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Biomedical subjects

M Bazin

Publications and source records attributed to M Bazin.

At least 19 recordsLinked to original sources
PubMed1995

Subcellular localization of and photosensitization by protoporphyrin IXhuman keratinocytes and fibroblasts cultivated with 5-aminolevulinic acid.

The subcellular localization of protoporphyrin (PP) has been studied by microspectrofluorometric techniques in NCTC 2544 keratinocytes incubated with 5-aminolevulinic acid (ALA) for times up to 42 h. Whereas the plasma membrane shows strong staining, fluorescent spots are observed within the cytoplasm especially in the perinuclear region. Although the topographic pattern of the PP distribution does not change much with the incubation time with ALA, the fluorescence spectra suggest that the PP microenvironments are quite different at short and long incubation times. Addition of 18 microM desferrioxamine almost doubles the ALA-induced PP concentration. Colocalization experiments with rhodamine 123, a mitochondrial probe, and lucifer yellow (LY) or neutral red (NR), two lysosome probes, demonstrate that at least some of these spots are of lysosomal origin. Study of the time evolution of the NR fluorescence under irradiation with visible light in the presence and absence of ALA demonstrates that lysosomes are damaged cells that have synthesized PP. No PP fluorescence can be detected in mitochondria after incubation with ALA. However, photosensitization of mitochondria occurs under irradiation with visible light. Very little formation of lipofuscins by photosensitization with exogenous PP or ALA-induced PP is observed with the NCTC 2544 keratinocytes, as compared to normal human fibroblasts.

Aminolevulinic Acid↗
PubMed1994

Glutathione peroxidase and glutathione reductase activities towards glutathione-derived antioxidants.

A new class of glutathione derivatives with antioxidant properties has been prepared by transformation of the NH2 group into a pyrrole ring with various substitutions at the 2 and 5 positions. Due to steric hindrance and/or hydrophobicity of the 2-5-disubstituted pyrrole ring, the reduced glutathione derivatives are poor substrates of the glutathione peroxidase and do not effectively compete with GSH. The oxidized glutathione derivatives are, in turn, relatively good substrates (Km = 1.5 mM) of the glutathione reductase as compared to natural oxidized glutathione (Km = 0.51 mM) but are not effective competitors of the enzyme. It can be considered that the new glutathione derivatives do not strongly interfere with the natural enzymatic defence against fatty acid hydroperoxides formed during an oxidative stress.

Animals↗
PubMed1994

A pulse radiolysis study of one-electron oxidation and .OH radical reaction of a new class of antioxidants: 2,5 dimethylpyrrole--amino acid N-conjugates.

A new class of 2,5 dimethylpyrrole functionalized amino acid and peptide antioxidants has been synthesized. Pulse radiolysis studies have been carried out on these compounds to determine their reactivity in one-electron oxidation reactions with Br2- and (SCN)2-, as well as their reactivity toward the water radicals .OH and eaq-. When compared to similar reactions with parent amino acids and peptides, both rate constants and transient spectra indicate considerable interaction between the electronic states of the pyrrole moiety and amino acid or peptide. Where comparison data is available, radical reactions are significantly more rapid with these derivatives than with the parent compounds. Exceptions occur in some cases of .OH attack where steric hindrance may be evoked to explain a lower rate constant in the derivative compounds. Electron attachment rates are also found to be enhanced by the presence of the pyrrole ring.

Amino Acids↗
PubMed1991

Type I and type II photosensitization by the antibacterial drug nalidixic acid. A laser flash photolysis study.

The 355 nm laser flash photolysis of nalidixic acid at pH 9.2 leads to the formation of the nalidixate anion triplet state (absorption lambda max = 620 nm; 5700 less than or equal to epsilon T less than or equal to 9000 M-1cm-1; 0.6 less than or equal to phi T less than or equal to 1). The first order triplet state decay (kT = 7.7 x 10(3) s-1) is accompanied by a diffusion controlled triplet-triplet annihilation. Oxygen efficiently quenches the triplet state (k = 3.2 x 10(9) M-1s-1). The nalidixate radical dianion (absorption lambda max = 650 nm; epsilon = 3000 M-1cm-1) is produced by the diffusion controlled reductive quenching of the triplet state by tryptophan and tyrosine. The superoxide anion (O2-.) is produced by diffusion controlled reaction of the radical dianion with oxygen. The O2-. is characterized by its reactions with ferricytochrome c and superoxide dismutase. The physiological form of nalidixic acid is thus a good Type I and Type II photosensitizer.

