bluuebunny/crossref_metadata_2025 huggingface dataset:
Identification of Genes Responsive to UV‐A Radiation in Human Lens Epithelial Cells Using Complementary DNA Microarrays¶
Authors: Andley Usha P. , Patel Harendra C. , XI Jing‐Hua , Bai Fang
Published: July 2010

UV‐A radiation produces cataract in animals, enhances photoaging of the lens and skin and increases the phototoxicity of drugs. However, the nature of genes that are activated or repressed after cellular exposure to UV‐A radiation remains enigmatic. Because lens epithelial cells exposed to UV‐A radiation undergo apoptosis 4 h after exposure to the stress, we sought to establish the change in gene expression in cells by UV‐A radiation using gene expression profiling using complementary DNA microarrays containing about 12000 genes. We identified 78 genes abnormally expressed in UV‐A‐irradiated cells (showing > 2.5‐fold change at P < 0.05). These genes are implicated in various biological processes, including signal transduction and nucleic acid binding, and genes encoding enzymes. A majority of the genes were downregulated. Our analysis revealed that the expression of genes for the transcription factors ATF‐3 and Pilot increased four‐fold, whereas the gene for the apoptosis regulator NAPOR‐1 decreased five‐fold. These changes were confirmed by real‐time quantitative reverse transcriptase‐polymerase chain reaction. The calpain large polypeptide 3 (CANP3) gene also increased nine‐fold after UV‐A radiation. In addition, peroxisomal biogenesis factor 7, glucocorticoid receptor‐α and tumor‐associated calcium signal transducer genes decreased three‐ to eight‐fold. Western blot analysis further confirmed the increase in protein expression of ATF‐3 and CANP3 and decreased expression of glucocorticoid receptor‐α in the irradiated cells. Surprisingly, most of these genes had not been previously shown to be modulated by UV‐A radiation. Our results show that human lens epithelial cells respond to a single dose of UV‐A radiation by enhancing or suppressing functionally similar sets of genes, some of which have opposing functions, around the time at which apoptosis occurs. These studies support the intriguing concept that activation of competing pathways favoring either cell survival or death is a means to coordinate the response of cells to UV‐A stress.
Fluorescence Lifetime Spectroscopy of Glioblastoma Multiforme¶
Authors: Marcu Laura , Jo Javier A. , Butte Pramod V. , Yong William H. , Pikul Brian K. , Black Keith L. , Thompson Reid C.
Published: July 2010

Fluorescence spectroscopy of the endogenous emission of brain tumors has been researched as a potentially important method for the intraoperative localization of brain tumor margins. We investigated the use of time‐resolved, laser‐induced fluorescence spectroscopy for demarcation of primary brain tumors by studying the time‐resolved spectra of gliomas. The fluorescence of human brain samples (glioblastoma multiforme, cortex and white matter: six patients, 23 sites) was induced ex vivo with a pulsed nitrogen laser (337 nm, 3 ns). The time‐resolved spectra were detected in a 360–550 nm wavelength range using a fast digitizer and gated detection. Parameters derived from both the spectral‐ (intensities from narrow spectral bands) and the time domain (average lifetime) measured at 390 and 460 nm were used for tissue characterization. We determined that high‐grade gliomas are characterized by fluorescence lifetimes that varied with the emission wavelength (>3 ns at 390 nm, <1 ns at 460 nm) and their emission is overall longer than that of normal brain tissue. Our study demonstrates that the use of fluorescence lifetime not only improves the specificity of fluorescence measurements but also allows a more robust evaluation of data collected from brain tissue. Combined information from both the spectraland the time domain can enhance the ability of fluorescencebased techniques to diagnose and detect brain tumor margins intraoperatively.
Laser Techniques in the Study of Drug Photochemistry¶
Authors: Cosa Gonzalo , Scalano J. C.
Published: July 2010

This review describes the application of different laser time‐resolved techniques, such as time‐resolved fluorescence, phosphorescence and laser flash photolysis, to elucidate the mechanism of drug photodegradation. The assignment of transient species based on their luminescence or absorption spectra and based on their reactivity to various quenchers is illustrated. The mathematical expressions used in fitting measured transient decays over time are also discussed. Practical situations found in the literature and relevant to drug photodegradation illustrate different examples. The importance of laser power, concentration and absorbance of the ground state, as well as other parameters intrinsic to laser techniques are discussed.
Phtodeceomposition of Pigment Yellow 74, a Pigment Used in Tattoo Inks¶
Authors: Cul Yanyan , Spann Andrew P. , Couch Letha H. , Gopee Neera V. , Evans Frederick E. , Churchwell Maona I. , Williams Lee D. , Doerge Daniel R. , Howard Paul C.
Published: July 2010

Tattooing has become a popular recreational practice among younger adults over the past decade. Although some of the pigments used in tattooing have been described, very little is known concerning the toxicology, phototoxicology or photochemistry of these pigments. Seven yellow tattoo inks were obtained from commercial sources and their pigments extracted, identified and quantitatively analyzed. The monoazo compound Pigment Yellow 74 (PY74; CI 11741) was found to be the major pigment in several of the tattoo inks. Solutions of commercial PY74 in tetrahydrofuran (THF) were deoxygenated using argon gas, and the photochemical reaction products were determined after exposure to simulated solar light generated by a filtered 6.5 kW xenon arc lamp. Spectrophotometric and high‐pressure liquid chromatography (HPLC) analyses indicated that PY74 photodecomposed to multiple products that were isolated using a combination of silica chromatography and reversed‐phase HPLC. Three of the major photodecomposition products were identified by nuclear magnetic resonance and mass spectrometry as N(2‐methoxyphenyl)‐3‐oxobutanamide (o‐acetoacetanisidide), 2‐(hydroxyimine)‐N‐(2‐methoxyphenyl)‐3‐oxobutanamide and N,N″‐bis(2‐methoxyphenyl)urea. These results demonstrate that PY74 is not photostable in THF and that photochemical lysis occurs at several sites in PY74 including the hydrazone and amide groups. The data also suggest that the use of PY74 in tattoo inks could potentially result in the formation of photolysis products, resulting in toxicity at the tattoo site after irradiation with sunlight or more intense light sources.
Molecular Mechanism(s) for UV‐B Irradiation‐Induced Glutathione Depletion in Cultured Human Keratinocytes¶
Authors: Zhu Ming , Bowden G. Tim
Published: July 2010

