UVB‐Induced Inflammatory Cytokine Release,DNA Damage and Apoptosis of Human Oral Compared with Skin Tissue Equivalents

People can get oral cancers from UV (290–400 nm) exposures. Besides high outdoor UV exposures, high indoor UV exposures to oral tissues can occur when consumers use UV‐emitting tanning devices to either tan or whiten their teeth. We compared the carcinogenic risks of skin to oral tissue cells after UVB (290–320 nm) exposures using commercially available 3D‐engineered models for human skin (EpiDerm?), gingival (EpiGing?) and oral (EpiOral?) tissues. To compare the relative carcinogenic risks, we investigated the release of cytokines, initial DNA damage in the form of cyclobutane pyrimidine dimers (CPDs), repair of CPDs and apoptotic cell numbers. We measured cytokine release using cytometric beads with flow cytometry and previously developed a fluorescent immunohistochemical assay to quantify simultaneously CPD repair rates and apoptotic cell numbers. We found that interleukin‐8 (IL‐8) release and the initial CPDs are significantly higher, whereas the CPD repair rates and apoptotic cell numbers are significantly lower for oral compared with skin tissue cells. Thus, the increased release of the inflammatory cytokine IL‐8 along with inefficient CPD repair and decreased death rates for oral compared with skin tissue cells suggests that mutations are accumulating in the surviving population of oral cells increasing people's risks for getting oral cancers.     

Oxidation of guanine in cellular DNA by solar UV radiation: biological role.

The formation of cyclobutane pyrimidine dimers (CPD) and 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodGuo) was investigated in Chinese hamster ovary cells upon exposure to either UVC, UVB, UVA or simulated sunlight (SSL). Two cell lines were used, namely AT3-2 and UVL9, the latter being deficient in nucleotide excision repair and consequently UV sensitive. For all types of radiation, including UVA, CPD were found to be the predominant lesions quantitatively. At the biologically relevant doses used, UVC, UVB and SSL irradiation yielded 8-oxodGuo at a rather low level, whereas UVA radiation produced relatively higher amounts. The formation of CPD was 10(2) and 10(5) more effective upon UVC than UVB and UVA exposure. These yields of formation followed DNA absorption, even in the UVA range. The calculated relative spectral effectiveness in the production of the two lesions showed that efficient induction of 8-oxodGuo upon UVA irradiation was shifted toward longer wavelengths, in comparison with those for CPD formation, in agreement with a photosensitization mechanism. In addition, after exposure to SSL, about 19% and 20% of 8-oxodGuo were produced between 290-320 nm and 320-340 nm, respectively, whereas CPD were essentially (90%) induced in the UVB region. However, the ratio of CPD to 8-oxodGuo greatly differed from one source of light to the other: it was over 100 for UVB but only a few units for UVA source. The extent of 8-oxodGuo and CPD was also compared to the lethality for the different types of radiation. The involvement of 8-oxodGuo in cell killing by solar UV radiation was clearly ruled out. In addition, our previously reported mutation spectra demonstrated that the contribution of 8-oxodGuo in the overall solar UV mutagenic process is very minor.     

Hydrochlorothiazide Enhances UVA‐Induced DNA Damage

The UVA is currently thought to be carcinogenic because, similar to UVB, it induces the formation of cyclobutane pyrimidine dimers (CPDs). Various drugs have been reported to cause photosensitive drug eruptions as an adverse effect. Although the precise mechanism of photosensitive drug eruption remains to be elucidated, it is generally accepted that free radicals and other reactive molecules generated via UV‐irradiated drugs play important roles in the pathogenesis of photosensitive drug eruptions. The waveband of concern for photo‐reactive drugs is UVA‐visible light, but some extend into the UVB region. We tested whether photosensitive drugs could enhance CPD formation after UVA exposure by using isolated DNA in the presence of several reported photosensitive drugs using high‐performance liquid chromatography. We found that the diuretic agent hydrochlorothiazide (HCT) significantly enhanced the production of TT dimers over a wide range of UVA. Furthermore, we investigated whether UVA plus HCT could enhance CPD production in xeroderma pigmentosum model mice defective in nucleotide excision repair. Immunofluorescence studies showed that CPD formation in the skin significantly increased after 365 nm narrow‐band UVA irradiation in the presence of HCT, compared with that in wild‐type mice. HCT could be used with caution because of its enhancement of UVA‐induced DNA damage.     

