetic N-substituted polyacrylamide (PAA) conjugates of histo-blood group (A di, A tri, B di, B tri, H di, SiaLe a, and SiaLe x) to human polymorphonuclear leukocytes (PMNs), and effects on H 2O 2 generation elicited by different agonists such as digitonin, N-formyl-Met-Leu-Phe (FMLP) and the galactoside-specific lectin from Viscum album L. (VAA) were assessed. PMNs expressed binding sites for blood group-related neoglycoconjugates in the range of N10 6–10 7/cell with KD-values in the μM range. Treatment of PMNs (2×10 6 cells/ml) with PAA-probes (50 μg/ml) for 5 min did not activate the “respiratory burst”. However, it led to suppression (range 20–70%) of H 2O 2 generation of cells in the presence of elicitors. In detail, the FMLP-induced response was significantly decreased by A di, A tri, B tri, H di, SiaLe a, and SiaLe x conjugates, whereas for digitonin one only by A di, A tri, B tri. All the seven tested PAA-probes were found to inhibit significantly VAA-mediated release of H 2O 2 from PMNs. In this case, interference can take place already, at the stage of initial binding, especially for B- and H-epitopes, but less prominently for A- and SiaLe-epitopes. These results support the notion that PAA-immobilized histo-blood group oligosaccharides can serve as effector molecules with the ability to reduce the H 2O 2-generation of PMNs, warranting further studies on the involved reaction pathway. 相似文献
8.
We measured the toxicity and mutagenicity induced in human diploid lymphoblasts by various radiation doses of X-rays and two internal emitters. [ 125I]iododeoxyuridine ([ 125I]dUrd) and [ 3H]thymidine ([ 3H]TdR), incorporated into cellular DNA. [ 125I]dUrd was more effective than [ 3H]TdR at killing cells and producing mutations to 6-thioguanine resistance (6TG R). No ouabain-resistant mutants were induced by any of these agents. Expressing dose as total disintegrations per cell (dpc), the D0 for cell killing for [ 125I]dUrd was 28 dpc and for [ 3H]TdR was 385 dpc. The D0 for X-rays was 48 rad at 37°C. The slopes of the mutation curves were approximately 75 × 10 −8 6TG R mutants per cell per disintegration for [ 125I]dUrd and 2 × 10 −8 for [ 3H]TdR. X-Rays induced 8 × 10 −8 6TG R mutants per cell per rad. Normalizing for survival, [ 125I]dUrd remained much more mutagenic at low doses (high survival levels) than the other two agents. Treatment of the cells at either 37°C or while frozen at −70°C yielded no difference in cytotoxicity or mutation for [ 125I]dUrd or [ 3H]TdR, whereas X-rays were 6 times less effective in killing cells at −70°C. Assuming that incorporation was random throughout the genome, the mutagenic efficiencies of the radionuclides could be calculated by dividing the mutation rate by the level of incorporation. If the effective target size of the 6TGR locus is 1000–3000 base pairs, then the mutagenic efficiency of [125I]dUrd is 1.0–3.0 and of [3H]TdR is 0.02–0.06 total genomic mutations per cell per disintegration. 125I disintegrations are known to produce localized DNA double-strand breaks. If these breaks are potentially lethal lesions, they must be repaired, since the mean lethal dose (D0) was 28 dpc. The observations that a single dpc has a high probability of producing a mutation (mutagenic efficiency 1.0–3.0) would suggest, however, that this repair is extremely error-prone. If the breaks need not be repaired to permit survival, then lethal lesions are a subset of or are completely different from mutagenic lesions. 相似文献
9.
Of production by homogenates and isolated membranes of E. coli has been examined. Approximately one-fourth of the O 2-generated by extracts in the prescence of NAD (P) H is attributable to the membranes. The autoxidizable membrane component is a member of the respiratory chain, since O 2-production is NADH-specific, amplified by cyanide, and absent from membranes lacking the respiratory NADH dehyd-rogenase. Other respiratory substrates (succinate, I -phosphoglycerol, D-lactate. and L-lactate) supported Or production at efficiencies between 3 and 30 O 2-released per 10.000 electrons transferred, under conditions of substrate saturation.
Membranes from quinoneless mutants quantitatively retain the ability to evolve O 2-. indicating that the dehydrogenases are the sites of O 2-production. Relative O 2-production was greater at low substrate concentrations, probably reflecting the facilitation of unpairing of electrons that may occur when enzymes with multiple redox centers are only partially reduced.
