bluuebunny/crossref_metadata_2025 huggingface dataset:
MINIMUM DOSES OF ULTRAVIOLET RADIATION REQUIRED TO INDUCE MURINE SKIN EDEMA and IMMUNOSUPPRESSION ARE DIFFERENT and DEPEND ON THE ULTRAVIOLET EMISSION SPECTRUM OF THE SOURCE*
Authors: Learn Douglas B. , Beasley Donathan G. , Giddens Lori D. , Beard John , Stanfield Joseph W. , Roberts Lee K.
Published: July 2010

Many photoimmunological studies have used UV radiation sources that emit nonsolar UV spectral energy and UV doses based on nonimmunological endpoints, e.g. erythema and skin edema. Interpretation of these data has led to misunderstanding when extrapolated to hypothetical effects in humans exposed to solar UV. The purpose of this study was to: (1) establish UV dose response relationships for murine skin edema and immunosuppression, and (2) determine how different UV spectra affect these relationships. Back skin and ear minimum edema doses (MEdD) for Kodacel‐filtered FS20 sunlamp UV (290–400nm) were greater than two‐fold higher than those for unfiltered FS20 sunlamp UV (250–400nm). Xenon arc solar simulator UV (295–400nm) MEdD were > 10‐fold higher than those for unfiltered sunlamp UV. Back skin and ear MEdD differed two‐ to five‐fold between C3H/ HeN, SWR/J and HRA/Skh‐1 mice. The minimum immunosuppression doses (MISD) in C3H mice showed similar UV source spectrum dependence. The solar simulator UV MISD was 5.4‐ and 1.5‐fold higher than for unfiltered and Kodacel‐filtered sunlamp UV MISD, respectively. Furthermore, MISD were from 3‐ to 50‐fold higher than the MEdD for the three UV sources. The UV bioeffectiveness spectra indicated that UVC energy (250–290nm) contributed 12% and 18%, respectively, of the total skin edema and immunosuppression UV energy. These data demonstrate the variability in UV sensitivity among mouse strains, the significant differences between murine MEdD and MISD and how these differences are influenced by nonsolar regions (below 295 nm) of the UV spectrum.
MODIFICATION OF α‐CHYMOTRYPSIN USING A WATER‐SOLUBLE PHOTO‐FENTON REAGENT
Authors: Kitano Hiromi , Maeda Yasushi , Furukawa Kanae , Yamamoto Takahiro , Izumida Rie , Matsugo Seiichi
Published: July 2010

Abstract—‐Modification of an enzyme, α‐chymotrypsin, was examined by using a water‐soluble photo‐Fenton reagent. By photoirradiation of the enzyme with the reagent, which can occupy a binding site of the enzyme, a tryptophan residue in the vicinity of the active site was oxidized toN‐formylkynurenine. Concurrently, the catalytic properties of the enzyme were largely changed: theKmwas increased and thekcatwas decreased. The decrease inkcatfor a specific amide substrate was the most significant among the esters and amides examined. The water‐soluble photo‐Fenton reagent would be useful to chemically modify relatively limited regions in biomolecules.
SITES OF PHOTODAMAGE in vivo and in vitro BY A CATIONIC PORPHYRIN
Authors: Kessel David , Woodburn Kathryn , Henderson Barbara W. , Chang C. K.
Published: July 2010

Abstract—Localization and photodynamic efficacy of a monocationic porphyrin (MCP) were assessed using murine leukemia cells in culture. This sensitizer localized at surface membrane loci and catalyzed selective photodamage to membrane structures. Although both cationic and hydrophobic, this porphyrin was not recognized by the multidrug transporter, which excludes many cationic agents from cells that express multidrug resistance. Photodynamic studies with the murine radiation‐induced fibrosarcoma tumor model indicated moderate photosensitization of neoplastic lesionsin vivoat 3 h, but not at 24 h after sensitizer administration. Pharmacokinetic studies indicate that plasma levels, not tissue levels were the major determinant of photodynamic therapy (PDT) response. Consistent with this observation, vascular damage and disturbances of tissue perfusion followed PDT. These effects were more pronounced in tumor‐bearing skin than in normal skin. The therapeutic response to MCP appeared to be related mainly to secondary, probably vascular, effects.
THE VALIDATION OF A NEW VASCULAR DAMAGE ASSAY FOR PHOTODYNAMIC THERAPY AGENTS
Authors: Bellnier David A. , Potter William R. , Vaughan Lurine A. , Sitnik Theresa M. , Parsons John C. , Greco William R. , Whitaker James , Johnson Patricia , Henderson Barbara W.
Published: July 2010

Abstract—The therapeutic effect of photodynamic therapy (PDT: photodynamic sensitizer + light) is partly due to vascular damage. This report describes a new vascular photodamage assay for PDT agents and a validation of the assay. The method described here quantitates changes in tissue blood perfusion based on the relative amount of injected fluorescein dye in treated and untreated tissues. A specially designed fluorometer uses chopped monochromatic light from an argon laser as a source for exciting fluorescein fluorescence. The fluorescent light emitted from the tissue is collected by a six element fiberoptic array, filtered and delivered to a photodiode detector coupled to a phase‐locked amplifier for conversion to a voltage signal for recording. This arrangement permits a rather simple, inexpensive construction and allows for the simultaneous use of the argon laser by other investigators.

