bluuebunny/crossref_metadata_2025 huggingface dataset:
CELLULAR DELIVERY AND RETENTION OF PHOTOFRIN: III. ROLE OF PLASMA PROTEINS IN PHOTOSENSITIZER CLEARANCE FROM CELLS
Authors: Korbelik Mladen
Published: January 2008

Abstract— Human plasma proteins, albumin, globulins and low density (LDL), high density (HDL) and very low density (VLDL) lipoproteins were tested for their effects on retention of Photofrin and three other photosensitizers in cultured cells. This was assessed by incubating the cells, subsequent to the exposure to Photofrin, in the photo‐sensitizer‐free medium containing various concentrations of different plasma proteins. Photofrin clearance levels differed with individual plasma proteins and also were dependent on concentration of these proteins in the incubation medium. All of the proteins except VLDL promoted clearance of Photofrin taken up by the cells in the presence of 5% human serum. Subsequent to some Photofrin exposure conditions (in the presence of 5% fetal bovine serum, or in protein‐free medium), albumin, in contrast to LDL, HDL and globulins, exhibited decreased capacity for promoting the photosensitizer clearance from the cells. The VLDL showed very little or no effect in promoting cellular clearance of Photofrin, tetraphenyl porphine tetrasulfonate (TPPS4), and di‐ and tetrasulfonated chloroaluminum phthalocy‐anine (AlPcS2 and AlPcS4, respectively). The LDL seem to be particularly effective in promoting clearance of Photofrin and AlPcS2 from the cells, whereas albumin and globulins were shown to be more effective than LDL and HDL in promoting the cellular clearance of TPPS4.
INFLUENCE OF HYDRATION ON THE INTERNAL DYNAMICS OF HEN EGG WHITE LYSOZYME IN THE DRY STATE
Authors: Shah Naina K. , Ludescher Richard D.
Published: January 2008

Abstract— Proteins exist in a predominately aqueous solvent environment. Hydration of the protein surface significantly affects many aspects of the protein's structure and function; these effects may be related to the molecular dynamics of the protein. We have examined the influence of hydration on the internal dynamics of hen egg white lysozyme using room‐temperature phosphorescence from the intrinsic tryptophan residues. Powders of lyophilized lysozyme were hydrated in a phosphorimeter using a flow system that allowed for continuous manipulation of relative humidity over the range 0–92%; this system allowed us to directly compare intensity differences that result from changes in hydration. Lysozyme phosphorescence intensity decreased as a function of hydration over the entire relative humidity range; the decrease was not linear but appeared to occur in distinct phases. The phosphorescence intensity decays were multiexponential over the hydration range studied, and hydration had the largest influence on the long lifetime component. These data suggest that the protein exists in multiple, static conformations in the dry state and that water binding to polar (as opposed to charged) sites on the protein surface induces local and/or global softening of the protein structure.
UPPER EXCITED STATE PHOTOCHEMISTRY OF DNA
Authors: Gut Ivo G. , Farmer Raymond , Huang Russel C. , Kochevar Irene E.
Published: January 2008

Abstract— The quantum yields for cyclobutylpyrimidine dimers, alkali‐labile sites, and frank strand breaks in double‐stranded DNA have been measured using low‐intensity radiation at 199.8, 217.8, and 239.5 nm from a Raman‐shifted frequency quadrupled Nd:YAG laser. The quantum yield for cyclobutylpyrimidine dimers was also measured using 254 nm radiation from a low‐pressure mercury lamp. The quantum yield for cyclobutylpyrimidine dimers is constant within a factor of two between 254 and 199.8 nm except for 239.5 nm, indicating that upper excited singlet states of bases convert efficiently to the lowest singlet state. The quantum yields for alkali‐labile sites and frank strand breaks both increase as the wavelength decreases but follow different patterns. These results indicate that alkali‐labile sites form from a higher excited state of the base, whereas frank strand breaks form by excitation of the sugar‐phosphate backbone.
STRESS PROTEIN EXPRESSION IN MURINE TUMOR CELLS FOLLOWING PHOTODYNAMIC THERAPY WITH BENZOPORPHYRIN DERIVATIVE
Authors: Curry P. Mark , Levy Julia G.
Published: January 2008

Abstract— Photodynamic therapy (PDT) has been proven as a method of tumor eradication and is currently being used clinically to treat a wide variety of malignancies. Although it is understood that the interaction of light and sensitizer results in the production of potentially damaging oxygen species, the mechanism by which tumors are destroyed has yet to be defined fully. Using a new porphyrin sensitizer, benzoporphyrin derivative(BPD), we examined protein expression in murine tumor cells following treatment as an indication of molecular changes to target tissue concurrent with PDT‐mediated damage. In order to assess the relevance of the results obtained using an in vitro PDT model, metabolic labeling of proteins synthesized subsequent to PDT was performed both in tumor cells grown and treated in tissue culture dishes and in cells explanted from PDT‐treated solid tumors. We observed that the oxidative stress associated with PDT‐resulted in the induction of a number or proteins corresponding to a set of heat‐shock or stress proteins, and that the pattern of expression was similar when tumor cells were treated in vitro and in vivo. These results support the use of in vitro models in the dissection of the molecular erects of PDT and provide the foundation for future experiments that will examine the role of the immune system in tumor eradication by PDT.
UV INDUCTION OF CYCLOBUTANE THYMINE DIMERS IN THE DNA OF CULTURED MELANOCYTES FROM FORESKIN, COMMON MELANOCYTIC NEVI and DYSPLASTIC NEVI
Authors: Noz Käthe C. , Roza Len , Bergman Wilma , Darroudi Firouz , Schothorst Albert A.
Published: January 2008