Respiration rate, cell volume, rates of membraneous and cytosolic O 2-production, and SOD levels were used to calculate a steady-state concentration of O 2-between 10 -- 10 and 10 -- 9 M in well-fed, aerobic, SOD-proficient cells. 相似文献
10.
研究了常规CO 2(350μl·L -1)和CO 2倍增(700μl·L -1)环境中生长的春小麦幼苗,渗透胁迫时叶片中活性氧含量的变化和质膜透性的变化.结果表明,生长在CO 2倍增环境中的春小麦幼苗,渗透胁迫时O 2·-及H 2O 2的增长幅度均小于常规CO 2浓度下生长的春小麦幼苗.质膜透性的增长幅度也是前者小于后者.据此认为,CO 2浓度倍增可以减轻渗透胁迫对质膜的氧化伤害,提高植物的抗旱力. 相似文献
11.
Thyroglobulin (Tg) was subjected to metal-catalyzed oxidation, and the oxidative degradation was analyzed by SDS-polyacrylamide gel electrophoresis under reducing conditions. In contrast to no effect of hydrogen peroxide (H 2O 2) alone on the Tg degradation, the inclusion of Cu 2+ (30 μM), in combination with 2 mM H 2O 2, caused a remarkable degradation of Tg, time- and concentration-dependent. The action of Cu 2+ was not mimicked by Fe 2+, suggesting that Tg may interact selectively with Cu 2+. A similar degradation of Tg was also observed with Cu 2+corbate system, and the concentration of Cu 2+ (5-10 μM), in combination with ascorbate, required for the effective degradation was smaller than that of Cu 2+ (10-30 μM) in combination with H 2O 2. In support of involvement of H 2O 2 in the Cu 2+ corbate action, catalase expressed a complete protection. However, hydroxyl radical scavengers such as dimethylsulfoxide or mannitol failed to prevent the oxidation of Tg whereas phenolic compounds, which can interact with Cu 2+, diminished the oxidative degradation, presumably consistent with the mechanism for Cu 2+-catalyzed oxidation of protein. Moreover, the amount of carbonyl groups in Tg was increased as the concentration (3-100 μM) of Cu 2+ was enhanced, while the formation of acid-soluble peptides was not remarkable in the presence of Cu 2+ up to 200 μM. In further studies, Tg pretreated with heat or trichloroacetic acid seemed to be somewhat resistant to Cu 2+-catalyzed oxidation, implying a possible involvement of protein conformation in the susceptibility to the oxidation. Based on these observations, it is proposed that Tg could be degraded non-enzymatically by Cu 2+-catalyzed oxidation. 相似文献
12.
In the present study, using the technique of EPR spin trapping with DMPO a spin trap, we demonstrated formation of thiyl radicals from thiol-containing angiotensin converting enzyme (ACE) inhibitor captopril (CAP) and from its stereoisomer epicaptopril (EPICAP), a non-ACE inhibitor, in the process of .OH radical scavenging. Splitting constants of DMPO/thiyl radical adducts were identical for both thiols and were a N = 15.3 G, and a H = 16.2 G. Bimolecular rate constants for the reaction of CAP and EPICAP with .OH radicals were close to a diffusion-controlled rate (≈ 2 × 10 10 M −1s −1). Our data also show that both CAP and EPICAP reduce Fe(III) ions and that their respective thiyl radicals are formed in this reaction. In the presence of Fe(III), H 2O 2, and CAP, or EPICAP, .OH radicals were produced by a thiol-driven Fenton mechanism. Copper(II) ions were also reduced by these thiols, but no thiyl radicals could be detected in these reactions, and no .OH or other Fenton oxidants were observed in the presence of H 2O 2. Our data show direct evidence that thiol groups of CAP and EPICAP are involved in scavenging of .OH radicals. The direct .OH radical scavenging, together with the reductive “repair” of other sites of .OH radical attack, may contribute to the known protective effect of CAP against ischemia/reperfusion-induced arrhythmias. The formation of reactive thiyl radicals in the reactions of the studied compounds with .OH radicals and with Fe(III) ions may play a role in some of the known adverse effects of CAP. 相似文献
13.