The routine assay for characterizing a specific photosensitizer at a standard dose consists of the sequential allocation of eight mice to a set of different light doses designed to span the dose‐response range of fluorescein fluorescence exclusion (measured 8–10 min after fluorescein injection). The assay validation experiment used an anionic photosensitizer, 2‐[l‐hexyloxyethyl]‐2‐devinyl pyropheophorbide‐a at a dose of 0.4 μmol/kg. The parameter estimates (n = 34 mice) from fitting the standard Hill dose‐response model to the data were: median fluorescence exclusion light dose FE50= 275 ± 8.3 J/cm2and Hill sigmoidicity parameter m =−3.66 ± 0.28. Subsets of the full data set randomly selected to simulate a standard eight mice experiment yielded similar parameter estimates. The new assay provides reliable estimates of PDT vascular damage with a frugal sequential experimental design.
DISTRIBUTION OF 5‐AMINOLEVULINIC ACID‐INDUCED PORPHYRINS IN NODULOULCERATIVE BASAL CELL CARCINOMA
Authors: Peng Qian , Warloe Trond , Moan Johan , Heyerdahl Helen , Steen Harald B. , Nesland Jahn M. , Giercksky Karl‐Erik
Published: July 2010

Abstract— Microscopic fluorescence photometry incorporating a light‐sensitive thermo‐electrically cooled charge‐coupled device (CCD) camera was employed to investigate the fluorescence distribution of 5‐aminolevulinic acid (ALA)‐induced porphyrins in 22 patients with a total number of 52 noduloul‐cerative basal cell carcinomas (BCC) after topical ALA application with or without dimethylsulfoxide (DMSO)/ethylenediaminetetraacetic acid (EDTA) or after intravenous administration of ALA. Both localization patterns and amounts of ALA‐induced porphyrins in the BCC were studied. The ALA‐induced porphyrins were localized only in the superficial layers of the noduloulcerative BCC lesions after topical application of 20% ALA alone for 3 h. However, both the penetration of ALA into deep lesions and the production of the ALA‐induced porphyrin fluorescence were increased after topical administration of 20% ALA and 20% DMSO/4% EDTA for 3 h. Prior treatment with 99% DMSO for 15 min further enhanced the ALA penetration into the BCC lesions after topical application of the ALA/DMSO/EDTA mixture and produced more ALA‐induced porphyrins by a factor of about three compared with those treated with ALA alone. The penetration of ALA into the deep BCC lesions could also be increased by prolonging the time of topical application of 20% ALA/4% EDTA to 29–48 h (without DMSO). Intravenous injection of ALA led to a more homogeneous distribution of the ALA‐derived porphyrins in the whole noduloulcerative BCC lesions.
cis‐UROCANIC ACID FAILED TO AFFECT in vitro HUMAN LANGERHANS CELL ALLOSTIMULATORY FUNCTION
Authors: Rattis Frédérique‐Marie , Péguet‐Navarro Josette , Courtellemont Pascal , Redziniak Gérard , Schmitt Daniel
Published: July 2010

Abstract— ‐Urocanic acid (UCA) represents the major ultraviolet B (UVB, 290–320 nm)‐absorbing component of the skin. Trans‐UCA is naturally produced in the stratum corneum and converts to the cis isomer upon UVB irradiation. In this study, we examined the effect of purified cis‐UCA (about 99% of cis isomer) on the human Langerhans cell (LC) allostimulatory function by using the mixed epidermal cell‐lymphocyte reaction (MELR). We found that addition of increasing amounts (6.5–400 μg/mL) of purified cis‐UCA or (rara‐UCA did not modify the T‐cell response supported by enriched LC (eLC: 8–25% LC) as well as purified LC (pLC: 70–90% LC) suspensions. Because cis‐UCA had no effect on the allostimulatory function of untreated LC, we investigated whether this compound could modify T‐cell proliferation induced by UVB‐irradiated LC. The UVB exposure of eLC or pLC to 100 J/m2 significantly inhibited the capacity of both suspensions to mount a T‐cell response. However, addition of cis‐UCA did not potentiate this UVB‐induced immunosuppression. The eLC or pLC were then incubated with cis‐UCA for 18 h at 37°C and washed before adding to allogeneic T cells. The obtained proliferative response was similar to that induced by control LC incubated in medium alone, demonstrating that pretreatment with cis‐UCA did not alter human LC function. In conclusion, these results strongly suggest that cis‐UCA has no direct effect on human LC antigen‐presenting function.
AN APPARATUS FOR STUDYING RAPID ELECTROPHYSIOLOGICAL RESPONSES TO LIGHT DEMONSTRATED ON Arabidopsis LEAVES
Authors: Spalding Edgar P.
Published: July 2010

Abstract— An apparatus for making high‐resolution measurements of electrophysiological changes induced by light in plant cells was constructed. Its main components were a xenon arc lamp, an electronic shutter, a liquid light‐guide, a computer equipped with an analog‐to‐digital converter and a computer program that controlled the shutter and data acquisition. The apparatus was used to examine transient changes in membrane potential (Vm) that occur upon illumination in Arabidopsis leaves. Light‐on induced a transient hyperpolarization of 4 mV after a lag time of 0.53 s. It was followed by a much larger transient depolarization that peaked 31s after light‐on. The Vm returned to near its original value after approximately 3 min. The early changes in Vm have been proposed to result from effects of photosyn‐thetically produced ATP on the activities of H+ ATPases and K+ channels at the plasma membrane. The kinetics of the initial hyperpolarization were found to be reasonably consistent with such a mechanism. It is expected that the apparatus described here will be useful in future investigations of this and other electrophysiological responses to light.
SPECTROSCOPIC STUDIES ON A RETINYLIDENE‐IMINE‐TETRAFLUOROBORATE
Authors: Baran Jan , Ilczyszyn Marek , Ratajczak Henryk , Baron Marie‐Hélène , Piart‐Goypiron Annie , Ratovelomanana Nicole , Zine Hassan , Sandorfy Camille
Published: July 2010

Abstract— N‐retinylidene‐butyl‐l‐aminium tetrafluoroborate (NRBH+BF4‐) was used to determine the frequencies of the free (non‐hydrogen‐bonded) v(N+‐H) and v(C=NH+) vibrations and their anharmon‐icity constants. For this purpose the Fourier‐transform IR and near‐IR (overtone) spectra were recorded. The visible‐UV spectra were also taken. The results locate the free v(N+‐H) band at about 3250 ± 50 cm‐1; the vibration has normal anharmonicity while v(C=NH+) is very nearly harmonic. The UV and IR spectra show that the Schiff base is protonated, but that the proton is only very weakly hydrogen‐bonded to the anion. Therefore, in this respect, NRBH+BF4‐ is a good model for the hydrogen‐bonding environment of the region around the nitrogen of rhodopsin chromophores.
Assessment of Ocular Exposure to Ultraviolet‐B for Population Studies
Authors: Duncan Donald D. , Munoz Beatriz , Bandeen‐Roche Karen , West Sheila K. ,   Salisbury Eye Evaluation Project
Published: July 2010