Abstract— We compared the induction of cyclobutane thymine dimers after exposure to 302 nm UV in foreskin‐derived melanocytes and melanocytes from nevocellular nevi, as well as in melanocytes cultured from dysplastic nevi, precursor lesions of melanoma, derived from four, three and four individuals, respectively. Cyclobutane thymine dimers were quantified in situ by means of an immunofluorescence assay with a specific monoclonal antibody. A method was developed to compare separately performed experiments in a standardized manner. For melanocytes from each source, we demonstrated a linear relationship between UV dose and immunofluorescence. In nevocellular and dysplastic nevi, two subpopulations could be detected, distinguished by their nuclear size. Large nucleated nevocellular nevus cells were most susceptible to the induction of thymine dimers (49% higher induction compared to induction in foreskin melanocytes), while in normal‐sized nuclei of these nevus cells the same induction of thymine dimers was found as in nuclei from foreskin melanocytes. In contrast, large nucleated dysplastic nevus melanocytes did not differ from the foreskin melanocytes, while normal‐sized nuclei of dysplastic nevus cells showed a lower induction (32% lower induction than in foreskin melanocytes).
TWO SPECTROSCOPICALLY AND PHOTOCHEMICALLY DISTINGUISHABLE PHYTOCHROMES IN ETIOLATED SEEDLINGS OF MONOCOTS AND DICOTS *
Authors: Sineshchekov Vitally
Published: January 2008

Abstract Fluorescence (F) emission spectra of the red‐absorbing phytochrome form (Pr) at 85 K, temperature dependence of the F intensity and the extent of the Pr F changes in the phototransformation of Pr into the first stable photoproduct (lumi‐R) at 85 K (γ1,) and into the far‐red‐absorbing form (Pfr) at 267 K (γ2) were investigated in etiolated shoots and roots of monocots (oat, maize, rice) and dicots (pea, cress). These characteristics monotonously changed as a function of the phytochrome content, [Ptot]: with its decrease to 3‐5% of the maximal values, the F spectrum shifts from 686 nm to 682 nm, its half‐band width rises from 22 nm to ca 25 nm, temperature dependence of Pr F changes its character, γ1, drops down from ca > 0.45 to ca 0.05‐0.10 and γ2 from 0.80–0.82 to ≤0.70. These data were interpreted in terms of two different phytochromes whose relative concentration varies with [Ptot]: (1) a longer wavelength type with the F maximum at 686 nm, low activation energy of the photoreaction (Ea ≤ 3–4 kj/ mol) and high extent of the phototransformation at 85 K (0.49 ± 0.03) and at 267 K (ca 0.85) (Pra); (2) a shorter wavelength type practically inactive at 85 K with F maximum at 682 nm, higher Ea (ca 35 kj/mol) and lower extent of the Pr & Pfr phototransformation (≤0.70) (Pri). [Pra] widely varies in different parts of the seedlings (up to 100 times) and Pra dominates when [Ptot] is high. The [Pri] is much more constant (variations, <10 times), and it becomes the major one when [Ptot] drops down. The two species are likely to belong to the labile (type 1) and stable pools of pigment and not to be connected with the localization of the pigment in the cell since red‐far‐red preillu‐mination, which is believed to bring about sequestering of the pigment, does not change their relative concentration and properties.
REACTIVITY OF SINGLET OXYGEN TOWARD AMINO ACIDS AND PEPTIDES
Authors: Michaeli Albert , Feitelson Jehuda
Published: January 2008

Quenching of singlet oxygen (1O2) in D2O‐ethanol by the amino acids tryptophan, tyrosine, histidine, methionine, cysteine and their derivatives was measured by exciting the sensitizers rose bengal or meso‐tetra (N‐methyl‐4‐pyridyl)porphyrin tetratosylate in the presence of oxygen and the above quenchers in solution. In our polar solvent, containing 75% D2O on a molar basis it was found that (1) substitution of the aromatic ring in indole, phenol and imidazole by the electron‐donating methyl group increases the total (i.e. nonreactive and reactive) quenching rate constant by a factor of five to eight. Free or blocked amino and carboxyl groups removed by two methylene groups from the ring counteract the above increase in the rate constant. The reactive quenching of singlet oxygen, which leads to oxidative destruction of the aromatic ring, correlates with the above substitution effects. It has been proposed that the quenching process takes place by formation of an exciplex between 1O2 and the quencher. Thus our results indicate that the better an electron donor the amino acid residue is the more pronounced is the charge transfer contribution in the exciplex formed with 1O2 and the more likely it is to lead to charge separation and hence to a chemical reaction. (2) Oligopeptides in solution or peptide bonds linked to the amino acid residue have only a minor effect on singlet oxygen. It can therefore be expected that the polypeptide chains per se in the protein network will not interact significantly with the single oxygen molecules present. The quenching of the latter should, to a first approximation, depend only on the presence of the above reactive amino acid residues and to their accessibility to 1O2 as well as on the effective dielectric constant within the protein structure.
HEMATOPORPHYRIN DERIVATIVE‐INDUCED PHOTODYNAMIC INHIBITION OF Na+/K+‐ATPase IN L929 FIBROBLASTS, CHINESE HAMSTER OVARY CELLS AND T24 HUMAN BLADDER TRANSITIONAL CARCINOMA CELLS
Authors: Penning Louis C. , Tijssen Karmi , VanSteveninck John , Dubbelman Tom M. A. R.
Published: January 2008

The photodynamically induced inhibition of Na+/K+‐ATPase with hematoporphyrin derivative as photosensitizer was studied in murine L929 fibroblasts, Chinese hamster ovary (CHO) cells and T24 human bladder transitional carcinoma cells. In T24 cells the inhibition of the enzyme activity appeared to be caused by ATP depletion rather than by direct damage from the enzyme itself. In L929 and CHO cells, on the other hand, the inhibition was caused by photodynamic damage from the enzyme molecule. For all three cell lines it was shown that a causal relationship between photodynamically induced reduction in Na+/K+‐ATPase activity and the loss of clonogenicity is highly unlikely.
ACTIVATION OF BENZOPORPHYRIN DERIVATIVE IN THE CIRCULATION OF MICE WITHOUT SKIN PHOTOSENSITIVITY
Authors: Richter Anna M. , Jain Ashok K. , Obochi Modestus , Meadows Howard , Canaan Alice J. , Levy Julia G.
Published: January 2008