Glutathione (GSH) plays a centxral role in maintenance of cellular redox homeostasis and protection against oxidative injury. Ultraviolet B (UV‐B) irradiation‐induced GSH depletion is believed to be involved in the pathogenesis of several cutaneous disorders. In this study, the molecular mechanism(s) of UV‐B‐induced GSH depletion was investigated in cultured human keratinocytes, HaCaT cells. We found that UV‐B irradiation caused GSH depletion in a dose‐ and timedependent manner in HaCaT cells. The mechanistic studies showed that UV‐B‐induced GSH depletion did not result from the GSH efflux. UV‐B irradiation appeared to cause a slight decrease in enzymatic activity of y‐glutamate cysteine ligase (GCL), a rate‐limiting enzyme in GSH biosynthesis. UV‐B irradiation resulted in the GCL cleavage through the activation of a caspase cascade. Inhibition of total caspase activity by the general caspase inhibitor, zVAD‐fmk, partially reversed the UV‐B‐induced GSH depletion. More importantly, we found that UV‐B irradiation could dramatically decrease the cystine uptake through the functional inhibition of the system Xc−, a cystine transporter on the cell membrane. The results suggest that the inactivation of cystine transporter system Xc− was a major contributor to the UV‐B‐mediated decrease of GSH levels in human keratinocytes.
Influence of Eicosapentaenoic Acid, an Omega‐3 Fatty Acid, on Ultraviolet‐B Generation of Prostaglandin‐E2 and Proinflammatory Cytokines Interleukin‐1β, Tumor Necrosis Factor‐α, Interleukin‐6 and Interleukin‐8 in Human Skin In Vivo¶
Authors: Shahbakhti Hassan , Watson Rachel E. B. , Azurdia Richard M. , Ferreira Christiana Z. , Garmyn Marjan , Rhodes Lesley E.
Published: July 2010

Dietary omega‐3 polyunsaturated fatty acids (ω‐3 PUFA) reduce sunburn, an acute inflammatory response, in humans, We assessed whether this may be mediated by reduced ultraviolet‐B (UV‐B) induction of proinflammatory mediators tumor necrosis factor‐α (TNF‐α), interleukin (IL)‐1β, IL‐6, IL‐8 and prostaglandin (PG)E2 in healthy skin. In a double‐blind, randomized study, 28 humans received 4 g daily of 95% ethyl esters of eicosapentaenoic acid (EPA) or oleic acid (OA) orally for 3 months. Skin biopsies and suction blister fluid were taken from unexposed and UV‐B‐exposed skin and examined for mediator expression immunohistochemically and quantitatively by immunoassay; plasma levels were also assayed. The subjects taking EPA, but not OA, showed a significant rise in their minimal erythemal dose (MED) (data reported elsewhere). Before supplementation, irradiation with 3X MED UV‐B increased blister fluid TNF‐α, IL‐6, IL‐8 and PGE2 at 16 h (all P < 0.001). No significant change occurred in baseline or UV‐B‐induced skin levels of cytokines after either supplement, whereas UV‐B induction of PGE2 was abolished after EPA but not OA. Immunohistochemical expression of the cytokines at baseline and after UV‐B was unaltered by EPA and OA; circulating cytokine and PGE2 levels were also unchanged. Hence, in healthy skin in vivo, there was no evidence that reduction of the sunburn response by EPA is mediated by the proinflammatory cytokines examined; abrogation of UV‐B‐generated PGE2, may play a role.
Comparative Sensitivity of Microvascular Endothelial Cells, Fibroblasts and Tumor Cells after In Vitro Photodynamic Therapy with meso‐Tetra‐Hydroxyphenyl‐Chlorin¶
Authors: Triesscheijn Martijn , Ruevekamp Marjan , Aalders Maurice , Baas Paul , Stewart Fiona A.
Published: July 2010

The phototoxic effect of meso‐tetra‐hydroxyphenyl‐chlorin (mTHPC)‐mediated photodynamic therapy (PDT) on human microvascular endothelial cells (hMVEC) was compared with that on human fibroblasts (BCT‐27) and two human tumor cell lines (HMESO‐1 and HNXOE). To examine the relationship between intrinsic phototoxicity and intracellular mTHPC content, we expressed cell survival as a function of cellular fluorescence. On the basis of total cell fluorescence, HNXOE tumor cells were the most sensitive and BCT‐27 fibroblasts the most resistant, but these differences disappeared after correcting for cell volume. Endothelial cells were not intrinsically more sensitive to mTHPC‐PDT than tumor cells or fibroblasts. Uptake of mTHPC in hMVEC increased linearly to at least 48 h, whereas drug uptake in the other cell lines reached a maximum by 24 h. No difference in drug uptake was seen between the cell lines during the first 24 h, but by 48 hMVEC had a 1.8‐ to 2.8‐ fold higher uptake than other cell lines. Endothelial cells showed a rapid apoptotic response after mTHPC‐mediated PDT, whereas similar protocols gave a delayed apoptotic or necrotic like response in HNXOE. We conclude that endothelial cells are not intrinsically more sensitive than other cell types to mTHPC‐mediated PDT but that continued drug uptake beyond 24 h may lead to higher intracellular drug levels and increased photosensitivity under certain conditions.
Energy Transfer and Fluorescence Quenching in Complexes of Polymethine Dyes with Human Serum Albumin¶
Authors: Tatikolov Alexander S. , Costa Sílvia M. B.
Published: July 2010