Ultraviolet B Inhibits Skin Wound Healing by Affecting Focal Adhesion Dynamics

As the most important interface between human body and external environment, skin acts as an essential barrier preventing various environmental damages, among which DNA‐damaging UV radiation from the sun remains the major environmental risk factor causing various skin diseases. It has been well documented that wavelengths in the ultraviolet B (UVB) radiation range (290–320 nm) of the solar spectrum can be absorbed by skin and lead to cutaneous injury and various other deleterious effects. During process such as wound healing, the orchestrated movement of cells in a particular direction is essential and highly regulated, integrating signals controlling adhesion, polarity and the cytoskeleton. Cell adhesion and migration are modulated through both of actin and microtubule cytoskeletons. However, little was known about how UVB affects skin wound healing and migration of epidermal keratinocytes. Here, we demonstrate that UVB can delay the wound healing progress in vivo with a murine model of full‐thickness skin wound. In addition, UVB significantly inhibited keratinocyte motility by altering focal adhesion turnover and cytoskeletal dynamics. Our results provide new insights into the etiology of UVB exposure‐induced skin damages.     

Fisetin Inhibits UVB‐induced Cutaneous Inflammation and Activation of PI3K/AKT/NFκB Signaling Pathways in SKH‐1 Hairless Mice

Solar ultraviolet B (UVB) radiation has been shown to induce inflammation, DNA damage, p53 mutations and alterations in signaling pathways eventually leading to skin cancer. In this study, we investigated whether fisetin reduces inflammatory responses and modulates PI3K/AKT/NFκB cell survival signaling pathways in UVB‐exposed SKH‐1 hairless mouse skin. Mice were exposed to 180 mJ cm^?2 of UVB radiation on alternate days for a total of seven exposures, and fisetin (250 and 500 nmol) was applied topically after 15 min of each UVB exposure. Fisetin treatment to UVB‐exposed mice resulted in decreased hyperplasia and reduced infiltration of inflammatory cells. Fisetin treatment also reduced inflammatory mediators such as COX‐2, PGE₂ as well as its receptors (EP1–EP4) and MPO activity. Furthermore, fisetin reduced the level of inflammatory cytokines TNFα, IL‐1β and IL‐6 in UVB‐exposed skin. Fisetin treatment also reduced cell proliferation markers as well as DNA damage as evidenced by increased expression of p53 and p21 proteins. Further studies revealed that fisetin inhibited UVB‐induced expression of PI3K, phosphorylation of AKT and activation of the NFκB signaling pathway in mouse skin. Overall, these data suggest that fisetin may be useful against UVB‐induced cutaneous inflammation and DNA damage.     

Cyclobutane Pyrimidine Dimer Density as a Predictive Biomarker of the Biological Effects of Ultraviolet Radiation in Normal Human Fibroblast

This study compared biological responses of normal human fibroblasts (NHF1) to three sources of ultraviolet radiation (UVR), emitting UVC wavelengths, UVB wavelengths, or a combination of UVA and UVB (solar simulator; emission spectrum, 94.3% UVA and 5.7% UVB). The endpoints measured were cytotoxicity, intra‐S checkpoint activation, inhibition of DNA replication and mutagenicity. Results show that the magnitude of each response to the indicated radiation sources was best predicted by the density of DNA cyclobutane pyrimidine dimers (CPD). The density of 6‐4 pyrimidine–pyrimidone photoproducts was highest in DNA from UVC‐irradiated cells (14% of CPD) as compared to those exposed to UVB (11%) or UVA–UVB (7%). The solar simulator source, under the experimental conditions described here, did not induce the formation of 8‐oxo‐7,8‐dihydroguanine in NHF1 above background levels. Taken together, these results suggest that CPD play a dominant role in DNA damage responses and highlight the importance of using endogenous biomarkers to compare and report biological effects induced by different sources of UVR.     