In vitro experiments with benzoporphyrin derivative monoacid ring A (BPD) confirmed earlier studies that it was taken up rapidly (within 30 min) to maximum concentrations by all cells tested. It was also confirmed that rapidly dividing tumor cell lines and mitogen‐activated murine T lymphocytes took up significantly more (5‐10‐fold) BPD than did normal splenic lymphocytes. Further experiments were undertaken to determine whether BPD could be activated by whole‐body irradiation with red light in the blood of animals, shortly after intravenous (i.v.) administration, in the absence of skin photosensitivity. It was found that shaved and depilated mice injected i.v. 60 min earlier with BPD at between 0.5 and 1.0 mg/kg could tolerate 160 J/cm2 of broad‐band red light (560‐900 nm) delivered, at a relatively low rate, over a 90 min time interval without developing skin photosensitivity or general phototoxicity. During the treatment time, plasma levels of BPD were between 0.7 and 1.0 μg/mL. The light treatment resulted in between 70 and 80% photoinactivation of circulating BPD. When LI 210 tumor cells were preincubated with BPD and injected i.v. into mice immediately before total‐body light treatment (160 J/cm2 of 590‐900 nm light delivered over 90 min), significant reductions in circulating clonogenic tumor cells were observed in blood samples taken immediately following treatment. This indicated that sufficient light was being delivered to BPD in the blood flowing in the peripheral vasculature to effect cytotoxicity to cells containing the photosensitizer without causing either vascular or skin photosensitivity. Thus, activation of this photosensitizer in the circulation can be achieved by transdermal light exposure without causing skin photosensitivity provided that light exposure is performed at a time when the first phase of plasma clearance is complete and when the drug has not yet accumulated in skin.
PYRIMIDINE DIMER INDUCTION AND REMOVAL IN THE EPIDERMIS OF HAIRLESS MICE: INEFFICIENT REPAIR IN THE GENOME OVERALL AND RAPID REPAIR IN THE H‐ras SEQUENCE
Authors: Lan Ruiting , Greenoak Gavin E. , Moran Chris
Published: January 2008

Excision repair of pyrimidine dimers was examined at the genome overall in three strains of hairless (hr/hr) and congenic wild‐type mice, as well as in the expressed H‐ras gene in hairless mice. The assay used a pyrimidine dimer‐specific endonuclease from Micrococcus luteus and alkaline agarose gel electrophoresis. From 0 to 25% of endonuclease‐sensitive sites were removed at the genome level in either hairy or hairless mice but about 50% were removed in the H‐ras gene in hairless mice by 24 h after exposure to 5.4 J/cm2 UV (290‐400 nm) irradiation. No differences were observed in the repair capacity between hairy and hairless mice, thus eliminating defective DNA repair as the explanation for the greater susceptibility to UV carcinogenesis in hairless mice.
SITES OF PHOTODYNAMICALLY INDUCED DNA REPAIR IN HUMAN CELLS
Authors: Kvam Egil , Stokke Trond
Published: January 2008

Abstract Human REH cells were incubated with the photosensitizers meso‐tetra(4‐sulfonatophenyl)porphyrin (TSPP=TPPS4) or meso‐tetra(3‐hydroxyphenyl)porphyrin (3‐THPP). The relatively hydrophilic TSPP was partly found in the cytoplasm and partly in the nuclei, whereas the lipophilic 3‐THPP was found apparently in membranes and not inside the nuclei. After illumination, sites of DNA repair were labeled by means of a monoclonal antibody against proliferating cell nuclear antigen (PCNA) bound in the nuclei. The amount of bound PCNA in non‐S‐phase cells was proportional to the light dose. The bound PCNA was homogeneously distributed in the nuclei 0.5 h after photodynamic treatment (PDT) with TSPP. In contrast, for cells given PDT with 3‐THPP, the periphery of the nuclei was selectively labeled, indicating that the initial DNA damage was localized close to the sensitizer at the nuclear membrane.
VISIBLE LIGHT PHOTOCHEMICAL RELEASE OF NITRIC OXIDE FROM S‐NITROSOGLUTATHIONE: POTENTIAL PHOTOCHEMOTHERAPEUTIC APPLICATIONS
Authors: Sexton Daniel J. , Muruganandam Arumugam , McKenney Donald J. , Mutus Bulent
Published: January 2008

Abstract Some aspects of the physiological role of NO may be mediated by stable NO‐carriers such as S‐nitrosoglutathione and related S‐nitrosothiols. In this report we show that irradiation of S‐nitrosoglutathione at either absorption band (λmax= 340 nm or 545 nm) results in the release of nitric oxide. Photolysis of S‐nitrosoglutathione at 545 nm exhibited a quantum yield of 0.056 ± 0.002 and was best approximated by a first‐order process with kobs= 4.9 × 10−7± 0.3 × 10−7 s−1. The photolytic release of NO from S‐nitrosoglutathione resulted in an enhanced cytotoxic effect of S‐nitrosoglutathione on HL‐60 leukemia cells. That the cytotoxic effect of S‐nitrosoglutathione was diminished by the addition of oxyhemoglobin strongly suggests that NO is the cytotoxic species. The finding that NO can be readily liberated from S‐nitrosoglutathione by visible radiation indicates that the photochemical properties of this compound in the visible spectrum must be considered in order to obtain meaningful data as to its physiological role and the S‐nitrosoglutathione and related compounds may find use as photochemotherapeutic agents.
THE PRESENCE OF PHOTOREACTIVE β‐CARBOLINE FLUOROPHORES IN WEATHERED WOOL KERATIN
Authors: Smith Gerald J. , Markham Kenneth R. , Melhuish W. Hugh
Published: January 2008

Abstract Solvent‐cleaned Merino wool, which was enzymatically digested with protease K, displayed a fluorescence maximum at 500 nm when excited at 430 nm. The yield of this emission was approximately 15 times greater for a fiber tips digest than for a digest of the mid‐length region of the same fibers.

Separation of the components in the wool tip digest by silica gel thin‐layer chromatography revealed the presence of several fluorescent species. The chromatographic mobility of these species was similar to the behavior observed in a preparation of authentic 1‐methyltetrahydro‐P‐carboline, 1,3‐dicarboxylic acid.