Abstract— We have developed an empirical model with which to estimate the ocular exposure in the UVB wavelength band. This model incorporates aspects of personal behavior, geographic location, season and wavelength that have been developed from population‐based data. This model, in conjunction with job history interviews, allows the estimate of cumulative exposures from age 30 in our population‐based study. We present data on average annual exposure by age, gender, race, education and reported photophobia. The exposures in the general population are considerably lower than exposures reported in previous works based on occupational groups. There is a statistically significant difference between males and females, with females having lower exposures. African‐Americans have median exposures similar to whites. Exposures decrease with education and with reported photophobia. These data provide the basis for characterizing lifetime exposure for the general population. The variability of exposure is much greater than found in occupational groups and includes numbers in the lowest exposure. The model should permit determination of a dose response relationship with eye disease, even to the lower dose ranges.
Nonlinear Effects and a Cascade of Radical Events Leading to Laser‐Specific Generation of Active Oxygen Species
Authors: Hancock‐Chen Tanya , Scaiano J. C.
Published: July 2010

Laser photolysis of benzoyl radical precursors (such as PhCOC[OH]R2) in oxygen‐containing solutions leads to a laser‐specific cascade of oxidative events in which benzoyl peroxyl radicals undergo self‐reaction, ultimately leading to the sequential formation of benzoyloxyl, phenyl and phenyl peroxyl radicals. Thus, photoreactions producing benzoyl (PheO) radicals lead to Ph?O under conditions where benzoyl radicals do not decarbonylate. These reactions require the high‐intensity conditions that can only be achieved under pulsed laser excitation; yet, they do not involve any multiphoton processes. Type I mechanisms of this type may be of importance as new laser‐based therapeutic technologies are developed.
Photophysical Properties and Photobiological Activity of the Furanochromones Visnagin and Khellin
Authors: Borges Marta L. , Latterini Loredana , Elisei Fausto , Silva Palmira F. , Borges Rui , Becker Ralph S. , Maçanita António L.
Published: July 2010

The larger photobiological activity of visnagin (VI) versus khellin (KH) toward several living organisms, including fungi, viruses, yeasts and bacteria, induced a detailed investigation of the photophysical properties of these naturally occurring furanochromones, using laser‐flash‐photolysis, photoacoustic calorimetry and fluorescence (steady‐state and time‐resolved) techniques in solvents with different polarity and content of water, including micelles and vesicles. The results have shown that the magnitude of all the three rate constants out of S1 (radiative, kf; internal conversion, kic and intersystem crossing, kisc) for VI and KH strongly depend on the solvent, namely on its hydrogen bonding ability and polarity. The changes of kf and kisc are due to the solvent‐assisted mixing and/or inversion of the two first singlet excited states (1n,π* and 1π,π*), while kic increases with a decrease of the So–S1 energy gap. As a consequence, the quantum yield of triplet formation (φT) strongly decreases from values of ˜0.8 in dioxane to < 0.05 in water for both compounds. The magnitude of solvent polarity/hydrogen bonding ability required, at which the state order is inverted and φT starts to decrease, is greater for VI than for KH and consequently φT (VI) > φT (KH) over a broad range of water content including that appropriate to the environment of the compounds in a living system. These facts account for the larger photobiological activity of VI with respect to KH, regarding both the fungus Fusarium culmorum L. and the wild strain of Escherichia coli, studied by us.
Photochemistry of Flavins in Micelles: Specific Effects of Anionic Surfactants on the Monomerization of Thymine Cyclobutane Dimers Photosensitized by Tetra‐O‐acyl Riboflavins*
Authors: Yasuda Masahide , Nishinaka Yoshimasa , Nakazono Takumi , Hamasaki Takeshi , Nakamura Nobuya , Shiragami Tsutomu , Pac Chyongjin , Shima Kensuke
Published: July 2010

It was found that tetra‐o‐acyl riboflavins efficiently photosensitize the monomerization of the cis, syn‐cyclobutane dimers of 1,3‐dimethylthymine and 1,3‐dimethyluracil in aqueous solution in the presence of such anionic surfactants as sodium dodecyl sulfate and sodium hexadecyl sulfate at concentrations higher than their critical micelle concentration, while little monomerization of the dimers was photosensitized by the flavins in the absence of the surfactants and even in the presence of cationic and nonionic surfactants.
Urocanic Acid Does not Photobind to DNA in Mice Irradiated with Immunosuppressive Doses of UVB*
Authors: Ijland Steven A. J. , Noonan Frances P. , Ceryak Susan , Steenvoorden David P. T. , Bouscarel Bernard , Hug Dan , Beijersbergen van Henegouwen Gerard M. J. , De Fabot† Edward C.
Published: July 2010