Anions↗
PubMed1991

Photoreactivity of 5-geranoxypsoralen and lack of photoreaction with DNA.

5-Geranoxypsoralen, commonly called bergamottin, a major furocoumarin contained in bergamot oil, is reported in vitro as a highly photoreactive psoralen. In ethanol, it exhibits quite a high triplet state quantum yield (approximately 0.37). The triplet state is involved in subsequent photochemistry which depends on the initial concentration and on the presence of oxygen. In contrast to most psoralens, absorption and fluorescence data suggest that 5-geranoxypsoralen does not interact with DNA in the dark. No UVA-induced interstrand cross-links in DNA were shown.

DNA↗
PubMed1990

Degradation of crystallins from a psoriatic patient undergoing PUVA therapy.

Comparative physico-chemical and spectroscopic analyses were carried out in human lens proteins obtained from extracts of normal, senile and PUVA cataracts. Mass recovery analysis reveals a large protein concentration loss in the PUVA cataract relative to the normal lens and senile cataract. This protein loss parallels an increase in the degraded polypeptide chains. However, the tryptophan content (2.1 mol/mol of 20 kDa protein subunit) and the apparent fluorescence quantum yield (phi f = 0.056) of the tryptophan residues which are believed to be involved in the development of UV-induced cataracts are unchanged after age-related alterations and/or in vivo photochemistry associated with psoralen (8MOP) photosensitized reactions.

Aging↗
PubMed1990

Ultra-structure and localisation of formazan formed by human neutrophils and amoebae phagocytosing virulent and avirulent Legionella pneumophila.

Legionella pneumophila (LP) strains of differing virulence were incubated with a solution of nitroblue-tetrazolium (NBT) at a concentration of 1 mg.ml-1 in the presence of Acanthamoeba polyphaga or human polymorphonuclear neutrophils (PMN). Reduction of NBT to formazan occurred at a faster rate in the presence of virulent strains. Reduction appeared to be temperature dependent; at 37 degrees C the reaction rate was higher than at 20 degrees C. On microscopic examination, deposits of formazan around Legionella cells were observed inside amoebae similar to those deposited in human neutrophils. Electron microscopy revealed electron-dense particles surrounding virulent legionellae, which appeared to be associated with formazan formation. Formazan formation inside amoebae may suggest the presence of a respiratory burst against LP, which is more intense with virulent strains.

Acanthamoeba↗
PubMed1990

Is rhodamine 123 a photosensitizer?

Because of conflicting reports on the photoxicity of rhodamine 123 (Rh 123), we have undertaken a study of Rh 123 photosensitization in several in vitro systems. First, Rh 123 is not a photodynamic agent and does not react with singlet oxygen. Second, when bound to cytochrome c (Cyt c), Rh 123 photosensitizes ferro Cyt c but not ferri Cyt c degradation by an oxygen-independent process. When delivered to skin fibroblasts where it specifically stains mitochondria, Rh 123 photosensitizes membrane damage. These results are consistent with the hypothesis that Rh 123 is a phototoxic stain. The lack of photosensitivity of Rh 123-stained mitochondria in some cell lines might therefore be due to specific structural features.

Animals↗
PubMed1989

Photosensitization with alpha-terthienyl: the formation of superoxide ion in aqueous media.

It is well known that alpha-terthienyl generates singlet oxygen in organic solvents with high quantum yields. In an aqueous medium, the production of superoxide radical-anion is readily detected by comparing the reduction of ferricytochrome c or nitro blue tetrazolium in the presence and in the absence of superoxide dismutase. Electron transfer reactions from the electronically excited sensitizer are also detected in an argon atmosphere.

Cytochrome c Group↗
PubMed1989

[Neutralizing activity against herpes simplex in maternal milk].

Human milk could play a role in the protection of the newborn from Herpes simplex virus 2 contamination. Neutralizing activity against Herpes simplex virus 2 was measured in 94 human milk samples and in 31 sera from healthy women. A weak neutralizing activity was detected in 48 milk samples. They were neither correlated to anti-Herpes simplex virus antibodies found in milk, nor to the neutralizing activity in sera. The origin and non immunologic nature of this activity are discussed.

Antibodies, Viral↗
PubMed1988

Determination of the dioxygen quenching constant for protein and model indole triplets.