Mass spectroscopy of the fluorescent compounds in the wool tip digest displayed molecular ions, [M + H]+ with m/z = 273.0890 and 257.0560, and molecular weights that define the formulae C14H12N2O4 and C11H8O4N2, respectively, which correspond to β‐carboline 1,3‐dicarboxylic acids. Mass spectral evidence also indicates the presence of two other β‐carbolines.
HYPOCRELLIN, FROM HYPOCRELLA BAMBUASE, IS PHOTOTOXIC TO HUMAN IMMUNODEFICIENCY VIRUS
Authors: Hudson J. B. , Zhou J. , Chen J. , Harris L. , Yip L. , Towers G. H. N.
Published: January 2008

Abstract Hypocrellin, a photodynamic perylene quinonoid isolated from the Chinese medicinal fungus Hypocrella bambuase, was evaluated for antiviral activity against the human immunodeficiency virus (HIV‐1). Hypocrellin was phototoxic to HIV‐1, almost as good as the structurally similar plant pigment hypericin, and like hypericin its activity required visible light. In contrast peroxyhypocrellin had little or no effect on the virus.
SUNSCREENS WITH BROAD‐SPECTRUM ABSORPTION DECREASE THE trans TO cis PHOTOISOMERIZATION OF UROCANIC ACID IN THE HUMAN stratum corneum AFTER MULTIPLE UV LIGHT EXPOSURES
Authors: Krien Paul M. , Moyal Dominique
Published: January 2008

Abstract The trans to cis photoisomerization of urocanic acid (UCA) in skin is considered to play an important role in the mechanism of immunosuppression. We have investigated the effects of skin type and various sunscreens with low sun protection factor (SPF) on the UV‐induced cis‐UCA formation in human skin after exposure to artificial IJV light. The rate of cis‐UCA formation depends little on the skin type and is reduced by topical application of sunscreens. The rate of cis‐UCA formation decreases with increasing SPF and only broad‐spectrum, highly protective sunscreens offer protection against the UV‐induced formation of cis‐UCA, which accumulates in the stratum corneum after multiple UV exposures. A theoretical approach to estimate the distribution of cis‐UCA after irradiation indicates that this compound may diffuse into the deeper layers of the epidermis with D ∼ 10−17 m2/s, and that its elimination from the stratum corneum is mainly due to desquamation.
PENETRATION OF SOLAR UV IRRADIATION IN COASTAL LAGOONS OF THE SOUTHERN BALTIC SEA AND ITS EFFECT ON PHYTOPLANKTON COMMUNITIES
Authors: Piazena H. , Häder D.‐P.
Published: January 2008

Abstract Direct measurements of solar UV irradiance at the surface and in the water column were made in several coastal lagoons near the islands of Hiddensee and Rügen in spring and summer 1993 using an internally frosted quartz sphere at the end of a 10 m quartz fiber cable connected to a double monochromator spectroradiometer. The measurements showed a nearly uniform vertical distribution of the absorption coefficient in the whole water column due to effective mixing, shallow depths and little temperature and salinity density stratifications. There was an increase in the absorption coefficient at 310 nm from 5 m−1 in the outer lagoons to 12 m−1 in the open inner lagoons and up to 29 m−1 in the inner lagoons without water exchange with the Baltic Sea as a result of different amounts of UV‐absorbing substances such as yellow substance (Gelbstoff) and seston. Depending on the specific concentration of the absorbents, the solar UV‐B irradiance decreases to 1% after penetration into the water column between 0.2 m (inner lagoons without water exchange), 0.4 m (inner lagoons with water exchange), and 1 m (outer lagoons), whereas UV‐A penetrates on the order of several meters. As phytoplankton populates the top layers of the water column it may be affected by increased solar UV irradiation even in these turbid coastal waters.
THEORETICAL STUDY OF COLOR CONTROL MECHANISM IN RETINAL PROTEINS. I. ROLE OF THE TRYPTOPHAN RESIDUE, TYROSINE RESIDUE and WATER MOLECULE
Authors: Beppu Yoshitaka , Kakitani Toshiaki
Published: January 2008

Abstract –We calculated the opsin shift due to the electrostatic interaction between tryptophan or tyrosine residues and the chromophore by the perturbation method for various mutual configurations. The obtained opsin shift maps for these configurations demonstrated that when the above residues reside around the ionone ring side, the positive opsin shift (bathochromic shift) is obtained, and when they reside around the Schiff‐base side, the negative opsin shift (hypsochromic shift) is obtained. These properties hold true, irrespective of the orientation of those residues, indicating that higher order multipoles of the group play a central role. The maximum value of the opsin shift by these groups amounts to several hundred wavenumbers. These results indicate that the location of some of those amino acid residues at proper positions around the chromophore can cause a considerable opsin shift. We also calculated opsin shift maps for the various mutual configurations between a water molecule and the chromophore for comparison.
ACTIVATION OF NF‐KB IN HUMAN SKIN FIBROBLASTS BY THE OXIDATIVE STRESS GENERATED BY UVA RADIATION
Authors: Vile Glenn F. , Tanew‐Iliitschew Adrian , Tyrrell Rex M.
Published: January 2008

Abstract—

We have examined the role of the nucleus and the membrane in the activation of nuclear factor (NF)‐KB by oxidant stress generated via the UVA (320–380nm) component of solar radiation. Nuclear extracts from human skin fibroblasts that had been irradiated with UVA at doses that caused little DNA damage contained activated NF‐KB that bound to its recognition sequence in DNA. The UVA radiation‐dependent activation of NF‐KB in enucleated cells confirmed that the nucleus was not involved. On the other hand, UVA radiation‐dependent activation of NF‐KB appeared to be correlated with membrane damage, and activation could be prevented by a‐tocopherol and butylated hydroxytol‐uene, agents that inhibited UVA radiation‐dependent peroxidation of cell membrane lipids. The activation of NF‐KB by the DNA damaging agents UVC (200–290nm) and UVB (290–320nm) radiation also only occurred at doses where significant membrane damage was induced, and, overall, activation was not correlated with the relative levels of DNA damage induced by UVC/UVB and UVA radiations. We conclude that the oxidative modification of membrane components may be an important factor to consider in the UV radiation‐dependent activation of NF‐KB over all wavelength ranges examined.
UVB‐INDUCED SUPPRESSION OF THE MIXED EPIDERMAL CELL LYMPHOCYTE REACTION IS CRITICALLY DEPENDENT ON IRRADIANCE
Authors: Hurks H. M. H. , Out‐Luiting C. , Vermeer B. J. , Claas F. H. J. , Mommaas A. M.
Published: January 2008