SPECTRAL DEPENDENCE OF UV-INDUCED IMMEDIATE AND DELAYED APOPTOSIS: THE ROLE OF MEMBRANE AND DNA DAMAGE

Abstract— The phototoxicity of each waveband region of UV radiation (UVR), i.e., UVA (32CM100 nm), UVB (290–320 nm) and UVC (200–290 nm), was correlated with an apoptotic mechanism using equilethal doses (10% survival) on murine lymphoma L5178Y-R cells. Apoptosis was qualitatively monitored for DNA "ladder" formation (multiples of 200 base pair units) using agarose gel electrophoresis, while the percentages of apoptotic and membrane-permeabilized cells were quantified over a postexposure time course using flow cytometry. The UVA1 radiation (340–400 nm) induced both an immediate (<4 h) and a delayed (>20 h) apoptotic mechanism, while UVB or UVC radiation induced only the delayed mechanism. The role of membrane damage was examined using a lipophilic free-radical scavenger, vitamin E. Immediate apoptosis and membrane permeability increased in a UVA1 dose-dependent manner, both of which were reduced by vitamin E. However, vitamin E had little effect on UVR-induced delayed apoptosis. In contrast, the DNA damaging agents 2,4- and 2,6-diaminotoluene exclusively induced delayed apoptosis. Thus, immediate apoptosis can be initiated by UVA 1-induced membrane damage, while delayed apoptosis can be initiated by DNA damage. Moreover, the results suggest that immediate and delayed apoptosis are two independent mechanisms that exist beyond the realm of photobiology.     

Protective Effect of Tropical Highland Blackberry Juice (Rubus adenotrichos Schltdl.) Against UVB‐Mediated Damage in Human Epidermal Keratinocytes and in a Reconstituted Skin Equivalent Model

Solar ultraviolet (UV) radiation, particularly its UVB (280–320 nm) spectrum, is the primary environmental stimulus leading to skin carcinogenesis. Several botanical species with antioxidant properties have shown photochemopreventive effects against UVB damage. Costa Rica's tropical highland blackberry (Rubus adenotrichos) contains important levels of phenolic compounds, mainly ellagitannins and anthocyanins, with strong antioxidant properties. In this study, we examined the photochemopreventive effect of R. adenotrichos blackberry juice (BBJ) on UVB‐mediated responses in human epidermal keratinocytes and in a three‐dimensional (3D) reconstituted normal human skin equivalent (SE). Pretreatment (2 h) and posttreatment (24 h) of normal human epidermal keratinocytes (NHEKs) with BBJ reduced UVB (25 mJ cm^?2)‐mediated (1) cyclobutane pyrimidine dimers (CPDs) and (2) 8‐oxo‐7,8‐dihydro‐2′‐deoxyguanosine (8‐oxodG) formation. Furthermore, treatment of NHEKs with BBJ increased UVB‐mediated (1) poly(ADP‐ribose) polymerase cleavage and (2) activation of caspases 3, 8 and 9. Thus, BBJ seems to alleviate UVB‐induced effects by reducing DNA damage and increasing apoptosis of damaged cells. To establish the in vivo significance of these findings to human skin, immunohistochemistry studies were performed in a 3D SE model, where BBJ was also found to decrease CPDs formation. These data suggest that BBJ may be developed as an agent to ameliorate UV‐induced skin damage.     

In Vitro Investigations on the Effect of Dermal Fibroblasts on Keratinocyte Responses to Ultraviolet B Radiation

Exposure to ultraviolet radiation is closely linked to the development of skin cancers in humans. The ultraviolet B (UVB) radiation wavelength (280–320 nm), in particular, causes DNA damage in epidermal keratinocytes, which are linked to the generation of signature premalignant mutations. Interactions between dermal fibroblasts and keratinocytes play a role in epidermal repair and regeneration after UVB‐induced damage. To investigate these processes, established two and three‐dimensional culture models were utilized to study the impact of fibroblast–keratinocyte crosstalk during the acute UVB response. Using a coculture system it was observed that fibroblasts enhanced keratinocyte survival and the repair of cyclobutane pyrimidine dimers (CPDs) after UVB radiation exposure. These findings were also mirrored in irradiated human skin coculture models employed in this study. Fibroblast coculture was shown to play a role in the expression and activation of members of the apoptotic cascade, including caspase‐3 and Bad. Interestingly, the expression and phosphorylation of p53, a key player in the regulation of keratinocyte cell fate postirradiation, was also shown to be influenced by fibroblast‐produced factors. This study highlights the importance of synergistic interactions between fibroblasts and keratinocytes in maintaining a functional epidermis while promoting repair and regeneration following UVB radiation‐induced damage.     

