摘要:

In neutral solutions polyamines are fully protonated, and hence are really polyammonium cations (PAC). Spermine, for example, carries four positive charges in a linear system, H3N+(CH2)3N+H2(CH2)4N+H2(CH2)3N+H3. There is a very powerful coulombic interaction between aqueous DNA and such cations, thus the cations are attracted to the DNA over large distances, and once close to the DNA normally remain there for long periods. A key issue is; are the cations mobile, or do they remain at one preferred site for significant periods? The latter is the currently preferred concept, but NMR and EPR evidence will be presented in favour of the former. If the former is correct, then PACs may be able to act as good drug delivery systems. In its simple form the concept is that any drug that acts directly on DNA can be chemically bound to a PAC. Once in the cell, this PAC-drug complex (PAC-D) will be carried to DNA and will move very rapidly along the exposed strands until it recognises the site of action. This may be some special base sequence region, a damaged site, or the PAC-D unit may simply be present prior to potential damage, so that this can be repaired very rapidly. Some of our current studies on these systems are described.

ISSN:1071-5762    

DOI:10.3109/10715769509147523

出版商:Taylor&Francis    

年代:1995

数据来源: Taylor

摘要:

The 5, 5-dimethyl-1-pyrroline-N-oxide (DMPO) spin adduct of myoglobin (Mb) or hemoglobin (Hb) was formed when metmyoglobin (MetMb) or methemoglobin (MetHb) reacted with H2O2in the presence of DMPO, and both decayed with half-life of a few minutes. The DMPO spin adduct of Mb decayed with biphasic kinetics with k1= 0.645 min−1and k2= 0.012 min−1, indicating that the spin adduct consisted of two kinetically heterogeneous species, stable and unstable ones. The DPMO spin adduct of Hb, however, was homogeneous. Decay of both spin adducts was accelerated in the presence of tyrosine, tryptophan or cysteine, but not phenylalanine, methionine or histidine. The decay obeyed the first order kinetics at varying concentrations of the spin adducts. The decay was accelerated by denaturation and proteolysis of protein moiety. The decay rate was not affected by the extra addition of MetMb or MetHb to each spin adduct. The decay rate of the spin adduct of Mb was increased by hematin in the presence of H2O2and decreased by catalase. Decay of stable spin adduct of Mb, however, was not significantly changed under any experimental conditions used. These results led us to conclude that instability of the DMPO-spin adducts of Mb and Hb is due to intramolecular redox reactions between the spin adducts and amino acid residues and/or products of the reaction between heme and H2O2.

ISSN:1071-5762    

DOI:10.3109/10715769509147524

出版商:Taylor&Francis    

年代:1995

数据来源: Taylor

摘要:

Free radical formation and subsequent lipid peroxidation may participate in the pathogenesis of tissue injury, including the brain injury induced by hypoxia or trauma and cardiac injury arising from ischemia and reperfusion. However, the exact cellular mechanisms by which the initial oxidative insult leads to the ultimate tissue damage are not known. A number of reports have indicated that protein kinase C (PKC) may be activated following oxidative stress and that this enzyme may play an important role in the steps leading to cellular damage. In this work, we have examined in a cell model whether PKC is activated following oxidative exposure. UC11MG cells, a human astrocytoma cell line, were treated with H2O2. Incubation with 0.5 mM H2O2increased malondialdehyde levels by as early as 15 minutes. To assess the effects of H2O2treatment on PKC activation, we measured phosphorylation of an endogenous PKC substrate, the MARCKS (myristoylatedalanine-richCkinasesubstrate) protein. Treatment of cells with 0.2-1.0 mM H2O2resulted in a rapid increase in MARCKS phosphorylation. Phosphorylation was stimulated approximately 2.5-fold following treatment with 0.5 mM H2O2for ten minutes. Treatment with phorbol 12-myristate 13-acetate, a PKC activator, increased MARCKS phosphorylation approximately 4-fold. The H2O2-induced MARCKS phosphorylation was inhibited by the addition of the kinase inhibitors H-7 and staurosporine. Furthermore, specific down-regulation of PKC by phorbol ester also inhibited H2O2-induced MARCKS phosphorylation. These results indicate that PKC is rapidly activated in cells following an oxidative exposure and that this cell system may be a good model to further investigate the role of PKC in regulating oxidative damage in the cell.

ISSN:1071-5762    

DOI:10.3109/10715769509147525

出版商:Taylor&Francis    

年代:1995

数据来源: Taylor

摘要:

To delineate the role of peroxisomes in the pathophysiology of hypoxia-reoxygenation we examined the functions of peroxisomes and mitochondria in cultured skin fibroblasts from controls and from patients with cells lacking peroxisomes (Zellweger cells). The loss of peroxisomal functions (lignoceric acid oxidation and dihydroxyacetonephosphate acyltransferase [DHAP-AT] activities) in control cells following hypoxia and hypoxia followed by reoxygenation, suggests that peroxisomes are sensitive to oxidative injury. The sensitivity of peroxisomes to oxidative stress was compared to that of mitochondria by examining the oxidation of palmitic acid (a function of both mitochondria and peroxisomes) in control and Zellweger cell lines, following hypoxia-reoxygenation. The greater loss of activity of palmitic acid oxidation observed in control cells as compared to that seen in Zellweger cells suggests that the peroxisomalβ-oxidation system is relatively more labile to hypoxia- reoxygenation induced oxidative stress. This data clearly demonstrates the difference in the response of mitochondria and peroxisomes to oxidative stress.