Ultraviolet B (UVB, 290–320 nm) radiation initiates in vivo a dose‐ and wavelength‐dependent down regulation of cell‐mediated immunity. An action spectrum for UV‐induced immunosuppression indicated that the photoreceptor for this effect is urocanic acid (UCA), which undergoes a trans to cis isomerization in the stratum corneum on UV exposure. An accumulation of evidence has supported this conclusion. However, evidence has a/so been presented that formation of thymine dimers in DNA is responsible for initiation of UV‐induced immunosuppression. Because photobinding of UCA to DNA in vitro forming cyclobutane‐type adducts has been shown, we sought to resolve this dilemma by investigating if UCA photobinds to DNA in vivo. The [14C]cis‐UCA, [14C]trans‐UCA or [3H]8‐MOP (8‐methoxypsoralen) was applied topically to BALB/c mice that were then irradiated with a dose of UV previously shown to cause systemic suppression of contact hypersensitivity. The DNA was prepared from epidermal cells by phenol extraction immediately after in vivo irradiation and bound radioactivity determined. Although photobinding of [3H]8‐MOP was readily demonstrable under these conditions (0.9 nmol/mg DNA), no significant binding of either isomer of UCA to DNA (between 1.2 × 10−3 and 2.1 × 10−3 ng/mg DNA) could be detected. Uptake studies in keratinocytes prepared from epidermis of untreated animals indicated that [3H]8‐MOP was taken up with a rate constant of 4.2 × 10−3 pmol/s/mg protein/μmol/L. In contrast, uptake of [14C]cis‐UCA was not statistically significant from zero and uptake of [14C]trans‐UCA was negligible (0.8 × 10−3± 0.08 × 10 3 pmol/s/mg protein/μmol/L). There was no significant difference between uptake of UCA isomers but uptake of [3H]8‐MOP was significantly greater than that of either UCA isomer (P < 0.01). These studies indicate that the photobinding of UCA to DNA does not play a role in UV‐induced immunosuppression.
p53 Mutations in Hairless SKH‐hr1 Mouse Skin Tumors Induced by a Solar Simulator
Authors: Ananthaswamy Honnavara N. , Fourtanier Anny , Evans Randall L. , Tison Sylvie , Medaisko Chantal , Ullrich Stephen E. , Kripke Margaret L.
Published: July 2010

In this study, we investigated whether the spectrum of p53 mutations in skin tumors induced in hairless SKH‐hr1 mice by a solar simulator (290–400 nm) are similar to those found in skin tumors induced in C3H mice by UV radiation from unfiltered (250–400 nm) and Kodacelfiltered (290–400 nm) FS40 sunlamps. Analysis of tumor DNA for p53 mutations revealed that 14 of 16 (87.5%) SkH‐hr1 skin tumors induced by the solar simulator contained mutations. Single C → T transitions at dipyrimidine sequences located on the nontranscribed DNA strand were the most predominant type of p53 mutation. Remarkably, 52% of all p53 mutations in solar simulator‐induced SKH‐hr1 skin tumors occurred at codon 270, which is also a hotspot in C3H skin tumors induced by unfiltered and Kodacel‐filtered FS40 sunlamps. However, T → G transversions, which are hallmarks of UVA‐induced mutations, were not detected in any of the solar simulator‐induced skin tumors analyzed. These results demonstrate that the p53 mutation spectra seen in solar simulator‐induced SKH‐hr1 skin tumors are similar to those present in unfiltered and Kodacel‐filtered FS40 sunlamp‐induced C3H skin tumors. In addition, our data indicate that the UVA present in solar simulator radiation does not play a role in the induction of p53 mutations that contribute to skin cancer development.
The Structure of the Aggregate Form of Bacteriochlorophyll c Showing the Qy Absorption above 740 nm as Determined by the Ring‐current Effects on 1H and 13C Nuclei and by 1H–1H Intermolecular NOE Correlations
Authors: Mizoguchi Tadashi , Sakamoto Shun , Koyama Yasushi , Ogura Kenji , Inagaki Fuyuhiko
Published: July 2010

13C‐enriched bacteriochlorophyll c (S[I, E] BChl cF) was suspended in a 1:3 mixture of methylene chloride and carbon tetrachloride to form an aggregate showing the Qy absorption above 740 nm; changes in the 13C chemical shifts were traced when methanol was titrated to dissolve the aggregate, and then, the changes were correlated to the ring‐current effects due to the neighboring macrocycles in the aggregate. A pair of aggregate structures has been proposed based on the ring‐current effects on both 1H and 13C nuclei; the monomeric units are stacked together to form an inclined column with different sliding directions, in which the y‐axis of the molecule is parallel to the long axis of the column. In order to confirm this pair of models, the ring‐current effects on the 1H and 13C nuclei were calculated based on both the magnetic‐dipole and the loop‐current approximations. Further, an application of three‐dimensional F1 13C‐edited F3 13C‐filtered heteronuclear single‐quantum correlation‐nuclear Overhauser effect spectroscopy to the above aggregate consisting of a 1:1 mixture of 13C‐labeled and unlabeled BChl c succeeded in detecting selectively the 1H–1H intermolecular nuclear Overhauser effect correlations, which established the coexistence of the above pair of stacked structures in the aggregate.
Protoporphyrin IX Fluorescence Induced in Basal Cell Carcinoma by Oral δ‐Aminolevulinic Acid*
Authors: Tope Whitney D. , Ross E. Victor , Kollias Nikiforos , Martin Alex , Gillies Robert , Anderson R. Rox
Published: July 2010

Limited depth of penetration significantly limits photodynamic therapy of nodular basal cell carcinoma (BCC) using topical δ(5)‐aminolevulinic acid (ALA). To demonstrate safety and efficacy of orally administered ALA in inducing endogenous protoporphyrin IX (PpIX) production in BCC, 13 patients with BCC ingested ALA in a dose‐escalation protocol. All dose ranges (10, 20 or 40 mg/kg single doses) resulted in formation of PpIX in human skin and BCC, measurable by in vivo fluorescence spectrophotometry. The PpIX fluorescence peaked in tumors before normal adjacent skin from 1 to 3 h after ALA ingestion. Gross fluorescence imaging of ex vivo specimens revealed greater PpIX fluorescence in tumor than normal skin only at the 40 mg/kg dose. Fluorescence microscopy confirmed this finding by showing distinct, full‐thickness PpIX fluorescence in all subtypes of BCC only after ALA given at 40 mg/kg. Side effects were dose dependent and self limited. Photosensitivity lasting less than 24 h and nausea coinciding with peak skin PpIX fluorescence occurred at 20 and 40 mg/kg doses. After 40 mg/kg ALA, serum hepatic enzyme levels rose to a maximum within 24 h, then resolved over 1–3 weeks. Transient bilirubinuria occurred in two patients.
Contribution of the cAMP‐Dependent Signal Pathway to Circadian Synchrony of Motility and Resting Membrane Potential in Paramecium
Authors: Hasegawa Kenji , Tsukahara Yasuo , Ishizaki Shigeo , Shimamoto Masakazu , Nakamura Takeshi , Sohma Masaki , Sato Toshiro
Published: July 2010