Electronic excitation energy transfer (EET) between molecules of polymethine dyes bound to human serum albumin (HSA) has been established and studied by absorption and fluorescence spectroscopy as well as by fluorescence decay measurements. In this system, excitation of the donor dye molecule leads to fluorescence of the acceptor dye molecule, both bound to HSA, with donor fluorescence quenching by the acceptor. The short distance between the donor and the acceptor (25‐28 Å) revealed from the Förster model of EET as well as some spectroscopic data show that both molecules are probably located in the same binding domain of HSA. The role of HSA is to bring donor and acceptor molecules together to a distance adequate to achieve EET as well as to increase the donor and acceptor fluorescence quantum yields. Efficient quenching of the intrinsic HSA fluorescence by some polymethine dyes (oxonols) is observed. The experimental results fit well a model for the formation of a weakly fluorescent dye‐HSA complex; the quencher in this complex should be located in the immediate vicinity of the HSA fluorophore group (Trp214).
The Effect of Molecular Environment on the Photoisomerization of Urocanic Acid¶
Authors: Wallis Richard A. , Smith Gerald J. , Dunford Cara L.
Published: July 2010

Urocanic acid, imidazole propenoic acid, is a metabolic product of histidine, which accumulates in skin and is excreted in sweat. It absorbs UV radiation at wavelengths shorter than 340 nm, and its principal photochemical reaction is a trans‐cis isomerization about the propenyl double bond. This isomerization to the biologically active cis isomer is implicated in the photo‐induced suppression of the immune system of skin. The kinetics of the trans→cis photoisomerization of urocanic acid has been determined in a number of solvents, spanning a range of polarities. The initial rates of isomerization and the photostationary trans‐cis compositions, in all solvents except water, correlate linearly with solvent polarity. This indicates that the isomerization proceeds through a polar intermediate that is stabilized by coulombic interactions with the molecular environment.
Photoinactivation of Hepatitis A Virus by Synthetic Porphyrins¶
Authors: Casteel Michael J. , Jayaraj K. , Gold Avram , Ball Louise M. , Sobsey Mark D.
Published: July 2010

Porphyrins are photosensitizers and may be applicable in situations where viral inactivation is required, as for in vitro inactivation of nonenveloped viruses in blood components or in other aqueous media. No study has examined the efficacy of porphyrin inactivation on human pathogens such as hepatitis A virus (HAV) in plasma or other liquids. Experiments were conducted to evaluate the effect of synthetic porphyrins on HAV in porphyrin‐containing human plasma and phosphatebuffered saline exposed to long‐wavelength (365 nm) UV light. Inactivation of bacteriophage MS2 (MS2) also was determined in some trials. Solutions containing cationic, anionic or amphiphilic porphyrins irradiated with an average light dose of 4.3 J/cm2 for 90 min resulted in >3 log10 (>99.9%) to >4 log10 (>99.99%) inactivation of both HAV and MS2. Viral inactivation may have been greater than observed because the limits of detection of the assay had been reached. Under ambient lighting conditions, none of the porphyrins was mutagenic in the Ames assay and only the congener with the longest chain‐length, tetrakis (N‐[n‐hexadecyl]‐4‐pyridiniumyl) porphyrin, was appreciably toxic to mammalian cells. Disinfection by photoactivated synthetic porphyrins therefore can offer an effective and relatively safe approach to removal of nonenveloped viruses from aqueous media.
Ground‐ and Excited‐state Proton Transfer Reaction of 3‐Hydroxyflavone in Dimyristoylphosphatidylcholine Liposome Membrane¶
Authors: Shyamala Thirunavukkuarasu , Mishra Ashok K.
Published: July 2010

3‐Hydroxyflavone (3HF), a molecule that exhibits excited‐state intramolecular proton transfer, has been studied for its fluorescence characteristics in dimyristoylphosphatidylcholine (DMPC) liposome membrane. 3HF partitions to the lipid bilayer membrane with a reasonably large partition coefficient. On excitation at 417 nm, a weak emission from the ground‐state anion species was observed at 483 nm, whereas excitation at absorption maxima (345 nm) gives the usual intense fluorescence of the phototautomeric emission at 530 nm. In this article, we report the observation of a ground‐state proton transfer reaction of 3HF in DMPC liposome membrane.
UV Absorption and Photoisomerixation of p‐Methoxycinnamate Grafted Silicone¶
Authors: Pattanaargson Supason , Hongchinnagorn Nantawan , Hirunsupachot Piyawan , Sritana‐anant Yongsak
Published: July 2010

p‐Methoxycinnamate moieties, UV‐B‐absorptive chromophores of the widely used UV‐B filter, 2‐ethylhexyl p‐methoxycinnamate (OMC), were grafted onto the 7 mol% amino functionalized silicone polymer through amide linkages. Comparing with OMC, the resulting poly [3(p‐methoxycinnamido)(propyl)(methyl)‐dimethyl] siloxane copolymer (CAS) showed less E to Z isomerization when exposed to UV‐B light. The absorption profiles of the product showed the maximum absorption wavelength to be similar to that of OMC but with less sensitivity to the type of solvent. Poly (methylhydrosiloxane) grafted with 10 mol%p‐methoxycinnamoyl moieties was prepared through hydrosilylations of 2‐propenyl‐p‐methoxycinnamate, in which the resulting copolymer showed similar results to those of CAS.
Radical Cation Generation from Singlet and Triplet Excited States of All‐trans‐Lycopene in Chloroform¶
Authors: Han Rui‐Min , Wul Yi‐Shi , Feng Juan , Ai Xi‐Cheng , Zhang Jian‐Ping , Skibsted Leif H.
Published: July 2010