The decay of the indole triplet of single tryptophan-containing proteins and model compounds can be readily determined at room temperature in solution by monitoring the triplet absorption or emission following an exciting laser pulse. The dioxygen triplet quenching constants, can be measured for all these molecules and compared to the analogous singlet values determined by fluorescence methods. The dioxygen triplet quenching constant (tkq) ranged from a high of 5.1.10(9) M-1.s-1 for the exposed indole of corticotropin to a low of 0.1.10(9) M-1.s-1 for the buried indole of asparaginase. The ratio of these values with their respective dioxygen singlet quenching constants (skq), tkq/skq, ranged from 0.3 to 0.6 for aqueous exposed polypeptide indoles. For globular proteins the tkq/skq value is observed to be 0.2 +/- 0.1. This lower value for protein indoles is not attributable to 'bulk' environmental or hydrogen bonding effects, since the magnitude of tkq/skq (= 0.5 +/- 0.1) for model indoles was independent of solvent dielectric constant, polarity, and proticity. Temperature-dependence studies were done to test whether tkq could be used to characterize the nature of the protein matrix. The activation energy (Ea) for tkq was found to be 11 +/- 2 kcal/mol for most proteins. This Ea was independent of whether the indole side-chain was solvent exposed or buried in the non-aqueous protein interior. Large Ea values were also obtained for model indoles, naphthalene and nalidixic acid, dissolved in water, whereas the same compounds dissolved in 95% ethanol exhibited much smaller Ea values. These data, in combination with the observation that the tkq of model indoles is insensitive to changes in solvent viscosity, indicate that dioxygen quenching at the triplet level can not be easily used to characterize the dynamics of proteins.

Indoles↗
PubMed1988

Characterization of the indole triplet excited state in proteins utilizing laser flash photolysis.

The triplet-triplet absorption spectrum of the sole indole side chain of human serum albumin and its decay kinetics were previously characterized, at room temperature, by using a conventional flash photolysis method [(1978) Proc. Natl. Acad. Sci. USA 75, 1172-1175]. Exploitation of this potentially useful long lived reporter group in protein studies was limited by the excessively large sample size required by that apparatus. The 265 nm laser flash instrument used in the present work avoids this problem at the price of a loss in photo-selectivity. We report that the latter concern can be mitigated. Melittin was studied first because this polypeptide contains a single aromatic residue (W-19), and because its monomeric and tetrameric forms are good models for solvent exposed and buried indole side chains of proteins. For both forms, the indole triplet and neutral radical absorption spectra could be readily time resolved and identified on the basis of shape and differential dioxygen sensitivity. The single tryptophan containing protein human serum albumin was studied next because it contains a large number of other 265 nm absorbing moieties whose transient spectra might complicate the detection of the indole triplet. These transients were shown to not interfere significantly in the wavelength region 450 nm to 600 nm, and, in contrast to the indole triplet, they were relatively dioxygen insensitive. Thus, a facile means is available by which the indole triplet of proteins may be characterized. Subsequently the question of whether this species could be detected in the presence of nuclei acid components was investigated by flashing the phage fd. The putative nucleic acid transients were shown not to interfere and the absorbance of the indole triplet was readily time resolved. The spectral assignment was persuasively confirmed by showing that the indole triplet absorption and phosphorescence emission spectra decay with the same lifetime. The present work thus provides additional evidence for the general applicability of the indole triplet excited state as a long lived intrinsic protein reporter group.

Bacteriophages↗
PubMed1987

Phototoxic effects of fluoranthene, a polycyclic aromatic hydrocarbon, on bacterial species.

Fluoranthene, a non-carcinogenic polycyclic aromatic hydrocarbon, inactivates Escherichia coli cells in the presence of near-ultraviolet light (NUV; 300-400 nm). E coli cells carrying defects in the uvrA6 or katF genes are sensitized to inactivation by the simultaneous treatment with fluoranthene and NUV, suggesting that DNA is a target and that hydrogen peroxide is generated. Haemophilus influenzae transforming DNA can be inactivated by the simultaneous treatment with fluoranthene and NUV confirming DNA as a target. Using the photooxidation of imidazole and histidine as probes, fluoranthene was found to generate singlet oxygen in organic and aqueous media. In water, it participated in electron transfer reactions, reducing nitro blue tetrazolium as well as ferricytochrome C. This reduction took place both in the presence of air, where superoxide anion was formed, and under argon. Simultaneous treatment with fluoranthene and NUV was incapable of inducing histidine-independent mutations. Simultaneous treatment with fluoranthene and NUV was incapable of inducing the uvrA gene product as evidenced by the absence of the induction of beta-galactosidase in an E coli operon fusion strain [uvrA215::Mud(Ap,lac)].

DNA Damage↗