It is known that the ciliated protozoan Paramecium multimicronucleatum has synchronized circadian rhythms of motility, resting membrane potential and cyclic adenosine 3′,5′‐monophosphate (cAMP) and cyclic guanosine 3′,5′‐monophosphate (cGMP) concentrations. The present study shows that (1) extracellularly added 4 mM tetraethylammonium (TEA)+ (a K+ channel blocker) almost completely abolishes the diurnal oscillation of intracellular cAMP concentrations; (2) even 32 mM TEA+ fails to abolish the circadian motility rhythm; but (3) the motility rhythm is highly damped when 4 mM TEA+ and 100 μM CdCl2 (a Ca2+ channel blocker) are added simultaneously. A cAMP analogue (N6‐monobutyryl‐cAMP) added extracellularly accelerates swimming velocity. Both a K+ channel blocker (e.g. TEA+) and an inhibitor (trifluoperazine) of adenylate cyclase (AC) suppress cAMP formation, supporting the hypothesis that AC in Paramecium has dual functions, as a K+ channel and as an enzyme for cAMP formation. It is hypothesized that the circadian synchrony is due to circadian fluctuations of AC causing separate circadian changes both in ciliary motion and membrane potential through a cAMP‐dependent signal pathway that forms a sophisticated network of second messengers to govern the synchrony together with Ca2+‐ and cGMP‐dependent pathways in a manner antiphasic and/or complementary to one another.
Subcellular Distributions and Excited‐State Processes of Hypericin in Neurons
Authors: English Doug. S. , Doyle Robert T. , Petrich Jacob W. , Haydon Philip G.
Published: July 2010

Abstract— The photodynamic drug, hypericin, is studied in fetal rat neurons using fluorescence microscopy. Hypericin has an extremely high affinity for the cell membrane and is found to a smaller extent in the nucleus. Fluorescent excitation of hypericin is shown to cause irreversible damage to the cell membranes of living neurons. Fixed cells were used to make ultrafast time‐resolved measurements to avoid the deleterious effects of long‐term exposure to intense light and room temperatures. To our knowledge, these are the first ultrafast time‐resolved measurements of the fluorescence lifetime of hypericin in a subcellular environment. Nonexponential fluorescence decay is observed in hypericin in the neurons. This nonexponential decay is discussed in terms of other examples where non‐exponential decay is induced in hypericin upon its binding to biomolecules. The nonaradiative processes giving rise to the nonexponential hypericin decay are attributed to excited‐state electron transfer, excited‐state proton transfer or both.
Can Cellular Phototoxicity be Accurately Predicted on the Basis of Sensitizer Photophysics?
Authors: Aveline Beatrice M. , Redmond Robert W.
Published: July 2010

Abstract— The phototoxicity of three structurally related photosensitizers (PS), deuteroporphyrin IX (DP) and monobromo (Br‐DP) and dibromo (Br2‐DP) derivatives, was studied in murine L1210 leukemia cells. These compounds were chosen on the basis of heavy‐atom‐induced differences in triplet yield, ΦT, and lifetime, TT, and used as tools to test a model for phototoxicity based on photophysical parameters. All three porphyrins were found to localize preferentially in the plasma membrane of L1210 cells by confocal fluorescence microscopy. A poor correlation was observed between the measured photodynamic efficacies of these PS and a model using photophysical parameters determined by laser flash photolysis in homogeneous solution. However, an excellent correlation was obtained when the same parameters measured directly in the cells were used. The biological microenvironment of the porphyrins in cells induces significant changes in the photophysics of the PS. Reduction in fluorescence yield, φF, and φT observed for Br2‐DP in cell suspensions arises from self association of the molecule due to increased hydrophobicity and high local concentrations. The photophysical model was also tested for its ability to handle variations in the oxygen dependence of cellular phototoxicity of these PS. The good correlation achieved between laser flash photolysis data determined in cells and the measured phototoxicity under air, 1.5% and 0.5% O2‐saturated conditions, proves the intermediacy of singlet oxygen. This study gives further credence to the direct use of photophysical techniques to elucidate photochemical mechanisms in biological media while highlighting the potential pitfalls of using solution data to predict photosensitizing potential.
Stratum Corneum Lipid Abnormalities in UVB‐lrradiated Skin
Authors: Meguro Shinichi , Aral Yoichi , Masukawa Katsunori , Uie Kazuyoshi , Tokimitsu Ichiro
Published: July 2010

Abstract— Stratum corneum (SC) lipids are of particular importance in maintaining the permeability barrier function. Although many studies have demonstrated that UVB irradiation of mammalian skin reduces barrier function, the responsible alterations in SC lipid profiles are not known. In this study, we investigated both compositional and morphological alterations in SC lipids with the development of barrier abnormalities caused by daily UVB irradiation in hairless rat skin. The UVB irradiation of suberythemal doses (0.5 minimal erythema dose) significantly increased transepidermal water loss (TEWL) relative to nonirradiated control, indicating a diminished barrier function. Under these conditions, the total amounts of major SC lipid species (ceramides, cholesterol, free fatty acids) in UVB‐irradiated SC did not differ from those in nonirradiated SC. However, electron microscopic observations revealed marked abnormalities in the intercellular domains of UVB‐irradiated SC, where naturally occurring intercellular multilamellar structures were often absent and leaving the area with the appearance of an empty space. Moreover, in UVB‐irradiated SC, individual corneocytes often showed small amounts of intercellular deposition product with abnormal lamellar structure, where lamellar body sphingomyelinase activity was present. These observations demonstrated a partial failure of lamellar body secretion in UVB‐irradiated SC and suggested that a defect in the secretion of lamellar body‐derived lipids and enzymes to SC intercellular space is, at least in part, responsible for the observed abnormal intercellular structure and barrier disruption.
Detection of Colonic Malignant Lesions by Digital Imaging of UV Laser‐Induced Autofluorescence
Authors: Chwirot Barbara W. , Michniewicz Zbigniew , Kowalska Malgorzata , Nussbeutel Joanna
Published: July 2010