On direct photoexcitation, subpicosecond time‐resolved absorption spectroscopy revealed that the 1Bu‐type singlet excited state of all‐trans‐lycopene in chloroform was about seven times more efficient than all‐trans‐β‐carotene in generating the radical cation. The time constant of radical cation generation from the 1Bu‐type state was found to be ∼0.14 ps, a value that was comparable for the two carotenoids. On anthracene‐sensitized triplet excitation, radical cation generation was found to be much less efficient for lycopene than for β‐carotene. A slow rising phase (20‐30 μs) in the bleaching of ground‐state absorption was common for both lycopene and β‐carotene in chloroform and was ascribed to an efficient secondary reaction with a solvent radical leading to the formation of carotenoid radical cations. The reverse ordering in the tendency of the excited states of different multiplicities for the two carotenoids to generate radical cations is discussed in relation to the two carotenoids as scavengers of free radicals.
Spatial Distribution of Upper‐room Germicidal UV Radiation as Measured with Tubular Actinometry as Compared with Spherical Actinometry¶
Authors: Rahn Ronald O.
Published: July 2010

Chemical actinometry of UV germicidal irradiation using sections of quartz tubing as compared with quartz spheres as irradiation vessels has been investigated. Vessels were either 3 mm inner diameter quartz tubing, 46 cm in length (tubular actinometry), or 1 cm quartz spheres (spherical actinometry). The vessels containing an iodide/iodate actinometric solution were suspended from the ceiling at 24 positions in a room (6 × 6 m) containing five germicidal lamp fixtures in the corners and in the center of the room. The lamp fixtures were louvered collimating the radiation in the horizontal (x, y) plane. Hence, the tubes, which span the depth of the radiation field, essentially integrate the radiation along the z‐axis for a given x, y position. The pseudospatial average fluence rate obtained using tubular actinometry was 18 mW/cm2 for the volume contained in the upper 46 cm (18 inch) of the room. Spherical actinometry, which measured the fluence rate in the center of the beam, provided an average value of 32 mW/cm2 over the volume of the beam. A comparison of the fluence rates obtained by these two methods allowed the average depth of the beam to be estimated as 26 cm. It is concluded that tubular actinometry is more advantageous than spherical actinometry for this application.
Probing the Active Site of Trypsin with Rose Bengal: Insights into the Photodynamic Inactivation of the Enzyme¶
Authors: Khajehpour Mazdak , Troxler Thomas , Vanderkool Jane M.
Published: July 2010

In this work the active site of trypsin has been probed with the dye rose bengal. The dye binds competitively to the enzyme, and it can be used as a probe of the active site of the enzyme. On the basis of the emission wavelength, the binding site of trypsin is relatively polar and is similar to that of acetone in its polarity. The triplet state of rose bengal is quenched by trypsin. This quenching may be caused by the tryptophan and tyrosine residues that are in the near vicinity of the trypsin active site. This quenching can compete with the formation of singlet oxygen from the excited triplet state of rose bengal. We demonstrate that the singlet oxygen involved in the photoinactivation of trypsin is produced by the free rose bengal in solution and the bound dye is incapable of producing singlet oxygen. This explains the lack of correlation between photoinactivation efficiency and sensitizer binding capability previously reported by Wade and Spikes.
The Phase‐dependent Photochemical Reaction Dynamics of Halooxides and Nitrosyl Halides¶
Authors: Cooksey Catherine C. , Reid Philip J.
Published: July 2010

Recent progress in understanding the phase‐dependent reactivity of halooxides and nitrosyl halides is outlined. Halooxide reactivity is represented by the photochemistry of chlorine dioxide (OCIO) and dichlorine monoxide (CIOCI). The gas phase photochemical dynamics of OCIO are contrasted with the dynamics in condensed environments. The role of excited‐state symmetry in defining the reaction dynamics and the observation of photoisomerization resulting in the production of CIOO are discussed. The current understanding of the excited‐state reaction dynamics of CIOCI and evidence for photoisomerization of this species resulting in the production of CICIO are outlined. Finally, the photochemical reaction dynamics of the nitrosyl halide CINO are presented. The main difference between the gas and condensed phase reaction dynamics of this species is that whereas photodissociation to form CI and NO dominates the gas phase reaction dynamics, photoisomerization resulting in CION production occurs to an appreciable extent in condensed environments. The observation of photoisomerization for OCIO, CIOCI and CINO suggests that this process is a general feature of the condensed phase reaction dynamics for smaller halooxides and nitrosyl halides. Finally, future areas for study in both halooxide and nitrosyl halide photoreactivity are outlined.
Lens α‐Crystallin and Hypericin: A Photophysical Mechanism Explains Observed Lens Damage¶†
Authors: Trevithick‐Sutton Colleen C. , Chin Khin K. , Contos Sotiria D. , Foote Christopher S.
Published: July 2010

Determining whether α‐crystallin (the major lens protein) affects the photophysics of hypericin, a photosensitizing agent found in various plants, such as St. John's Wort, is important. Hypericin shows promise in cancer and human immunodeficiency virus therapy but may harm individuals taking St. John's Wort extracts (for mild to moderate depression). Hypericin causes hypericism, which is characterized by cellular damage in light‐exposed areas. Ocular tissues are at risk for photosensitized damage; thus, we investigated the effects on hypericin photophysics by α‐crystallin. We measured the transient absorption spectra and the 1270 nm luminescence of singlet (1Δg) oxygen produced from hypericin in the presence of α‐crystallin. α‐Crystallin complexes hypericin, extending the lifetime of its triplet excited state; the Stern‐Volmer slope is negative, but not linear, after a saturation curve. Damage to the lens protein by hypericin is known to occur via singlet oxygen, which oxidizes methionine, tryptophan and histidine residues. Binding to α‐crystallin does not inhibit singlet oxygen formation by hypericin. α‐Crystallin reacts with singlet oxygen with a rate constant of 1.3 × 108M−1 s−1. Thus, we anticipate that hypericin will be an effective photosensitizer in the lens.
Binding of Metal Ions to Melanin and Their Effects on the Aerobic Reactivity¶
Authors: Hong Lian , Liu Yan , Simon John D.
Published: July 2010