The effect of different oxygen radical-generating systems on NAD(P)H was determined by incubating the reduced forms of the pyridine coenzymes with either Fe 2+-H 2O 2 or Fe 3+-ascorbate and by analyzing the reaction mixtures using a HPLC separation of adenine nucleotide derivatives. The effect of the azo-initiator 2,2'-azobis(2-methylpropionamidine)dihydrochloride was also tested. Results showed that, whilst all the three free radical-producing systems induced, with different extent, the oxidation of NAD(P)H to NAD(P) +, only Fe 2+-H 2O 2 also caused the formation of equimolar amounts of ADP-ribose(P) and nicotinamide. Dose-dependent experiments, with increasing Fe 2+ iron (concentration range 3-180 μM) or H 2O 2 (concentration range 50-1000 μM), were carried out at pH 6.5 in 50 mM ammonium acetate. NAD(P) +, ADP-ribose(P) and nicotinamide formation increased by increasing the amount of hydroxyl radicals produced in the medium. Under such incubation conditions NAD(P) +/ADP-ribose(P) ratio was about 4 at any Fe 2+ or H 2O 2 concentration. By varying pH to 2.0, 3.0, 4.0, 4.5, 5.0, 5.5, 6.0, 7.0 and 7.4, NAD(P) +/ADP-ribose(P) ratio changed to 5.5, 3.2, 1.8, 1.6, 2.0, 2.5, 3.0, 5.4 and 6.5, respectively. Kinetic experiments indicated that 90-95% of all compounds were generated within 5s from the beginning of the Fenton reaction. Inhibition of ADP-ribose(P), nicotinamide and NAD(P) + production of Fe 2+-H 2O 2-treated NAD(P)H samples, was achieved by adding mannitol (10-50 mM) to the reaction mixture. Differently, selective and total inhibition of ADP-ribose(P) and nicotinamide formation was obtained by performing the Fenton reaction in an almost completely anhydrous medium, i.e. in HPLC-grade methanol. Experiments carried out in isolated postischemic rat hearts perfused with 50 mM mannitol, showed that, with respect to values of control hearts, this hydroxyl radical scavenger prevented reperfusion-associated pyridine coenzyme depletion and ADP-ribose formation. On the basis of these results, a possible mechanism of action of ADP-ribose(P) and nicotinamide generation through the interaction between NAD(P)H and hydroxyl radical (which does not involve the C-center where “conventional” oxidation occurs) is presented. The implication of this phenomenon in the pyridine coenzyme depletion observed in postischemic tissues is also discussed. 相似文献
14.
The oxalate catalyzed iron(III) transfer from a trihydroxamate siderophore ferrioxamine B, [Fe(Hdfb) +], to ethylenediaminetetraacetic acid (H 4edta) has been studied spectro-photometrically in weakly acidic aqueous solutions at 298 K and a constant 2.0 M ionic strength maintained by NaClO 4. The results reveal that oxalate is a more efficient catalyst than the so far studied synthetic monohydroxamic acids. Any role of reduction of Fe(Hdfb) + by oxalate in the catalysis has been rejected by the experimentally observed preservation of the oxalate concentration during the reaction time. Therefore, catalysis has been proposed to be a substitution based process. Under our experimental conditions Fe(Hdfb) + is hexacoordinated and addition of oxalate results in the formation of Fe(H 2dfb)(C 2O 4), Fe(H 3dfb)(C 2O 4) −2 and Fe(C 2O 4) 3−3. Therefore, catalysis was proposed to be accomplished by the intermediate formation of the ternary and tris(oxalato) complexes. All three complexes react with H 2edta 2− to form thermodynamically stable Fe(edta) − as a final reaction product. Whereas the formation of the ternary complexes is fast enough to feature a pre-equilibrium process to the iron exchange reaction, the formation of Fe(C 2O 4) 3−3 is slow and is directly involved in the rate determining step of the Fe(edta) − formation. Nonlinear dependencies of the rate constant on the oxalate and the proton concentrations have been observed and a four parallel path mechanism is proposed for the exchange reaction. The rate and equilibrium constants for the various reaction paths were determined from the kinetic and equilibrium study involving the desferrioxamine B- (H 4dfb +), oxalate- and proton-concentration variations. The observed proton catalysis was attributed to the fast monoprotonation of ferrioxamine B as well as of the oxalate ligand. The observed catalysis of iron dissociation from the siderophore has been discussed in view of its significance with respect to in vivo microbial iron transport. 相似文献
15.