Diel Cycles of DNA Damage and Repair in Eggs and Larvae of Northern Anchovy, Engraulis mordax, Exposed to Solar Ultraviolet Radiation

Abstract— Damage from UVB radiation (280–320 nm) in the form of cyclobutane pyrimidine dimers (CPD) in DNA and the capacity for their repair were measured in newly spawned eggs and yolk-sac larvae of northern anchovy, Engraulis mordax, exposed to natural diel cycles of sunlight. The CPD were measured by a newly developed chemiluminescent immunoblot assay capable of measuring CPD in samples as small as 50 ng DNA. Eggs and yolk-sac larvae exposed to full irradiance levels died. At lower dose rates, equivalent to deeper more natural locations in the water column, there was a diel cycle of dimer concentration that tracked solar intensity. This diel cycle was due to the interaction of damage and repair processes. Repair of CPD in anchovy eggs and larvae could be attributed to true photodependent repair that could be stopped by moving samples into the dark. The CPD present at sunset remained until the following morning. The diel cycles of damage and repair were maintained over at least 4 days without a long-term upward or downward trend in dimer concentration. This indicates that at the UVB doses used for these experiments, there was no long-term accumulation of CPD nor an induction of increased repair capacity. Unhatched embryos spawned in the dark also exhibited a strong photorepair response, suggesting that photolyase expression was innate and not dependent on previous light exposure. The diel cycle observed here indicates that, at least for northern anchovy, the CPD concentration at the time of sampling is a good indicator of dose rate but a poor indicator of cumulative dose (i.e. late afternoon samples have the highest cumulative dose but relatively low CPD concentrations). The CPD immunoassay described here has the required sensitivity for measuring DNA damage in wild populations of ichthyoplankton exposed to natural sunlight. These results will guide the collection and interpretation of field data on natural levels of CPD in wild larvae collected at different depths and times of the day.     

UVB-induced Cyclobutane Pyrimidine Dimer Frequency Correlates with Skin Cancer Mutational Hotspots in p53

Abstract— Ultraviolet light has been identified as the major carcinogen in skin cancer and the p53 tumor suppressor gene is a major target for UV-induced mutations. The mutations are probably caused by unrepaired UV-induced cyclobutane pyrimidine dimers (CPD) and possibly by the less frequent pyrimidine (6-4) pyrimidone photoproducts. While hot spots for p53 mutations in human nonmela-noma skin tumors correspond quite well to slow spots for CPD repair in cultured cells irradiated with the model mutagen 254 nm UVC (which is not present in terrestrial sunlight), they do not all coincide with sequences that are initially frequently damaged by 254 nm UVC. Using LMPCR (ligation-mediated polymerase chain reaction), we show that environmentally relevant UVB light induces CPD at CC and PyrmC positions much more frequently than does UVC light, and that all eight skin cancer hot spots in p53 are also hot spots for UVB-induced CPD. Our results show that methylation of dipyrimidine sites (PyrmCpG) is associated with an increase rate of CPD formation upon UVB irradiation. Consequently, DNA methylation may increase the mutagenic potential of UVB and explains that several p53 mutation hot spots are found at PyrmCpG. The distribution patterns of CPD formation and the photofootprint patterns found along exons 5 and 6 of p53 gene are suggestive of DNA folding into nucleosomes.     

Quantification of Cyclobutane Pyrimidine Dimers Induced by UVB Radiation in Conidia of the Fungi Aspergillus fumigatus,Aspergillus nidulans,Metarhizium acridum and Metarhizium robertsii

Conidia are responsible for reproduction, dispersal, environmental persistence and host infection of many fungal species. One of the main environmental factors that can kill and/or damage conidia is solar UV radiation. Cyclobutane pyrimidine dimers (CPD) are the major DNA photoproducts induced by UVB. We examined the conidial germination kinetics and the occurrence of CPD in DNA of conidia exposed to different doses of UVB radiation. Conidia of Aspergillus fumigatus, Aspergillus nidulans and Metarhizium acridum were exposed to UVB doses of 0.9, 1.8, 3.6 and 5.4 kJ m⁻². CPD were quantified using T4 endonuclease V and alkaline agarose gel electrophoresis. Most of the doses were sublethal for all three species. Exposures to UVB delayed conidial germination and the delays were directly related both to UVB doses and CPD frequencies. The frequencies of dimers also were linear and directly proportional to the UVB doses, but the CPD yields differed among species. We also evaluated the impact of conidial pigmentation on germination and CPD induction on Metarhizium robertsii. The frequency of dimers in an albino mutant was approximately 10 times higher than of its green wild-type parent strain after exposure to a sublethal dose (1.8 kJ m⁻²) of UVB radiation.     