Abstract— The objective of our study was to investigate whether digital imaging of autofluorescence could be applied in the detection of colonic malignancies. Autofluorescence was excited with a 325 nm line from a He‐Cd laser. Images were recorded in vitro in six spectral bands. The material for study was 50 resected specimens for which images of 64 areas were recorded. The main result is the observation that for a majority of malignant and premalignant lesions the intensity of autofluorescence was lower than for the corresponding normal mucosa in all of the spectral bands selected for imaging. The spectral bands centered around 440 nm and 475 nm seem to be most promising in terms of possible future clinical applications.
Salt Water Bathing Prior to UVB Irradiation Leads to a Decrease of the Minimal Erythema Dose and an Increased Erythema Index without Affecting Skin Pigmentation
Authors: Schempp Christoph M. , Müller Knut , Schulte‐Mönting Jürgen , Schöpf Erwin , Simon Jan C.
Published: July 2010

Abstract— The combination of salt water baths and solar radiation is known as an effective treatment for patients with psoriasis and atopic dermatitis. To determine whether increased susceptibility to UVB radiation may contribute to this therapeutic effect we have studied the effect of bathing the skin in salt water prior to UVB irradiation. Twelve subjects were phototested on the volar aspects of their forearms with increasing doses of UVB radiation. One forearm was exposed to 5% salt water prior to irradiation. The minimal erythema dose (MED) was determined and the erythema index and skin pigmentation were assessed by photometric measurement. The combination of salt water bath and irradiation yielded a significant decrease of the MED when compared to UVB alone (median 90 mJ/cm2vs 130 mJ/cm2P < 0.01). Analysis of variance showed a significant influence of salt water bath on erythema (P < 0.05) but not on skin pigmentation. Within the MED test area the erythema index of the salt water exposed forearms was elevated significantly (P < 0.05) while skin pigmentation was not affected. Thus, bathing the skin in salt water leads to a decreased threshold level for the elicitation of UVB‐induced erythema and a selective increase of the erythemal response. This sensitization to the effects of shortwave UVB radiation may increase immunosuppressive effects of UVB radiation and may lead to an increased efficacy of UVB phototherapy. However, there is also an increased sunburn risk when salt water baths are followed by exposure to UV radiation.
Protoporphyrin IX‐Sensitized Photoinactivation of 5‐Aminolevulinate‐Treated Leukemia Cells: Effects of Exogenous Iron
Authors: Lin Fubao , Geiger Peter G. , Korytowski Witold , Girotti Albert W.
Published: July 2010

Abstract— Photodynamic therapy with 5‐aminolevulinic acid (ALA) is based on metabolism of ALA to a photosensitizing agent, protoporphyrin IX (PpIX), in tumor cells. Photo‐sensitivity of target cells may be influenced by mitochondrial iron levels because ferrochelatase‐catalyzed insertion of Fe2+ into PpIX converts it to heme, a nonsensitizer. To investigate this prospect, we exposed L1210 cells (∼106/mL in 1% serum‐containing medium) to a lipophilic iron chelate, ferric‐8‐hydroxyquinoline (Fe[HQ]2, 0.5 μM), prior to treating with ALA (0.2 mM, 4 h) and irradiating with broadband visible light. When Fe(HQ)2 was added to cells immediately or 1 h before ALA, the initial rate of photokilling, as measured by thiazolyl blue (mitochondrial dehydrogenase) assay, was markedly less than that of non‐iron controls. The HPLC analysis of cell extracts indicated that ALA‐induced PpIX was at least 50% lower after this Fe(HQ)2 treatment, presumably explaining the drop in photolethality. By contrast, cells treated with ALA and light 20 h after being exposed to Fe(HQ)2 contained the same amount of PpIX as non‐iron controls and were photoinactivated at nearly the same rate. The 20 h delayed cells contained ∼12 times more immunodetectable ferritin heavy subunit than controls or 1 h counterparts, which could account for the disappearance of iron's antisensitization effects in the former. Consistent with this idea, the short‐term effects of Fe(HQ)2 on ALA‐induced sensitization were found to be blunted significantly in ferritin‐enriched cells. The Fe(HQ)2 produced strikingly different results when cells were sensitized with exogenous PpIX, stimulating photokilling after short‐term contact but inhibiting it after long‐term contact while having no significant effect on the level of cell‐associated PpIX in either case. Thus, iron can have diverse effects on PpIX‐mediated photokilling, depending on contact time with cells and whether the porphyrin is metabolically derived or applied as such.
Unrepaired Cyclobutane Pyrimidine Dimers Do Not Prevent Proliferation of UV‐B‐irradiated Cultured Human Fibroblasts¶
Authors: Courdavault Sophie , Baudouin Caroline , Sauvaigo Sylvie , Mouret Stéphane , Candéias Serge , Charveron Marie , Favier Alain , Cadet Jean , Douki Thierry
Published: July 2010