Oxygen radical generating systems, namely, Cu(II)/ H 2O 2, Cu(II)/ascorbate, Cu(II)/NAD(P)H, Cu(II)/ H 2O 2/catecholamine and Cu(II)/H 2O 2/SH-compounds irreversibly inhibited yeast glutathione reductase (GR) but Cu(II)/H 2O 2 enhanced the enzyme diaphorase activity. The time course of GR inactivation by Cu(II)/H 2O 2 depended on Cu(II) and H 2O 2 concentrations and was relatively slow, as compared with the effect of Cu(II)/ascorbate. The fluorescence of the enzyme Tyr and Trp residues was modified as a result of oxidative damage. Copper chelators, catalase, bovine serum albumin and HO ˙ scavengers prevented GR inactivation by Cu(II)/H 2O 2 and related systems. Cysteine, N-acetylcysteine, N-(2-dimercaptopropi-onylglycine and penicillamine enhanced the effect of Cu(II)/H 2O 2 in a concentration- and time-dependent manner. GSH, Captopril, dihydrolipoic acid and dithiotreitol also enhanced the Cu(II)/H 2O 2 effect, their actions involving the simultaneous operation of pro-oxidant and antioxidant reactions. GSSG and try-panothione disulfide effectively protected GR against Cu(II)/H 2O 2 inactivation. Thiol compounds prevented GR inactivation by the radical cation ABTS* +. GR inactivation by the systems assayed correlated with their capability for HO* radical generation. The role of amino acid residues at GR active site as targets for oxygen radicals is discussed. 相似文献
16.
[Ru II(Me 2edda)(H 2O) 2] (1), Me 2edda 2− = N, N′-dimethylethylenediaminediacetate, exhibits a sterically-controlled molecular recognition in forming η 2 and η 4 olefin complexes. 1 exists with an N 2O 2 in-plane set of chelate donors and axial H 2O ligands. The two CH 3 functionalities of Me 2edda 2− are poised above and below the N 2O 2 plane of the glycinato rings. Studies herein of the 2,2′-bipyridine complex, [Ru II(Me 2edda)(bpy)], with bidentate bpy chelation as established via 1H NMR and electrochemical methods show 1 to be ligated in the S,S configuration with the glycinato rings in-plane as a cis-O form. 1 is sterically discriminating in forming η 2 complexes with smaller olefins (ethylene, 2-propene, cis-2-butene, methyl vinyl ketone and 3-cyclohexene-1-methanol), but rejects larger decorated ring structures and branched olefins (1,2-dimethyluracil, cyclohexene-1-one 2-methyl-2-propene). η 2 complexes of 1 have characteristic Ru II/III DPP waves near 0.55 V which vary slightly with olefin structure. Potentially bidendate dienes (1,3-butadiene, 1,3-cyclohexadiene and 2,5-norbornadiene (nbd) form η 4 complexes as shown by Ru II/III waves between 0.94 and 1.30 V, indicate of a highly stabilized Ru II center by π-backboning. An η 2η 4 ‘equilibrium’ with apparent K = 22 at 25 °C is observed for nbd coordinated to 1. (The η 2 and η 4 distribution may be a kinetic one and not a thermodynamic one). To allow formation of the cis η 4 complexes, 1 must undergo a shift of one or both glycinato donors from the N 2O 2 plane into the axial site away from the dimethyl functionalities. η 4 chelation by 1,3-butadiene has been confirmed by 1H NMR spectral assignments of two [Ru II(Me 2edda)] isomers, one in the axial rans-O glycinato configuration, e.g. 1,3-butadiene is bidentate in the original N 2O 2 plane and a second unsymmetrical glycinato arrangement with in-plane and axial glycinato as well as in-plane and axial η 4-1,3-butadiene coordination. [Ru II(hedta)(H 2O)] − (2), hedta 3− = N-hydrpxyethylenediaminetriacetate, is less discriminating for olefin structures, forming η 2 complexes with all eleven olefins and dienes mentioned for studies with 1. However, 2 does not undergo displacement of a carboxylate donor by the second olefin unit of a diene [Ru II(hedta)(diene)] − complexes possess a pendant non-coordinated olefin and on η 2-bound olefin in the complex, indicated by a normal Ru II(pac)(olefin)Ru II/III wave near 0.55 V. 相似文献
17.