Action Spectra for UVB Impacts on Blepharisma japonicum Motility and Photobehavior

Abstract— The ciliate Blepharisma japonicum was exposed to artificial polychromatic and monochromatic UV radiation to evaluate the relative roles of UVB (280–315 nm UV radiation) and UVA (315–400 nm UV radiation) in altering its motility and photobehavior and to determine absolute weighting coefficients for these effects in the UVB range. Under polychromatic UV irradiation B. japonicum cells showed a severe reduction of cell speed and of the capability to respond to light stimuli. At low doses, however, UV caused a significant increase in the average velocity of a cell population. The UVB exclusion experiments indicated that UVA does not significantly alter motility and photoresponsiveness. The increase and the subsequent decrease in cell velocity was observed also under monochromatic irradiation at 281, 290 and 300 nm, whereas at 310 nm cells swim faster up to the highest photon flux density used. The cell capability of reacting to photic stimuli, conversely, steadily declined with increasing photon flux density at all the tested UVB wavelengths. The action spectra for the alteration of cell velocity and the impairment of photoresponsiveness show that the lower the irradiation wavelength, the more remarkable are the UVB effects and suggest different targets for the increase and the decrease in cell velocity.     

Photoreactivation in Paramecium tetraurelia under conditions of various degrees of ozone layer depletion

Photoreactivation (PR) is an efficient survival mechanism that helps protect cells against the harmful effects of solar-ultraviolet (UV) radiation. The PR mechanism involves photolyase, just one enzyme, and can repair DNA damage, such as cyclobutane-pyrimidine dimers (CPD) induced by near-UV/blue light, a component of sunlight. Although the balance of near-UV/blue light and far-UV light reaching the Earth's surface could be altered by the atmospheric ozone layer's depletion, experiments simulating this environmental change and its possible effects on life have not yet been performed. To quantify the strength of UVB in sunlight reaching the Earth's surface, we measured the number of CPD generated in plasmid DNA after UVB irradiation or exposure to sunlight. To simulate the increase of solar-UV radiation resulting from the ozone layer depletion, Paramecium tetraurelia was exposed to UVB and/or sunlight in clear summer weather. PR recovery after exposure to sunlight was complete at a low dose rate of 0.2 J/m2 x s, but was less efficient when the dose rate was increased by a factor of 2.5 to 0.5 J/m2 x s. It is suggested that solar-UV radiation would not influence the cell growth of P. tetraurelia for the reason of high PR activity even when the ozone concentration was decreased 30% from the present levels.     

Molecular Regulation of UV‐Induced DNA Repair

Ultraviolet (UV) radiation from sunlight is a major etiologic factor for skin cancer, the most prevalent cancer in the United States, as well as premature skin aging. In particular, UVB radiation causes formation of specific DNA damage photoproducts between pyrimidine bases. These DNA damage photoproducts are repaired by a process called nucleotide excision repair, also known as UV‐induced DNA repair. When left unrepaired, UVB‐induced DNA damage leads to accumulation of mutations, predisposing people to carcinogenesis as well as to premature aging. Genetic loss of nucleotide excision repair leads to severe disorders, namely, xeroderma pigmentosum (XP), trichothiodystrophy (TTD) and Cockayne syndrome (CS), which are associated with predisposition to skin carcinogenesis at a young age as well as developmental and neurological conditions. Regulation of nucleotide excision repair is an attractive avenue to preventing or reversing these detrimental consequences of impaired nucleotide excision repair. Here, we review recent studies on molecular mechanisms regulating nucleotide excision repair by extracellular cues and intracellular signaling pathways, with a special focus on the molecular regulation of individual repair factors.     

Effects and Mechanism of Nicotinamide Against UVA‐ and/or UVB‐mediated DNA Damages in Normal Melanocytes

Melanoma incidences are increasing rapidly, and ultraviolet (UV) radiation from the sun is believed to be its major contributing factor. UV exposure causes DNA damage in skin which may initiate cutaneous skin cancers including melanoma. Melanoma arises from melanocytes, the melanin‐producing skin cells, following genetic dysregulations resulting into hyperproliferative phenotype and neoplastic transformation. Both UVA and UVB exposures to the skin are believed to trigger melanocytic hyperplasia and melanomagenesis. Melanocytes by themselves are deficient in repair of oxidative DNA damage and UV‐induced photoproducts. Nicotinamide, an active form of vitamin B3 and a critical component of the human body's defense system has been shown to prevent certain cancers including nonmelanoma skin cancers. However, the mechanism of nicotinamide's protective effects is not well understood. Here, we investigated potential protective effects and mechanism of nicotinamide against UVA‐ and/or UVB‐ induced damage in normal human epidermal melanocytes. Our data demonstrated an appreciable protective effect of nicotinamide against UVA‐ and/or UVB‐ induced DNA damage in melanocytes by decreasing both cyclobutane pyrimidine dimers and 8‐hydroxy‐2′‐deoxyguanosine levels. We found that the photoprotective response of nicotinamide was associated with the activation of nucleotide excision repair genes and NRF2 signaling. Further studies are needed to validate our findings in in vivo models.     