Mutagenic and carcinogenic UV‐B radiation is known to damage DNA mostly through the formation of bipyrimidine photoproducts, including cyclobutane dimers (CPD) and (6‐4) photoproducts ((6‐4) PP). Using high‐performance liquid chromatography coupled to tendem mass spectrometry, we investigated the formation and repair of thymine‐thymine (TT) and thymine‐cytosine (TC) CPD and (6‐4) PP in the DNA of cultured human dermal fibroblasts. A major observation was that the rate of repair of the photoproducts did not depend on the identity of the modified phyrimidines. In addition, removal of CPD was found to significantly decrease with increasing applied UV‐B dose, whereas (6‐4) PP were efficiently repaired within less than 24 h, irrespective of the dose. As a result, a relatively large amount of CPD remained in the genome 48 h after the irradiation. Because the overall applied doses (<500 J m−2) were chose to induce moderate cytotoxicity, fibroblasts could recover their proliferation capacities after transitory cell cycle arrest, as shown by 5‐bromo‐2′‐deoxyuridine (BrdUrd) incorporation and flow cytometry analysis. It could thus be concluded that UV‐B‐irradiated cultured primary human fibroblasts normally proliferate 48 h after irradiation despite the presence of high levels of CPD in their genome. These observations emphasize the role of CPD in the mutagenic effects of UV‐B.
Comparative Study of Photodynamic Properties of 13, 15‐N‐cycloimide Derivatives of chlorin p6¶
Authors: Feofanov Alexei , Sharonov George , Grichine Alexei , Karmakova Tatyana , Pljutinskaya Anna , Lebedeva Victoria , Ruziyev Ramezes , Yakubovskaya Raisa , Mironov Andrei , Refregier Matthieu , Maurizot Jean‐claude , Vigny Paul
Published: July 2010

Comparative study of 13,15‐[N‐(2‐hydroxyethyl)]cycloimide chlorin p6 (2), 13,15‐(N‐acetoxy)cycloimide chlorin p6 (3), 13,15‐(N‐hydroxy)cycloimide chlorin p6 methyl ester (4) and 13,15‐(N‐methoxy)cycloimide chlorin p6 methyl ester (5) together with the previously investigated 13,15‐[N‐(3‐hydroxypropyl)]cycloimide chlorin p6 (1) was performed. The dependence of the key photodynamic properties of 1–5 on the introduced substituents was analyzed. The photoinduced cell‐killing activity of 4 is 100‐ and 280‐fold higher than that of chlorin p6 and Photogem, respectively, as estimated on A549 human lung adenocarcinoma cells. The activity is reduced eight times in the order 4 > 5 > 1 > 2 > 3. The intracellular accumulation of 1–5 occurs in cytoplasm in a monomeric form bound to the lipids of cellular membranes. This form of 1,2,3, 4 and 5 is characterized by the high quantum yield of singlet oxygen generation, which depends on the introduced substituents, 0.66, 0.59, 0.35, 0.51 and 0.73, respectively. The photostability is two‐fold less for 1 and four‐fold less for 2, 3 and 5 than for 4. The rates of cellular uptake and efflux of 1–5 vary widely, thus providing the way to optimize the pharmacological properties of the photosensitizer (PS) using the respective substituents. Modifying the substituents, 1–5 were targeted to different cellular organelles. The enhanced accumulation in the Golgi apparatus and mitochondria complemented with diffuse staining of intracellular membranous structures is a property of 1–4. Compound 5 accumulates selectively in the lipid droplets and stains weakly perinuclear structures. Temperature‐sensitive mechanisms of transport are responsible for the 1–4 uptake. Diffusion can play a role in the internalization of 5 but not of 1–4. Endocytosis via caveolae, clathrin‐dependent and adenosine triphosphate‐dependent pathways are not noticeably involved in the 1–5 internalization. Independently from their intracellular localization 1,4 and 5 are highly efficient near‐IR PS, which induce predominantly an apoptotic type of cell death under conditions providing ca 50% level of phototoxicity and necrosis at the 100% level of phototoxicity.
Development of a Test System for Mutagenicity of Photosensitizers Using Drosophila melanogaster¶
Authors: Smijs Threes G. M. , Nivard Madeleine J. M. , Schuitmaker Hans J.
Published: July 2010

In the past few years, there has been an increase in the application of photosensitizers for medical purposes. A good standardized test system for the evaluation of the mutagenic potentials of photosensitizers is therefore an indispensable device. In the standard Ames test, white light itself was proven to be mutagenic and the result influenced by the light source. Lack of a reliable positive control is another problem in many genotoxicity test systems used for the evaluation of mutagenicity of photosensitizers. Based on the validated somatic mutation and recombination test, known as SMART, and using Drosophila melanogaster, we developed the Photo‐SMART and demonstrated that methylene blue, known to induce photomutagenicity, can act as a positive control in the presented test system. The SMART scores for the loss of heterozygosity caused predominantly by homologous mitotic recombination. The Photo‐SMART can be used to detect photogenotoxicity caused by short‐lived photoproducts or by stable photoproducts or both. We demonstrated the Photo‐SMART to be a good standardized test system for the evaluation of mutagenic potentials of the photosensitizer 5,10,15‐tris(4‐methylpyridinium)‐20‐phenyl‐[21H,23H]‐porphine trichloride (TPP). We demonstrated that TPP was mutagenic using the Photo‐SMART. For hematoporphyrin, the results of the Photo‐SMART indicate the absence of mutagenicity.
Survival and Recovery of Yeast Cells After Simultaneous Treatment of UV Light Radiation and Heat¶
Authors: Kim Jin Kyu , Petin Vladislav G. , Tkhabisimova Marianna D.
Published: July 2010

Cell survival, synergistic interaction, liquid‐holding recovery (LHR) kinetics and inactivation forms after the simultaneous treatment with UV light (254 nm) and various high temperatures were studied in diploid yeast cells Saccharomyces cerevisiae. The synergistic interaction was observed within a certain temperature range in which there was a temperature that maximizes the synergistic effect. The LHR study revealed that both the extent and the rate of recovery greatly decreased with the increase in exposure temperature. A quantitative approach describing the LHR process as a decrease in the effective radiation dose was used to estimate the probability of recovery per unit time and the irreversible component of damage. Using the experimental data obtained and the mathematical model described, it was shwon that the irreversible component, i.e. the fraction of cells incapable of recovery, increased with the exposure temperature, whereas the recovery constant, i.e. the probability of recovery per unit time, was independent of the exposure temperature. The increase in the irreversible component was accompanied by an increase in cell death without postirradiation division. It is concluded based on this that the synergistic interaction of UV light radiation and hyperthermia in yeast cells is not related to the impairment of the recovery process itself and that it may be attributed to an increased yield of the irreversible damage.
Personal solar UV Exposure Measurements Employing Modified Polysulphone with an Extended Dynamic Range†¶
Authors: Parisi Alfio V. , Kimlin Michael G.
Published: July 2010