The binding of Mg(II), Ca(II), Zn(II), Cu(II) and Fe(III) to ETDA‐washed Sepia melanin is quantified by inductively coupled plasma mass spectrometry. By monitoring the solution pH change associated with metal uptake, it is concluded that Mg(II), Ca(II) and Zn(II) bind to carboxylic acid groups in melanin, Cu(II) binds to hydroxyl (OH) groups and Fe(III) binds to OH or amine groups. The aerobic reactivity of melanins with different metal contents is analyzed by examining their ability to cause strand breaks in super‐coiled pUC18 DNA. Cu(II)‐ and Fe(III)‐enriched melanins induce the most damage. Hydroxyl radical, *OH, is proposed to be one of the reactive oxygen species responsible.
Phototoxicity in Human Lens Epithelial Cells Promoted by St. John's Wort¶
Authors: He Yu‐Ying , Chignell Colin F. , Miller David S. , Andley Usha P. , Roberts Joan E.
Published: July 2010

St. John's Wort (SJW), an over‐the‐counter antidepressant, contains hypericin, which absorbs light in the UV and visible ranges and is phototoxic to skin. To determine if it also could be phototoxic to the eye, we exposed human lens epithelial cells to 0.1–10 μM hypericin and irradiated them with 4 J/cm2 UV‐A or 0.9 J/cm2 visible light. Neither hypericin exposure alone nor light exposure alone reduced cell viability. In contrast, cells exposed to hypericin in combination with UV‐A or visible light underwent necrosis and apoptosis. The ocular antioxidants lutein and N‐acetyl cysteine did not prevent damage. Thus, ingested SJW is potentially phototoxic to the eye and could contribute to early cataractogenesis. Precautions should be taken to protect the eye from intense sunlight while taking SJW.
Effects of Ultraviolet B Exposure on the Expression of Proliferating Cell Nuclear Antigen in Murine Skin¶
Authors: Moore Julian O. , Palep Sapna R. , Saladl Rao N. , Gao Dayuan , Wang Yongyin , Phelps Robert G. , Lebwohl Mark G. , Wei Huachen
Published: July 2010

Proliferating cell nuclear antigen (PCNA) is an active nuclear protein involved in DNA replication, recombination and repair. PCNA is found throughout the basal layer in normal skin and in all layers of the epidermis in malignancy. This study evaluates PCNA's expression after acute and chronic UV‐B irradiation. Skh‐1 hairless mice exposed to 1.5 and 4.5 kJ/m2 of UV‐B were sacrificed at 6, 12, 24, 48, 72 and 168 h. Immunohistochemical analysis revealed PCNA expression throughout the basal layer of untreated skin, with diminished expression at 6 h, indicative of immediate UV damage, and evidenced by the observable upregulation in pyrimidine dimer formation early on. Subsequently, PCNA immunoreactivity progressively increased, demonstrating an aberrant upward epidermal migratory pattern in association with chronic exposure. The 4.5 kJ/m2 group exhibited prolonged recovery in staining and also demonstrated this altered migratory pattern with chronic exposure. Progressive reactivation of PCNA expression occurs with repair. PCNA migration to upper layers of the epidermis indicates proliferation and possibly a subsequent increased malignant potential. We conclude that PCNA can serve as a marker of DNA repair and indirectly as an indicator of UV‐B‐induced damage, expression being time dependent and dose related. Specific immunoreactivity patterns and the observable atypia in keratinocytes are relevant in elucidating malignant potentiation.
Ultraviolet‐B Radiation Causes an Upregulation of Survivin in Human Keratinocytes and Mouse Skin¶
Authors: Aziz Moammir Hasan , Ghotra Amaninderapal S. , Shukla Yogeshwer , Ahmad Nihal
Published: July 2010

Understanding of the mechanism of ultraviolet (UV)‐mediated cutaneous damages is far from complete. The cancer‐specific expression of Survivin, a member of the inhibitor of apoptosis family of proteins, coupled with its importance in inhibiting cell death and in regulating cell division, makes it a target for cancer treatment. This study was designed to investigate the modulation of Survivin during UV response, bothin vitroandin vivo. We used UV‐B‐mediated damages in normal human epidermal keratinocytes (NHEK) cells as anin vitromodel and SKH‐1 hairless mouse model for thein vivostudies. Forin vitrostudies, NHEK were treated with UV‐B and samples were processed at 5, 15, 30 min, 1,3,6,12 and 24 h after treatment. Our data demonstrated that UV‐B exposure (50 mJ/cm2) to NHEK resulted in a significant upregulation in Survivin messenger RNA (mRNA) and protein levels. We also observed that UV‐B exposure to NHEK resulted in significant (1) decrease in Smac/DIABLO and (2) increase in p53. Forin vivostudies, the SKH‐1 hairless mice were subjected to a single exposure of UV‐B (180 mJ/cm2), and samples were processed at 3, 6, 12 and 24 h after UV‐B exposure, UV‐B treatment resulted in a significant increase in protein or mRNA levels (or both) of Survivin, phospho‐Survivin and p53 and a concomitant decrease in Smac/DIABLO in mouse skin. This study demonstrated, for the first time, the involvement of Survivin (and the associated events) in UV‐B responsein vitroandin vivoin experimental models regarded to have relevance to human situations.
A far‐red fluorescent contrast agent to image epidermal growth factor receptor expression
Authors: Hsu Elizabeth R. , Anslyn Eric V. , Dharmawardhane Su , Alizadeh‐Naderi Reza , Aaron Jesse S. , Sokolov Konstantin V. , El‐Naggar Adel K. , Gillenwater Ann , Richards‐Kortum Rebecca R.
Published: July 2010