Abstract—

The mixed epidermal cell lymphocyte reaction (MECLR) is a commonly used method to study the effects of ultraviolet B (UVB) radiation on the skin immune system. In UVB experiments dosimetry is very important. The influence of irradiance on the MECLR was studied in vitro using Philips FS40 lamps with variable UV intensities. Irradiation of isolated epidermal cells with high irradiance impaired the alloactivating capacity more than irradiation with low irradiance. In vivo, the influence of long‐term UVB exposure on the MECLR was studied by treating normal healthy volunteers with suberythemagenic doses of UVB thrice weekly during 4 weeks. The first set of experiments, using low irradiance Sylvania UV‐21 F75/85 W lamps, resulted in a decrease of MECLR responses of 83.1%. In the second set of experiments performed a year later, employing an identical protocol except for the use of high irradiance Waldmann UV‐21 F85/100 W lamps, an increase of MECLR responses of 99.7% was observed. Volunteers of both sets of experiments received equal doses of UVB. In conclusion, this study shows that in vitro UVB‐induced suppression of the MECLR is critically dependent on irradiance and therefore might explain contradictory results described in the literature. The in vivo data suggest that, comparable to the in vitro experiments, irradiance may influence the effects of UVB irradiation in vivo. Further experiments should prove whether this is indeed the case.
PHOTOCHEMISTRY OF 2‐MERCAPTOPYRIDINES. PART 3. EPR STUDY OF PHOTOPRODUCTION OF HYDROXYL RADICALS BY N‐HYDROXYPYRIDINE‐2‐THIONE USING 5,5‐DIMETHYL‐1‐PYRROLINE N‐OXIDE IN AQUEOUS SOLUTIONS*
Authors: Reszka Krzysztof J. , Chignell Colin F.
Published: January 2008

Abstract— N‐Hydroxypyridine‐2‐thione, 2‐S‐PyrNOH, a potent antimicrobial, antifungal and anticancer agent, is photochemically active and upon UV irradiation generates free radicals. We have employed EPR and the spin‐trap 5,5‐dimethyl‐l‐pyrroline TV‐oxide (DMPO) to investigate the photochemistry in aqueous solutions of 2‐S‐PyrNOH (used here in the form of a sodium salt, 2‐S‐PyrNONa). We found that upon photoactivation 2‐S‐PyrNONa can follow two different pathways: it can produce hydroxyl radicals and/or it can act as a photoreducing agent. The capacity of 2‐S‐PyrNONa to produce “OH” radicals has been demonstrated by: (1) EPR detection of the DMPO/OH adduct in UV‐irradiated samples; (2) inhibition of the DMPO/OH formation by OH scavengers such as methyl alcohol, formate and DMSO and (3) by detection of EPR signals of DMPO adducts with radicals derived from reaction of OH with these inhibitors. The photoreductive capacity of 2‐S‐PyrNONa was deduced from the observation that the amplitude of the EPR signal of the spin adduct DMPO/OH decreased on UV irradiation in air‐free pH 7.0 buffers and that the signal recovered in the dark and after aeration. The ability to generate free radicals upon UV irradiation suggests that 2‐S‐PyrNONa can be regarded as a potential photocytotoxic agent. This feature may be relevant to the biological action of this compound. Our findings also emphasize that caution should be used when 2‐S‐PyrNOH is employed as a source of OH radicals in biological or chemical systems.
TWO GENES CONTRIBUTE TO DIFFERENT EXTENTS TO THE HEME OXYGENASE ENZYME ACTIVITY MEASURED IN CULTURED HUMAN SKIN FIBROBLASTS AND KERATINOCYTES: IMPLICATIONS FOR PROTECTION AGAINST OXIDANT STRESS
Authors: Applegate Lee Ann , Noël Alexandre , Vile Glenn , Frenk Edgar , Tyrrell Rex M.
Published: January 2008

Abstract— Activation of expression of the heme oxygenase (HO) gene appears to be involved in a cellular defense system in mammalian cells. We now demonstrate that while HO‐1 mRNA levels are strongly inducible in dermal fibroblasts they are barely inducible in human epidermal keratinocytes following oxidative stress (UVA radiation and hydrogen peroxide). Paralleling this result was the observation that HO‐2 mRNA levels were low in dermal fibroblasts but were high in epidermal keratinocytes. In neither case was the HO‐2 gene inducible. The expression of the two HO genes led to enzymatic activity in both types of skin cells with an approximately 2.5‐fold higher level of enzymatic activity present in keratinocytes compared with fibroblasts derived from the same biopsy. In addition, ferritin levels, which have been found to be augmented via the HO‐dependent release of iron from endogenous heme sources, were two‐ to three‐fold higher in keratinocytes compared with matching fibroblasts. This higher ferritin pool would result in an enhancement of cellular iron sequestering capacity that may confer increased resistance to oxidative stress. Indeed, keratinocytes showed less UVA radiation‐dependent cell membrane damage than fibroblasts. These results are consistent with the hypothesis that HO expression in human epidermis and dermis is related to cellular defense mechanisms that operate in human skin.
Cis‐UROCANIC ACID SYNERGIZES WITH HISTAMINE FOR INCREASED PGE2 PRODUCTION BY HUMAN KERATINOCYTES: LINK TO INDOMETHACIN‐INHIBITABLE UVB‐INDUCED IMMUNOSUPPRESSION
Authors: Jaksic Aleksandra , Finlay‐Jones John J. , Watson Christopher J. , Spencer Lyn K. , Santucci Irma , Hart Prue H.
Published: January 2008