Polysulphone dosimeters using a simple to use filter have been developed and tested to provide an extended dynamic measurement range of personal solar W exposures over an extended period (3 to 6 days). At a Southern Hemisphere subtropical site (27.6°S, 151.9°E), the dynamic range of the filtered polysulphone allowed measurements of erythemal exposures to approximately 100 minimum erythemal dose (MED) for a change in optical absorbance at 330 nm (AA330) of 0.35. In comparison, unfiltered polysulphone dosimeters were exposed to approximately 8 MED for the same AA330. The error associated with the use of the filtered polysulphone dosimeters is of the order of ±15%, compared with ±10% of the unfiltered variety. The developed filtered polysulphone dosimeter system allowed the measurement of erythemal UV exposures over 3 to 6 days at a subtropical site without the need to replace the dosimeters because of saturation. The results show that longer‐term measurement programs of personal solar UV have been made more feasible with the use of these polysulphone dosimeters with an extended dynamic range compared with unfiltered polysulphone dosimeters.
Kinetics and Mechanism of the Vitamin B2‐sensitized Photooxidation of Isoproterenol¶
Authors: Massad Walter A. , Bertolotti Sonia , Garcia Norman A.
Published: July 2010

The sensitized photooxidation promoted by daylight‐absorbing compounds appears as a plausible course to produce the photodegradation of catecholamines. We report the kinetics and mechanism of vitamin B2 (reiboflavin [Rf])‐sensitized photooxidation of isoproterenol (Isoproterenol (Iso), a synthetic sympathomimetic drug structurally related to epinephrine, using water as a solvent. A week dark complex Rf‐Iso is formed, only detectable at relatively high Iso concentrations (>10 mM), with a mean value of 13 ± 3 M−1 for the apparent association constant. Under aerobic sensitizing conditions (Rf ∼ 0.02 mM and Iso ∼ 0.5 mM) two oxidative mechanisms operate, mediated by singlet molecular oxygen (O2(Δg)) and superoxide radical anion (O2•−). Our analysis shows that the main reaction pathway is an electron transfer‐mediated quaenching of Rf excited triplet state (3Rf*) by Iso. It produces the species Iso•+ and Rf•−. The latter, in a subsequent reaction path, generates O2•−, which is mainly responsible for Iso Photooxygenation. In a less‐important process, energy transfer of the 3Rf* to dissolved oxygen generates O2(Δg). The kinetic balance between chemical and physical quenching of O2(1Δg) by Iso indicates that the process is largely dominated by the physical, not chemical, interaction. The results, which can be extrapolated to an in vivo condition, show the susceptibility of Iso to undergo visible light‐induced photodegradation in the presence of dye sensitizers present in the environment.
Flexibility in Proteins: Tuning the Sensitivity to O2 Diffusion by Varying the Lifetime of a Phosphorescent Sensor in Horseradish Peroxidase¶
Authors: Nibbs Janna , Vinogradov Sergei A. , Vanderkool Jane M. , Zelent Bogumil
Published: July 2010

The heme in horseradish peroxidase (HRP) was replaced by phosphorescent Pt‐mesoporphyrin IX (PtMP), which acted as a phosphorescent marker of oxygen quenching and allowed comparison with another probe, Pd‐mesoporphyrin IX (Khajehpour et al. (2003) Proteins 53, 656–666). Benzohydroxamic acid (BHA), a competitive inhibitor of the enzyme, was also used to monitor its effects on phosphorescence quenching. With the addition of BHA, in the presence of oxygen, the phosphorescence intensity of the protein increased. In contrast, the addition of BHA, in the absence of oxygen, reduced the phosphorescence intensity of the protein. Kd= 18 μM when BHA binds to PtMP‐HRP. The effect of BHA can be explained by two factors: (1) BHA reduces the accessibility of O2 to the protein interior and (2) BHA itself quenches the phosphorescence. Consistent with this, the oxygen quenching of the phosphorescence of PtMP‐HRP gave a quenching constant of kq= 234 mm Hg−1 s−1 in the absence of BHA and kq= 28.7 mm Hg−1 s−1 in the presence of BHA. The quenching rate of BHA is 4000 s−1. The relative quantum yield of the phosphorescence of the Pt derivative is about six times that of the Pd derivative, whereas the phosphorescence lifetime is approximately eight times shorter. The high quantum yield and suitable lifetime make Pt‐porphyrins appropriate as sensors of O2 diffusion and flexibility in heme proteins.
Virus Particles Monitored by Fluorescence Spectroscopy: A Potential Detection Assay for Macromolecular Assembly¶
Authors: Alimova Alexandra , Katz A. , Podder Rakhi , Minko Glenn , Wei Hul , Alfano R. R. , Gottlieb Paul
Published: July 2010

Native fluorescence spectroscopy was used for in situ investigations of two lipid‐containing bacteriophages from the cystovirus family as well as their Pseudomonad host cells. Both the viruses φ6 and φ12 and their bacterial host proteins contain the amino acid tryptophan (trp), which is the predominant fluorophore in UV. Within proteins, trp's structural environment differs, and the differences are reflected in their spectroscopic signatures. It was observed that the peak of the trp emission from both viruses was at 330 nm, a significantly shorter wavelength than trp in either the Pseudomonad host cells or the amino acid's chemical form. This allowed us to monitor the viral attachment process and subsequent lytic release of progeny virus particles by measurement of the trp emission spectra during the infection process. This work demonstrates that fluorescence may offer a novel tool to detect viruses and monitor viral infection of cells and may be part of a biodefense application.