Recent developments in optical technologies have the potential to improve the speed and accuracy of screening and diagnosis of curable precancerous lesions and early cancer, thereby decreasing the costs of detection and management of epithelial malignancies. The development of molecular‐specific contrast agents for markers of early neoplastic transformation could improve the detection and molecular characterization of premalignant lesions. In the oral cavity, epidermal growth factor receptor (EGFR) overexpression has been identified in early stages of premalignant lesions of the oral squamous cell carcinoma; therefore, real‐time assessment of EGFR expression could serve as a biomarker for oral neoplasia. The purpose of our study was to develop a molecular‐specific optical contrast agent targeted against EGFR for in vivo assessment of epithelial neoplasia using a monoclonal antibody and the far‐red fluorescent dye, Alexa Fluor® 660 streptavidin. In addition to demonstrating the specificity of the contrast agent for EGFR in cell lines, we document the ability to achieve penetration through 500 μm thick epithelial layers using multilayer tissue constructs and permeability‐enhancing agents. Finally, using the fluorescence intensity of the contrast agent on fresh oral cavity tissue sections, we were able to distinguish abnormal from normal oral tissue. This contrast agent should have important clinical applications for use in conjunction with fluorescence spectroscopy or imaging (or both) to facilitate tumor detection and demarcation.
Effects of acid and alkali on the light absorption, energy transfer and secondary structures of core antenna subunits CP43 and CP47 of photosystem II
Authors: Shukui Guo , Chongqin Tang , Zhenle Yang , Liangbi Li , Tingyun Kuang , Yandao Gong , Nanming Zhao
Published: July 2010

The effects of acid and alkali treatment on the light absorption, energy transfer and protein secondary structure of the photosystem II core antenna CP43 and CP47 of spinach were investigated by the absorption spectra, fluorescence emission spectra and ciruclar dichroism spectra. It has been found that acid treatment caused the appearance of absorption characteristic of pheophytin a (Pheo a), whereas alkali treatment induced a new absorption peak at 642 nm. The energy transfer between β‐carotene and chlorophyll a (Chl a) in CP43 was easily disturbed by alkali, whereas in CP47 was readily affected by acid. As to the effects on the secondary structure of proetins in CP43 and CP47, effects of acid were far less than those of alkali. Both acid and alkali disturbed the microenvironment of Chl a and interfered exciton interaction between Chl a molecules. It was suggested that acid and alkali affect the light absorption, energy transfer and protein secondary structure of CP43 and CP47 in a differenty way. H+ can permeate into the internal space of α‐helix, change Chl a into Pheo a and disturb the microenvironment of pgiments without damaging the secondary structure of protein, whereas OH− can induce the protein unfolding at first, then saponify Chl a to chlorophyllide and disturb the microenvironment of pigments.
Angiogenesis Induced by Photodynamic Therapy in Normal Rat Brain¶
Authors: Jiang Feng , Zhang Zheng Gang , Katakowski Mark , Robin Adam M , Faber Michelle , Zhang Fan , Chopp Michael
Published: July 2010

Angiogenesis promotes tumor growth and invasiveness in brain. Because brain injury often induces expression of angiogenic‐promoting molecules, we hypothesize that oxidative insult induced by photodynamic therapy (PDT) could lead to an endogenous angiogenic response, possibly diminishing the efficacy of PDT treatment of tumors. Therefore, we sought to establish whether PDT induced an angiogenic response within the nontumored brain. PDT using Photofrin as a sensitizer at an optical dose of 140 J/cm2 was performed on normal rat brain (n = 30). Animals were sacrificed at 24 h, and 1,2,3 and 6 weeks after PDT treatment. Fluorescein isothiocyanatedextran perfusion was performed, and brains were fixed for immunohistological study. Immunostaining revealed that vascular endothelial growth factor (VEGF) expression increased within the PDT‐treated hemisphere 1 week after treatment and remained elevated for 6 weeks. Three‐dimensional morphologic analysis of vasculature within PDT‐treated and contralateral brain demonstrated PDT‐induced angiogenesis, as indicated by a significant increase in vessel connectivity (P < 0.001) concomitant with decreased (P < 0.05) mean segment length compared with vessels within the contralateral hemisphere. Volumetric measurement of angiogenic regions indicate that neovascular expansion continued for 4 weeks after PDT. These data demonstrate that PDT induces VEGF expression and neovascularization within normal brain. Because angiogenesis promotes growth and invasiveness of tumor, antagonizing this endogenous anglogenic response to PDT may present a practical means to enhance the efficacy of PDT.
Determination of the Minimal Erythema Dose and Colorimetric Measurements as Indicators of Skin Sensitivity to UV‐B Radiation¶
Authors: Dornelles Sérgio , Goldim José , Cestarl Tania
Published: July 2010

There is a strong relation between chronic UV‐B‐induced sunburns and the development of skin cancer. Therefore, it is important to obtain a method that can be reproduced easily to detect individuals with similar skin color but different sensitiveness to sun exposure. The study evaluated 193 healthy volunteers (68% women; the average age was 38 years). They were divided into six groups of at least 30 subjects, according to skin type. The minimal erythema dose (MED) was assessed in two non‐sun‐exposed areas (thorax‐infra‐axillary area and on the buttocks), using a UV‐B source (0.5 mW/cm2), with openings of 1 cm2, in increasing doses. The same areas were evaluated with a Minolta CR 300 Chromameter (L*a*b* system). The MED values ranged from 13 to 156 mJ/cm2; the coordinate L* (brightness) ranged from 75.96 to 30.15. The correlation between the MED and the brightness was negative in both areas (Pearson's correlation r =−0.91, P < 0.05). Color measurements, especially brightness, can be used to quickly assess skin sensibility. Considering the MED, there is a substantial overlapping of adjacent phototypes, but they could be separated into two groups: more sensitive individuals (Types I, II, III and IV) and less sensitive ones (Types V and VI).
5‐Aminolevulinic Acid‐Based Photodynamic Therapy in Leukemia Cell HL60¶
Authors: Zhang Su‐Juan , Zhang Zhen‐Xi
Published: July 2010