Abstract— There is considerable evidence that suppression of the immune system by UVB (280–320 nm UV) irradiation is initiated by UVB‐dependent isomerization of a specific skin photoreceptor, urocanic acid (UCA), from the trans to the cis form. Previous studies have confirmed that cis‐UCA administration to mice 3–5 days prior to hapten sensitization at a distant site, suppresses the contact hypersensitivity (CHS) response upon challenge. This study demonstrates in mice that cis‐UCA, like UVB, suppresses CHS to trinitrochlorobenzene by a mechanism partly dependent on prostanoid production. In vitro experimentation showed that human keratinocytes, isolated from neonatal foreskin, increased prostaglandin E2 (PGE2) production in response to histamine but not UCA alone. However, cis‐UCA synergized with histamine for increased PGE2 production by keratinocytes. cis‐urocanic acid also increased the sensitivity of keratinocytes for PGE2 production in response to histamine. Prostaglandin E2 from keratinocytes exposed to cis‐UCA and histamine may contribute directly, or indirectly, to the regulation of CHS responses by UVB irradiation.
ADDITIONAL EVIDENCE FOR BLEPHARISMIN PHOTORECEPTOR PIGMENT MEDIATING STEP‐UP PHOTOPHOBIC RESPONSE OF UNICELLULAR ORGANISM, Blepharisma
Authors: Matsuoka Tatsuomi , Watanabe Yoshiya , Sagara Yasuhiro , Takayanagi Miyuki , Kato Yoji
Published: January 2008

Abstract— In the ciliated protozoan, Blepharisma japonicum, the pink‐colored pigment (blepharismin) contained in the pigment granules is believed to be the photoreceptor pigment responsible for the step‐up photophobic response. When the cells partially bleached by extrusion of the pigment granules caused by cold shocks were subsequently cultured under illuminated conditions, the pigment‐less granules regenerated and the cells were further bleached (pigment content below 0.5%). The photosensitivity of such colorless cells disappeared completely. In contrast, the blepharismin pigment regenerated gradually when such colorless cells were transferred to darkness. The photosensitivity of the cells also recovered with regeneration of the pigment. We found that blepharismin pigment was not photobleached in the absence of O2. The step‐up photophobic response was also completely repressed in the absence of O2. These results strongly confirm that blepharismin is a photoreceptor pigment mediating photobehavior of Blepharisma and that O2 is required for the early step in the phototransduction of the light‐excited pigment.
FORMATION OF 7,8‐DIHYDRO‐8‐OXOGUANINE IN THE 1,2‐DIOXETANE‐INDUCED OXIDATION OF CALF THYMUS DNA: EVIDENCE FOR PHOTOSENSITIZED DNA DAMAGE BY THERMALLY GENERATED TRIPLET KETONES IN THE DARK
Authors: Adam Waldemar , Saha‐Möller Chantu R. , Schönberger André , Berger Maurice , Cadet Jean
Published: January 2008

Abstract— Isolated calf thymus DNA was treated with the 1,2‐dioxetanes 3‐acetoxymethyl‐3,4,4‐tri‐methyl‐1,2‐dioxetane, 2,3‐dimethylbenzofuran dioxetane, 3‐hydroxymethyl‐3,4,4‐trimethyl‐1,2‐dioxeta‐ne (HTMD), 3,3,4,4‐tetramethyl‐1,2‐dioxetane and 3,4,4‐trimethyl‐1,2‐dioxetane (TrMD), which on thermal decomposition generate triplet‐excited carbonyl products. To monitor quantitatively the formation of the mutagenic oxidation product 7,8‐dihydro‐8‐oxoguanine (8‐oxoGua), a sensitive and selective HPLC electrochemical assay was used after acidic hydrolysis (HF/pyridine) of the dioxetane‐treated DNA. High yields of 8‐oxoGua (up to ca 4% of the available guanine) were obtained for HTMD and TrMD. Both were investigated in detail with respect to effects of concentration, time and temperature. The oxidative reactivity of 1,2‐dioxetanes was compared with several type I (benzophenone and riboflavin) and type II (methylene blue and rose bengal) photooxidants and disodium 1,4‐etheno‐2,3‐ben‐zodioxin‐1,4‐dipropionate as a chemical source of singlet oxygen. The persistence of 8‐oxoGua towards oxidation by HTMD was examined in the reaction with 7,8‐dihydro‐8‐oxo‐2'‐deoxyguanosine (8‐oxodGuo) and with oxidized DNA. It was shown that, indeed, 8‐oxoGua is consumed in the oxidized DNA on prolonged exposure to an excess of HTMD. The reaction of 8‐oxodGuo with HTMD afforded the two 4R* and 4S* diastereomers of 9‐(2‐deoxy‐ß‐D‐erythropentofuranosyl)‐4,8‐dihydro‐4‐hydroxy‐8‐oxoguanine as main oxidation products. Trapping experiments with teft‐butanol confirmed that hydroxyl radicals are not involved, whereas the use of the triplet quenchers sodium 9,10‐dibromo‐anthra‐cene‐2‐sulfonate and 2,3‐diazabicyclo[2.2.1]hept‐2‐ene established that triplet‐excited states are mainly responsible for the observed DNA oxidation through type I action (electron transfer chemistry). The role of singlet oxygen was tested by means of deuterium isotope effects in D2O versus H2O, but no definitive conclusion could be reached in regard to the involvement of 102 in these oxidations. The present results reveal that 1,2‐dioxetanes are efficient DNA oxidants and excellent tools to study photooxidation reactions of DNA in the dark.
SPONTANEOUS RECOVERY OF FLUORESCENCE BY PHOTOBLEACHED SURFACE‐ADSORBED PROTEINS
Authors: Stout Andrea L. , Axelrod Daniel
Published: January 2008