ISSN:1071-5762    

DOI:10.3109/10715769509147526

出版商:Taylor&Francis    

年代:1995

数据来源: Taylor

摘要:

Nitric oxide reacts with nitronyl nitroxides (NNO) to form imino nitroxides (INO) and this transformation can be monitored using electron spin resonance spectroscopy. Recently, Akaikeet al., reported that NNO such as 2-phenyl-4,4,5,5-tetramethylimidazoline-3-oxide-1-oxyl (PTIO) and its derivatives (e.g., carboxy-PTIO) react with nitric oxide (·NO) in a 1:1 stoichiometry forming 2-phenyl-4,4,5,5-tetra-methylimidazoline-1-oxyl (PTI) or the respective product (e.g., carboxy-PTI) together with nitrite and nitrate (Akaikeet al., Biochemistry 32, 827-832, 1993). In this paper, we reevaluate their results and show that the stoichiometry of the reaction between PTIO and·NO is 0.63±0.06:1.0. The reason for this discrepancy is due to an erroneous assumption by Akaikeet al., that the stoichiometry for the reaction between·NO and O2is 2:1 in aqueous solution. If the data reported by Akaikeet al., were recalculated using a 4:1 stoichiometry established for the aqueous oxidation of·NO, the reaction between·NO and PTIO would give a stoichiometry of 0.5:1.0 in closer agreement with our data. We propose a mechanism for the reaction between PTIO and·NO in aqueous solution. This mechanism predicts that the stoichiometry between carboxy-PTIO and·NO is dependent on the rate of generation of·NO and is 1:1 only at low rates of·NO generation (i.e., 10−13M/s). However the stoichiometry approaches 0.5:1.0 at higher rates of·NO production or when it is added as a bolus. The ratio between nitrite and nitrate also varies as a function of the rate of generation of·NO. The model agrees with previous experimental observations that the aqueous oxidation of·NO in air saturated solutions will exclusively form nitrite and predicts that·NO will only generate substantial amounts of nitrate if it is released at a rate less than 10−17M/s. This may have important consequences in cellular systems where the concentration of·NO is typically measured from nitrite production.

ISSN:1071-5762    

DOI:10.3109/10715769509147527

出版商:Taylor&Francis    

年代:1995

数据来源: Taylor

摘要:

High concentrations of total vitamin C have been measured in the plasma of premature infants. At these concentrations ascorbic acid inhibited the ferroxidase activity of caeruloplasmin measured directlyin vitro. The degree of inhibition was dependent on the ratio of ascorbic acid: caeruloplasmin. Values for the ratio of vitamin C: caeruloplasmin measured in premature babies would be predicted to inhibit ferroxidase activity by up to at least 80%. Ferroxidase activity measured in the plasma of premature babies increased from birth but was significantly lower than in plasma collected from adults (<0.001). Plasma ferroxidase activity was correlated with plasma caeruloplasmin concentration and, in premature babies only, showed a negative correlation with the ratio of vitamin C to caeruloplasmin. High levels of vitamin C in premature babies may compromise antioxidant mechanisms and exacerbate oxidant damage.

ISSN:1071-5762    

DOI:10.3109/10715769509147528

出版商:Taylor&Francis    

年代:1995

数据来源: Taylor

摘要:

During oxidative stress, iron traces are supposed to be released from normal storage sites and to catalyse oxidative damage by Fenton-type reactions. This type of damage is difficult to preventin vivoexcept by the use of strong iron chelators such as deferoxamine (affinity constant for Fe(III): log K = 30.8). However, strong iron chelating agents are also suspected to mobilize iron from various storage and transport proteins thereby leading to toxic effects. In contrast, N,N' -bis-dibenzyl ethylenediaminediacetic acid (DBED) is an iron chelator with relatively low affinity for iron (affinity constant for Fe(III): log K<15). In the present paper, we show that, in situations mimicking oxidative stressin vitro, DBED is site-specifically oxidized into new species with strong iron binding capacity. Indeed, in the presence of ascorbate as a reductant, the iron chelate of DBED reacts with H2O2in aqueous solution to yield a purple chromophore with minor release of free HO·in the medium, as measured by aromatic hydroxylation assay. The formation of these purple species is not suppressed by the presence of HO·scavengers at high concentration. The visible spectrum of these species is consistent with a charge transfer band from a phenolate ligand to iron. N-2-hydroxybenzyl N '-benzyl ethylenediaminediacetic acid (HBBED) was identified in the medium as one of the oxidation products of DBED. Therefore, these results suggest that the iron chelate of DBED undergoes an intramolecular aromatic hydroxylation by HO·leading to 2-OH derivatives and hence that DBED is a site-specific HO- scavenger. Moreover, since the measured affinity for Fe(III) of HBBED (log K = 28) is at least 13 orders of magnitude higher than that of DBED and since ferric HBBED chelate is not a catalyst of Fenton chemistry, DBED may be looked as an“oxidative stress activatable”iron chelator, e.g. which increase in affinity for iron is triggered in the presence of H2O2and an electron donor. Therefore it is proposed that DBED and related derivatives may be interesting as protective compounds against oxygen radicals toxicity, especially for chronic use.

ISSN:1071-5762    

DOI:10.3109/10715769509147529

出版商:Taylor&Francis    

年代:1995

数据来源: Taylor

ISSN:1071-5762    

DOI:10.3109/10715769509147530

出版商:Taylor&Francis    

年代:1995

数据来源: Taylor

ISSN:1071-5762    

DOI:10.3109/10715769509147531

出版商:Taylor&Francis    

年代:1995

数据来源: Taylor