A study to explore the optimal experimental parameters and the photosensitization of 5‐aminolevulinic acid (ALA)‐based photodynamic therapy (PDT) in promyelocytic leukemia cell HL60 has been conducted, in which HL60 cells and their control groups, peripheral blood mononuclear cell (PBMC), first are incubated with different concentrations of ALA in dark for different periods of time and then followed by irradiating with different wavebands for different fluences. Fluorescence microscope and spectrofluorometer have been used to detect the fluorescence of protoporphyrin IX (PpIX) endogenously produced by ALA. The response of the cells to ALA‐PDT was evaluated by the 3‐(4,5‐dimethyl‐2‐thiazolyl)‐2‐5‐diphenyl‐2H‐tetrazolium bromide (MTT) assay (interval between irradiation and the MTT assay is 24 h) and by flow cytometry (the length of time between irradiation and the flow assay is 30 min). MTT results will reflect the relative number of metabolically active mitochondria in the population. Propidium iodide uptake in flow cytometry will test for membrane damage. The results of parameter experiments were obtained: 1 × 105/mL HL60 cell was first incubated with 1 mmol/L ALA in dark for 4 h and the maximum fluorescence of PpIX level appeared; then irradiated with 410 nm (4 mW/cm2) for 14.4 J/cm2 and maximum photodamage to membrane and mitochondrial function of HL60 cell resulted. With the normal granulocytes, such response was not detected. Therefore a hypothetical idea can be brought forward that ALA‐based PDT can be used for inactivation of leukemia cell HL60 and these optimal parameters may be useful for clinical application.
Reduction in DNA Synthesis During Two‐photon Microscopy of Intrinsic Reduced Nicotinamide Adenine Dinucleotide Fluorescence¶
Authors: Nichols Michael G. , Barth Erin E. , Nichols Jennifer A.
Published: July 2010

Two‐photon laser scanning microscopy (TPLSM) of endogenous reduced nicotinamide adenine dinucleotide (NAD(P)H) provides important information regarding the cellular metabolic state. When imaging the punctate mitochondrial fluorescence originating from NAD(P)H in a rat basophilic leukemia (RBL) cell at low laser powers, no morphological changes are evident, and photobleaching is not observed when many images are taken. At higher powers, mitochondrial NAD(P)H fluorescence bleaches rapidly. To assess the limitations of this technique and to quantify the extent of photodamage, we have measured the effect of TPLSM on DNA synthesis. Although previous reports have indicated a threshold power for “safe” two‐photon imaging, we find the laser power to be an insufficient indicator of photodamage. A more meaningful metric is a two‐photon‐absorbed dose that is proportional to the number of absorbed photon paris. A temporary reduction of DNA synthesis in RBL cells occurs whenever a threshold dose of approximately 2 × 1053 photon2 cm−4 s−1 is exceeded. This threshold is independent of laser intensity when imaging with average powers ranging from 5 to 17 mW at 740 nm. Beyond this threshold, the extent of the reduction is intensity dependent. DNA synthesis returns to control levels after a recovery period of several hours.
Self‐sensitized Photodegradation of Membrane‐bound Protoporphyrin Mediated by Chain Lipid Peroxidation: Inhibition by Nitric Oxide with Sustained Singlet Oxygen Damage
Authors: Niziolek Magdalena , Korytowski Witold , Girotti Albert W.
Published: July 2010

In the presence of exciting light, iron and reductants, the singlet oxygen (1O2)‐generating sensitizer protoporphyrin IX (PpIX) induces free radical lipid peroxidation in membranes, but gradually degrades in the process. We postulated that NO, acting as a chain‐breaking antioxidant, would protect PpIX against degradation and consequently prolong its ability to produce1O2. This idea was tested by irradiating PpIX‐containing liposomes (LUVs) in the presence of iron and ascorbate, and monitoring the cholesterol hydroperoxides 5α‐OOH and 7α/β‐OOH as respective1O2and free radical reporters. 5α‐OOH accumulation, initially linear with light fluence, slowed progressively after prolonged irradiation, whereas 7α/β‐OOH accumulation only accelerated after an initial lag. The active, but not spent, NO donor spermine NONOate (0.4 mM) virtually abolished 7α/β‐OOH buildup as well as 5α‐OOH slowdown. Increasing membrane phospholipid unsaturation hastened the onset of rapid chain peroxidation and 5α‐OOH slowdown. Accompanying the 5α‐OOH effect was a steady decrease in1O2quantum yield and PpIX fluorescence at 632 nm, both of which were inhibited by NO. An NO‐inhibitable decay of PpIX fluorescence was also observed during dark incubation of 5α‐OOH‐bearing LUVs with iron and ascorbate, confirming a link between chain peroxidation and PpIX loss. By protecting PpIX in irradiated membranes, NO might select for and prolong purely1O2‐mediated damage. Supporting this was our observation that1O2‐mediated photoinactivation of a nonmembrane target, lactate dehydrogenase, slowed concurrently with 5α‐OOH accumulation and that spermine NONOate prevented this. Thus, NO not only protected membrane lipids against PpIX‐sensitized free radical damage, but PpIX itself, thereby extending its1O2‐generating lifetime. Consistent findings were obtained using porphyrin‐sensitized COH‐BR1 cells. These previously unrecognized effects of NO could have important bearing on 5‐aminolevulinate‐based photodynamic therapy in which PpIX is metabolically deposited in tumor cells.
Chemopreventive Action of Xanthone Derivatives on Photosensitized DNA Damage¶
Authors: Hirakawa Kazutaka , Yoshida Maim , Nagatsu Akito , Mizukami Hajime , Rana Virendra , Rawat Mohan Singh Maniyari , Oikawa Shinji , Kawanishi Shosuke
Published: July 2010