Abstract‐Fluorescence photobleaching of a carboxyfiuorescein‐labeled protein (erythrocyte cytoskel‐etal protein 4.1) immobilized on bare glass is found to be spontaneously reversible, provided that the sample is deoxygenated. After a short (hundredths of seconds) photobleaching laser flash, the subsequent fluorescence excited by a dim probe beam partly recovers on a long (tenths of second) time scale, even in the absence of chemical exchange or diffusion processes. Neither the fraction of the fluorescence that bleaches reversibly nor its recovery rate is a strong function of fluorophore surface concentration. At a fixed surface concentration, the reversibly photobleached fraction and its recovery rate decreases with increasing duration or intensity of the bleaching flash. On the other hand, nondeoxygenated air‐equilibrated samples exhibit almost total irreversible bleaching on this time scale. Quantitative fluorescence microscopy experiments occasionally require deoxygenation to avoid photochemical crosslinking or photobleaching or to enhance the triplet state population. The observations presented here indicate that fluorescence recovery after photobleaching (FRAP) experiments performed under deoxygenated conditions for measuring diffusion or chemical kinetics should be interpreted with caution: fluorescence recoveries may be due to intrinsic photochemical processes rather than fluorophore mobility. The recovery effect appears too slow to be ascribed simply to a relaxation of a triplet state; other possible explanations are offered.
PHOTOIONIZATION OF CRYSTAL VIOLET IN AQUEOUS SOLUTION
Authors: Bhasikuttan A. C. , Sapre A. V. , Rao K. V. S. Rama , Mittal J. P.
Published: January 2008

Abstract— Laser flash photolysis studies were carried out on a triphenylmethane dye, crystal violet (CV+), at 248 nm in aqueous solutions. The results show that CV+ undergoes photoionization and the resulting transients CV‐2+, hydrated electrons (e‐aq) and CV+ radical formed by the reaction of e‐aq with CV+ have been characterized. Studies using suitable scavengers were done to support the characterization of the transient species. Laser intensity effects show that the ionization is biphotonic. Two mechanisms are proposed to explain the observed photoionization involving higher excited singlet state and/ or another long‐lived excited state of the dye.
ALTERATIONS IN ERYTHROCYTE BAND 3 ORGANIZATION INDUCED BY THE PHOTOSENSITIZER, HEMATOPORPHYRIN DERIVATIVE
Authors: Beaton S. , McPherson R. A. , Tilley L.
Published: January 2008

Abstract— Photosensitization of erythrocytes in the presence of hematoporphyrin derivative causes cross‐linking of membrane proteins. This cross‐linking is associated with partial lysis of the cells and an increased susceptibility to heat‐induced membrane fragmentation. The effect of photosensitization on the organization of erythrocyte band 3 was monitored using the technique of time‐resolved phosphorescence anisotropy. Band 3 rotational diffusion was somewhat restricted upon photooxidation, indicating aggregation of this major integral membrane protein.
A METHOD FOR THE ELECTROCHEMICAL INITIATION OF THE in vitro BACTERIAL LUCIFERASE REACTION
Authors: Karatani Hajime , Shizuki Takahiro , Halon Rozana , Nakayama Eiichiro
Published: January 2008

Abstract— We have developed an electrochemical initiation method for the in vitro bacterial luciferase reaction using Vibrio harveyi luciferase. The cathodic conversion of oxidized riboflavin 5′‐phosphate (FMN) to its reduced form (FMNH2), on which the kinetics for the overall light‐emitting reaction depend, is followed by the bacterial luciferase reaction. The electrochemical initiation method can be used for both turnover and nonturnover luciferase reactions, depending on the FMN electroreduction methods.
Transcutaneous Photodynamic Therapy Alters the Development of an Adoptively Transferred Form of Murine Experimental Autoimmune Encephalomyelitis
Authors: Leong Simon , Chan Agnes H. , Levy Julia G. , Hunt David W. C.
Published: January 2008

Abstract— Transcutaneous photodynamic therapy (PDT), utilizing benzoporphyrin derivative monoacid ring A (BPD, ver‐teporfin) and whole‐body light exposure, was assessed for its capacity to modify the course of adoptively transferred experimental autoimmune encephalomyelitis (EAE) in PL mice. Using a novel cell culture technique to facilitate the induction of this neurodegenerative condition, disease signs commenced 3–4 weeks after the transfer of myelin basic protein (MBP)‐reactive lymph node or spleen cells to naive syngeneic recipients. Mice administered MBP‐sensitized lymph node cells preincu‐bated with BPD followed by whole‐body 690 nm light irradiation (15 J/cm2) did not display symptoms of EAE. Although almost all animals given MBP‐sensitized spleen cells developed EAE, mice given BPD (1 mg/kg) and the light treatment 24, 48 or 120 h after spleen cell transfer exhibited significantly less severe disease symptoms than control animals. Mice given the photodynamic treatment 24 h after spleen cell transfer also exhibited a significantly later disease onset than the control animals. Treatment of mice with PDT 24 h prior to spleen cell transfer did not influence subsequent disease severity but modestly delayed its onset. In the absence of directed light, BPD did not influence the development of EAE. Spinal cord tissues were evaluated for the presence of T cell receptor (TCR) Vα4 mRNA transcripts that specifically encode for the TCR ot‐chain of MBP‐reactive T cells of PL mice. Using the polymerase chain reaction, Va4 TCR mRNA transcripts were present in spinal cord samples prepared from almost all control mice but in only about one‐half of spinal cord samples prepared from mice treated with PDT 24 h after spleen cell transfer. These observations indicated that PDT had limited the expansion of MBP‐specific Vot4+ T cells within the central nervous system. Transcutaneous PDT represents a new technique with which to approach the treatment of autoimmune disease.
A Negative Regulatory Factor for the Dark Repression of Arabidopsis thaliana cab1 Gene*
Authors: Choi Dong‐Kug , Park Hee‐Jin , Kim In‐Soo
Published: January 2008