ACTIVATION OF NF-KB IN HUMAN SKIN FIBROBLASTS BY THE OXIDATIVE STRESS GENERATED BY UVA RADIATION

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.     

Effects of cytosine methylation on pyrimidine dimer formation in DNA

The relative induction of cyclobutane pyrimidine dimers (CPD) and pyrimidine (6-4)pyrimidone photoproducts ([6-4]PD) was quantified in the duplex homopolymers polydeoxyadenosine:polydeoxythymidine, polydeoxyguanosine:polydeoxycytidine and polydeoxyguanosine:polydeoxy-5-methylcytidine irradiated with UVC or UVB radiation. Cytosine methylation significantly increased the yield of cytosine (6-4)PD after irradiation with UVC light and of cytosine CPD and (6-4)PD after irradiation with UVB light. The data suggest that CPD and (6-4)PD are preferentially induced at 5-methylcytosine bases in DNA of cells exposed to sunlight and comprise a major component of the mutation spectrum leading to the initiation of sunlight-induced skin cancer.     

Targeted Inactivation of DNA Photolyase Genes in Medaka Fish (Oryzias latipes)

Proteins of the cryptochrome/photolyase family (CPF) exhibit sequence and structural conservation, but their functions are divergent. Photolyase is a DNA repair enzyme that catalyzes the light‐dependent repair of ultraviolet (UV)‐induced photoproducts, whereas cryptochrome acts as a photoreceptor or circadian clock protein. Two types of DNA photolyase exist: CPD photolyase, which repairs cyclobutane pyrimidine dimers (CPDs), and 6‐4 photolyase, which repairs 6‐4 pyrimidine–pyrimidone photoproducts (6‐4PPs). Although the Cry‐DASH protein is classified as a cryptochrome, it also has light‐dependent DNA repair activity. To determine the significance of the three light‐dependent repair enzymes in recovering from solar UV‐induced DNA damage at the organismal level, we generated mutants in each gene in medaka using the CRISPR genome editing technique. The light‐dependent repair activity of the mutants was examined in vitro in cultured cells and in vivo in skin tissue. Light‐dependent repair of CPD was lost in the CPD photolyase‐deficient mutant, whereas weak repair activity against 6‐4PPs persisted in the 6‐4 photolyase‐deficient mutant. These results suggest the existence of a heretofore unknown 6‐4PP repair pathway and thus improve our understanding of the mechanisms of defense against solar UV in vertebrates.     

Formation of cyclobutane pyrimidine dimers and 8-oxo-7,8-dihydro-2'-deoxyguanosine in mouse and organ-cultured human skin by irradiation with broadband or with narrowband UVB

Narrowband UVB (NB-UVB) is a newly developed UVB source that, in addition to the previously used broadband UVB (BB-UVB), has been effectively used in phototherapy of various skin diseases. Besides its therapeutic effectiveness, NB-UVB also has some adverse effects that should be evaluated. As with all phototherapies, the photocarcinogenic potential of NB-UVB is the major concern. To assess the carcinogenic potential we measured the DNA damage induced by the two UVB sources because exposure of cells to UVB directly or indirectly induces DNA damage such as cyclobutane pyrimidine dimers (CPD) or 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodGuo), respectively. These types of DNA damage cause mutations of oncogenes and tumor suppressor genes, which can lead to photocarcinogenesis. In the present study we measured the yield of CPD and the oxidative DNA damage marker, 8-oxodGuo, in organ-cultured human skin and in mouse skin after exposure to NB-UVB or BB-UVB at therapeutically equivalent doses. We show that a 10-fold higher dose of NB-UVB yields a similar amount of CPD compared with BB-UVB in two types of samples examined. In contrast to CPD, the formation of 8-oxodGuo after irradiation with NB-UVB at a 10-fold higher dose is 1.5-3 times higher than that caused by BB-UVB. These results suggest that although NB-UVB at equivalent erythema-edema doses is not more potent in inducing CPD formation than is BB-UVB, NB-UVB may generate a higher yield of oxidized DNA damage.