We examined the electrophysiological effect of pituitary adenylate cyclase activating polypeptide (PACAP) in isolated Xenopus laevis oocytes in vitro. In conventional two-electrode voltage clamp experiments, PACAP (1–10 μM) activated an inward rectifier current at membrane potentials more negative than −60 mV without causing any significant change in currents at potentials more positive than −60 mV both in the follicle-enclosed oocyte and in the defolliculated oocyte. This current reversed at −22.5 mV, close to the theoretical value of Cl − equilibrium potential and the reversal potential of this current was shifted positively by reducing [Cl −] o. This current was blocked by Cl − channel blocker SITS and Ba 2+. Furthermore, VIP and adenylate cyclase activator forskolin did not elicit the currents. In conclusion, PACAP elicited the hyperpolarization-activated Cl − current in Xenopus laevis oocytes. This current may modulate the membrane potential of the oocyte, thereby affecting the oocyte physiology. 相似文献
18.
Heme catalases are considered to degrade two molecules of H 2O 2 to two molecules of H 2O and one molecule of O 2 employing the catalatic cycle. We here studied the catalytic behaviour of bovine liver catalase at low fluxes of H 2O 2 (relative to catalase concentration), adjusted by H 2O 2-generating systems. At a ratio of a H 2O 2 flux (given in μM/min - 1) to catalase concentration (given in μM) of 10 min - 1 and above, H 2O 2 degradation occurred via the catalatic cycle. At lower ratios, however, H 2O 2 degradation proceeded with increasingly diminished production of O 2. At a ratio of 1 min - 1, O 2 formation could no longer be observed, although the enzyme still degraded H 2O 2. These results strongly suggest that at low physiological H 2O 2 fluxes H 2O 2 is preferentially metabolised reductively to H 2O, without release of O 2. The pathways involved in the reductive metabolism of H 2O 2 are presumably those previously reported as inactivation and reactivation pathways. They start from compound I and are operative at low and high H 2O 2 fluxes but kinetically outcompete the reaction of compound I with H 2O 2 at low H 2O 2 production rates. In the absence of NADPH, the reducing equivalents for the reductive metabolism of H 2O 2 are most likely provided by the protein moiety of the enzyme. In the presence of NADPH, they are at least in part provided by the coenzyme. 相似文献
19.
The ability of oxidative stress to induce apoptosis (programmed cell death), and the effect of Trolox, a water soluble vitamin E analog, on this induction were studied in vitro in mouse thymocytes. Cells were exposed to oxidative stress by treating them with 0.5–10 μM hydrogen peroxide (H 2O 2) for 10 min, in phosphate-buffered saline supplemented with 0.1 mM ferrous sulfate. Cells were resuspended in RPMI 1640 medium with 10% serum and incubated at 37°C under 5% CO 2 in air. Electron microscopic studies revealed morphological changes characteritic of apoptosis in H 2O 2-treated fragmented the DNA in a manner typical of apoptotic cells, producing a ladder pattern of 200 base pair increments upon agarose gel electrophoresis. The percentage of DNA fragmentation (determined fluorometrically) increased with increasing doses of H 2O 2 and postexposure incubation times. Pre- or posttreatment of cells with Trolox reduced H 2O 2-induced DNA fragmentation to control levels and below. The results indicate that oxidative stress induces apoptosis in thymocytes, and this induction can be prevented by Trolox, a powerful inhibitor of membrane damage. 相似文献
20.
Two new multi-cobalt-containing polyoxotungstates K 4Na 6Co 2(H 2O) 12{Co(H 2O) 4[Co 2(H 2O) 10Co 4(H 2O) 2( B--SiW 9O 34) 2] 2} · 40H 2O (1) and K 10Na 2[Co 4(H 2O) 2(GeW 9O 34) 2] · 20H 2O (2) have been obtained by the routine synthetic reactions in aqueous solution. The polyoxoanion framework of 1 consists of two sandwich-type polyoxoanions [Co 4(H 2O) 2( B--SiW 9O 34) 2] 12− connected together by a [CoO 2(H 2O) 4] cluster to constitute the sandwich dimer, and then, four isolated Co(H 2O) 5 cations coordinate to the dimer through four μ2-O atoms. The polyoxoanion 2 is isomorphic to the sandwich-type polyoxoanion [Co 4(H 2O) 2( B--SiW 9O 34) 2] 12− in 1. The magnetic property of compound 1 has been studied by measuring its magnetic susceptibility in the temperature range 2.0–300.0 K, indicating the existence of intramolecular ferromagnetic Co–Co interactions, and, the electrochemical properties of 1 and 2 are detected in the pH 4 buffer solution. 相似文献
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