Abstract— A protein factor and its binding site involved in light‐responsive gene expression of Arabidopsis thaliana cabl were investigated. Mobility shift assays were performed to identify a nuclear protein factor and its binding sites on the cab1 promoter. For the binding assay, the Arabidopsis cab1 promoter was cleaved with endonucleases into small fragments (65‐200 bp) and end‐labeled with Klenow fragments. Nuclei were prepared from the light‐grown plants and nuclear proteins were prepared by extracting the purified nuclei with 0.5 M ammonium sulfate. The binding site of the nuclear protein factor was scattered throughout the whole promoter region from the transcription start site to the far upstream region of the promoter. To identify the binding sites that are involved in the light responsiveness, mobility shift assays were performed between the cab1 promoter fragments and the nuclear extracts prepared from the 2 day dark‐adapted sample. The mobility shift assay of the 65 bp (‐318/ ‐254) fragment with nuclear extract from the dark‐adapted sample showed an additional band, not seen with the light‐grown sample. Because the new band was present only in the dark‐adapted sample that repressed cabl expression, it may represent a negative regulatory factor (NRF). The NRF was separable on a heparin‐Sepharose column from the other factor present in both the light‐grown and dark‐adapted samples. The implications of the presence of the NRF have been discussed with respect to gene products of the photosignal transduction Arabidopsis mutants.
Solar‐simulated Ultraviolet Irradiation Induces Selective Influx of CD4+ T Lymphocytes in Normal Human Skin
Authors: Nuzzo Sergio Di , de Rie Menno A. , van der Loos Chris M. , Bos Jan D. , Teunissen Marcel B. M.
Published: January 2008

Abstract— The proportion and composition of the human cutaneous CD3+ T lymphocyte population was determined in situ following a single exposure to physiological, erythema‐inducing doses of simulated solar radiation, mainly consisting of UV radiation. Biopsies were taken 1, 2 and 7 days after local irradiation of normal volunteers with 1,2 and 4 MED by a xenonarc lamp and immunohistochemistry was performed on cryostat sections. Ultraviolet radiation caused an initial decrease of intraepidermal CD3+ T‐cell numbers or even could lead to T‐cell depletion 24 and 48 h postirradiation, and this was followed by an infiltration of T cells in the epidermis as determined 1 week after UV exposure. The number of dermal CD3+ T ceDs was increased 24 h after irradiation, reached a maximum at 48 h and subsequently declined at day 7, though remained significantly higher than the unirradiated control Double staining demonstrated that the CD3+ T cells, which immigrated into the (epi)dermis upon UV exposure, coexpressed CD4 but not CD8. Therefore the CD4/CD8 ratio in skin was markedly increased during the first week upon UV exposure. Our time course study shows that UV radiation affects die T‐cell population within human skin by depleting the majority of epidermal T cells and initiating a selective influx of CD4+ T cells.
The Influence of Photosynthetically Active Radiation on the Effects of Ultraviolet‐B Radiation on Arabidopsis thaliana
Authors: Berkelaar Edward J. , Ormrod Douglas P. , Hale Beverley A.
Published: January 2008

Ultraviolet‐B (UVB;280–320 nm) radiation is a small but biologically significant portion of the solar spectrum reaching the earth's surface. Research interests have been fostered because UVB has been increasing in recent years due to depletion of stratospheric ozone. Ultraviolet‐B that penetrates into plant tissue may damage important cellular macromolecules. Although there has been considerable research on the effects of UVB on plants, the influence of the level of photosynthetically active radiation (PAR;400–700 nm) on effects of UVB requires further definition as a prelude to studies of UVB sensitivity and defense mechanisms. Arabidopsis thaliana wildtype ecotype Landsberg erecta (LER), which is relatively insensitive to UVB, and the relatively sensitive LER‐based mutant transparent testa‐5 (tt5), were grown under 100 or 250 μmol m−2 s−1 PAR and then exposed to O or 7 kJ m−2 day −1 UVBBE under these PAR levels. Plants exposed to UVB had reduced dry weight and leaf area and higher levels of UV‐absorbing compounds in leaf tissue. The level of PAR did influence the effects of UVB, with the higher level of PAR prior to UVB exposure reducing sensitivity of LER to UVB. In contrast to other studies, higher PAR supplied simultaneously with UVB increased rather than decreased sensitivity of both genotypes to UVB. These results demonstrate the importance of controlling and comparing PAR levels when undertaking studies of UVB sensitivity, as effects of UVB on plants are influenced by the PAR levels plants are growing under prior to and during exposure to UVB.
Role of Hydrogen Peroxide in the Cytotoxic Effects of UVA/B Radiation on Mammalian Cells
Authors: Bertling Chad J. , Lin Fubao , Girotti Albert W.
Published: January 2008

Effects of selenium (Se) deficiency on the sensitivity of murine leukemia L1210 cells to broad band UVA/B radiation (310–400nm) have been investigated. Cells rendered glutathione peroxidase (GPX) deficient by shortterm (2–3week) growth in 1% serum/RPMI medium without added Se [LSe(‐) cells] were found to be much less resistant to clonally assessed UVA/B lethality than Se‐supplemented controls [LSe(+) cells]. By contrast, long‐term (>20 week) Se‐deprived [L'Se(‐)] cells whose catalase (CAT) activity was elevated >100‐fold were far more resistant to UVA/B than LSe(+) cells. Similar trends were observed for cells irradiated in 1% serum/RPMI or Hank's medium. Whereas the CAT inhibitor 3‐amino‐1,2,4‐triazole had no effect on LSe(+) photosensitivity, it produced a large increase in L' Se(‐) photosensitivity. These findings are consistent with H2O2 in‐termediacy in photokilling and suggest that L1210 cells depend mainly on GPX for protection against this species but switch to overexpressed CAT after chronic Se deprivation. In agreement with this, steady‐state H2O2 levels measured by H2O2 electrode during UVA/B exposure were higher in LSe(‐) than LSe(+) suspensions but much lower (barely detectable) in L' Se(‐) suspensions. Cytotoxic effects of UVA/B and variations thereof resulting from Se manipulation could be mimicked by treating cells with glucose oxidase in the presence of D‐glucose, providing further support for H2O2 involvement. Whether UVA/B‐generated H2O2 is directly cytotoxic or gives rise to a more damaging species such as hydroxyl radical (HO) is presently unknown.