Photosensitized DNA damage participates in solar‐UV carcinogenesis, photogenotoxicity and phototoxicity. A chemoprevention of photosensitized DNA damage is one of the most important methods for the above phototoxic effects. In this study, the chemopreventive action of xanthone (XAN) derivatives (bellidifolin [BEL], gentiacaulein [GEN], norswertianin [NOR] and swerchirin [SWE]) on DNA damage photosensitized by riboflavin was demonstrated using [32P]‐5′‐end‐labeled DNA fragments obtained from genes relevant to human cancer. GEN and NOR effectively inhibited the formation of piperidine‐labile products at consecutive G residues by photoexcited riboflavin, whereas BEL and SWE did not show significant inhibition of DNA damage. The four XAN derivatives decrease the formation of 8‐oxo‐7,8‐dihydro‐2′‐deoxyguanosine (8‐oxodGuo), an oxidative product of G, by photoexcited riboflavin. The preventive action for the 8‐oxodGuo formation of these XAN derivatives increased in the following order: GEN > NOR ≫ BEL > SWE. A fluorescence spectroscopic study and ab initio molecular orbital calculations suggested that the prevention of DNA photodamage is because of the quenching of the triplet excited state of riboflavin by XAN derivatives through electron transfer. This chemoprevention is based on neither antioxidation nor a physical sunscreen effect; rather, it is based on the quenching of a photosensitizer. In conclusion, XAN derivatives, especially GEN, may act as novel chemopreventive agents by the quenching mechanism of an excited photosensitizer.
Evaluation of a Single‐monochromator Diode Array Spectroradiometer for Sunbed UV‐radiation Measurements¶
Authors: Ylianttila Lasse , Visuri Reijo , Huurto Laura , Jokela Kari
Published: July 2010

The suitability of a new technology single‐monochromator diode array spectroradiometer for UV‐radiation safety measurements, in particular for sunbed measurements, was evaluated. The linearity, cosine response, temperature response, wavelength scale, stray‐light and slit function of the spectroradiometer were determined and their effects on the measurement accuracy evaluated. The main error sources were stray‐light and nonideal cosine response, for which correction methods are presented. Without correction, the stray‐light may reduce the accuracy of the measurement excessively, particularly in the UV‐B range. The expanded uncertainty of the corrected UV measurements is estimated to be 14%, which is confirmed with the comparative measurements carried out with a well‐characterized double‐monochromator spectroradiometer. The measurement accuracy is sufficient for sunbed measurements, provided that all corrections described above have been done and the user of the instrument has a good understanding of the instrument's operating principles and potential error sources. If these requirements are met, the tested spectroradiometer improves and facilitates market surveillance field measurements of sunbeds.
The Photochemistry of Thymidylyl‐(3′‐5′)‐5‐methyl‐2′‐deoxycytidine in Aqueous Solution¶
Authors: Celewicz Lech , Mayer Moriz , Shetlar Martin D.
Published: July 2010

The photochemistry of the dinucleoside monophosphate thymidylyl‐(3′‐5′)‐5‐methyl‐2′‐deoxycytidine (Tpm5dC) has been studied in aqueous solution using both 254 nm and UV‐B radiation. A variety of dinucleotide photoproducts containing 5‐methyl‐cytosine (m5C) have been isolated and characterized. These include two cyclobutane dimers (CBD) (thecis‐syn [c,s]andtranssynforms), a (6–4) adduct and its realted Dewar isomer, and two isomers of a product in which the m5C moiety was converted into an acrylamidine. Small amounts of thymidylyl‐(3′‐5′)‐thymidine (TpT) were also formed, presumably as a secondary photoreaction product. In addition, a photoproduct was characterized in which the m5C moiety was lost, thus generating 3′‐thymidylic acid esterified with 2′‐deoxyribose at the 5‐hydroxyl on the sugar mioety. Thec,sCBD of Tpm5dC readily undergoes deamination to form the corresponding CBD of TpT. The kinetics of this deamination process has been studied; the corresponding enthalpy and entropy of activation for the reaction have been evaluated at pH 7.4 as being, respectively, 73.4 kJ/mol and −103.5J/K mol. Deamination was not observed for the other characterized photoproducts of Tpm5dC.
UV Exposure of Elementary School Children in Five Japanese Cities¶
Authors: Ono Masaji , Munakata Nobuo , Watanabe Shaw
Published: July 2010

A 1 week UV‐exposure measurement and outdoor‐activity pattern survey was conducted for elementary school children for four seasons at five sites in Japan, i.e. Sapporo (43°05′N, altitude 40 m), Tsukuba (36°05′N, 20 m), Tokyo (35°40′N, 45 m), Miyazaki (31°60′N, 40 m) and Naha (26°10′N, 5 m), and UV exposure was measured directly and estimated using outdoor‐activity records. The study site with largest UV exposure was Miyazaki, a southern rural area. Comparing the results for boys and girls, UV exposure was larger in boys. UV exposure was large in spring and summer and small in winter. The total amount of UV exposure in spring and summer contributed 57.7–73.4% of total exposure for the year. As a whole, 8.1% and 1.8% of the schoolchildren were exposed to more than 1 minimum erythemal dose (MED) and 2 MED of solar UV in a day, respectively. The estimated yearly UV exposure ranged from 49 207 J/m2 in Miyazaki to 31 520 J/m2 in Tsukuba. The actual UV exposure correlated to potential UV exposure, estimated using outdoor‐activity records and ambient UV irradiance, but the ratio differed by season and site. The yearly average of percent UV exposure to ambient UV on a horizontal plane ranged from 9.9% in Tokyo to 4.0% in Naha. In the questionnaire survey on outdoor‐activity pattern, a short question “How long did you spend time outdoors between 0900 and 1500 h?” gives the best estimates of UV exposure.