Ultraviolet Radiation-induced DNA Damage Research Articles

マイクロピラー基板を用いた核の力学的拘束が細胞の紫外線耐性に与える影響

DNA damage induced by the radiation, including ultraviolet (UV) light, exerts adverse effects on genome stability, alters the normal state of life, and causes many kinds of diseases. Thus, a biochemical or biomechanical method in DNA damage repair and protection is well required. Hear we investigated the effects of mechanical factors, such as mechanical deformation of the nucleus, on UV radiation resistance of DNA in epithelial-like cells derived from Xenopus laevis (XTC-YF). XTC-YF cells spread normally in the spaces between micropillars whose diameter, length, and center to center spacing was 3, 9, and 9 μm, respectively. Their nuclei showed remarkable deformation and appeared to be “trapped” mechanically on the array of pillars. We compared the cells cultured on the normal flat substrates and on the pillar substrates and found that the UV radiation-induced DNA damage estimated by the fluorescent intensity of the phospho-histone γ-H2A.X, was significantly inhibited in the cells cultured on the pillar substrates. The significant positive correlation was observed between fluorescent intensity of intranuclear DNA and γ-H2A.X in the cells cultured on the flat substrates following UV irradiation, while in the cells on the pillar substrate, their correlation became lower. These results indicate that the inhibition of UV radiation-induced DNA damages might be resulted from structural change of DNA caused by the mechanical stress of the nucleus of the cells on the pillars. Our study first demonstrated the nuclear stress-induced inhibition of DNA damages in living cells.

Infrared A radiation promotes survival of human melanocytes carrying ultraviolet radiation-induced DNA damage.

The link between solar radiation and melanoma is still elusive. Although infrared radiation (IR) accounts for over 50% of terrestrial solar energy, its influence on human skin is not well explored. There is increasing evidence that IR influences the expression patterns of several molecules independently of heat. A previous invivo study revealed that pretreatment with IR might promote the development of UVR-induced non-epithelial skin cancer and possibly of melanoma in mice. To expand on this, the aim of the present study was to evaluate the impact of IR on UVR-induced apoptosis and DNA repair in normal human epidermal melanocytes. The balance between these two effects is a key factor of malignant transformation. Human melanocytes were exposed to physiologic doses of IR and UVR. Compared to cells irradiated with UVR only, simultaneous exposure to IR significantly reduced the apoptotic rate. However, IR did not influence the repair of UVR-induced DNA damage. IR partly reversed the pro-apoptotic effects of UVR via modification of the expression and activity of proteins mainly of the extrinsic apoptotic pathway. In conclusion, IR enhances the survival of melanocytes carrying UVR-induced DNA damage and thereby might contribute to melanomagenesis.

XPC, XPF, TP53 and GSTP1 polymorphisms in prognosis of cutaneous melanoma patients.

9038 Background: XPC, XPF, TP53 and GSTP1 genes act on repair and detoxification of ultraviolet (UV) radiation-induced DNA damage, which are related to cutaneous melanoma (CM) development and progr...

Nicotinamide enhances repair of arsenic and ultraviolet radiation-induced DNA damage in HaCaT keratinocytes and ex vivo human skin.

Arsenic-induced skin cancer is a significant global health burden. In areas with arsenic contamination of water sources, such as China, Pakistan, Myanmar, Cambodia and especially Bangladesh and West Bengal, large populations are at risk of arsenic-induced skin cancer. Arsenic acts as a co-carcinogen with ultraviolet (UV) radiation and affects DNA damage and repair. Nicotinamide (vitamin B3) reduces premalignant keratoses in sun-damaged skin, likely by prevention of UV-induced cellular energy depletion and enhancement of DNA repair. We investigated whether nicotinamide modifies DNA repair following exposure to UV radiation and sodium arsenite. HaCaT keratinocytes and ex vivo human skin were exposed to 2μM sodium arsenite and low dose (2J/cm2) solar-simulated UV, with and without nicotinamide supplementation. DNA photolesions in the form of 8-oxo-7,8-dihydro-2′-deoxyguanosine and cyclobutane pyrimidine dimers were detected by immunofluorescence. Arsenic exposure significantly increased levels of 8-oxo-7,8-dihydro-2′-deoxyguanosine in irradiated cells. Nicotinamide reduced both types of photolesions in HaCaT keratinocytes and in ex vivo human skin, likely by enhancing DNA repair. These results demonstrate a reduction of two different photolesions over time in two different models in UV and arsenic exposed cells. Nicotinamide is a nontoxic, inexpensive agent with potential for chemoprevention of arsenic induced skin cancer.

Extreme tolerance and developmental buffering of UV-C induced DNA damage in embryos of the annual killifish Austrofundulus limnaeus.

Free-living aquatic embryos are often at risk of exposure to ultraviolet radiation (UV-R). Successful completion of embryonic development depends on efficient removal of DNA lesions, and thus many aquatic embryos have mechanisms to reverse DNA lesions induced by UV-R. However, little is known of how embryos that are able to enter embryonic dormancy may respond to UV-R exposure and subsequent DNA damage. Embryos of the annual killifish Austrofundulus limnaeus are unique among vertebrates because their normal embryonic development includes (1) a complete dispersion of embryonic blastomeres prior to formation of the definitive embryonic axis, and (2) entry into a state of metabolic depression and developmental arrest termed diapause. Here, we show that developing and diapausing embryos of A. limnaeus have exceptional tolerance of UV-C radiation and can successfully complete embryonic development after receiving substantial doses of UV-C, especially if allowed to recover in full-spectrum light. Recovery in full-spectrum light permits efficient removal of the most common type of DNA lesion induced by UV-R: cyclobutane pyrimidine dimers. Interestingly, whole-mount embryo TUNEL assays suggest that apoptosis may not be a major contributor to cell death in embryos UV-C irradiated during dispersion/reaggregation or diapause. We also observed embryo mortality to be significantly delayed by several weeks in diapausing embryos irradiated and allowed to recover in the dark. These atypical responses to UV-R induced DNA damage may be due to the unique annual killifish life history and provide insight into DNA damage repair and recognition mechanisms during embryonic dormancy.

Xpc, Xpf and P53 Polymorphisms Associated with Dna Repair and Prognosis in Cutaneous Melanoma

ABSTRACT Aim: XPC, XPF and P53 genes act on nucleotide excision repair of ultraviolet radiation-induced DNA damage and are involved in cutaneous melanoma (CM) development. The DNA repair ability is variable in humans.Variant C and wild Arg alleles of XPC A2920C and P53 Arg72Pro polymorphims encode proteins with lower activities in DNA repair than those encoded by other alleles, respectively. Variant C allele of XPF T30028C influences mRNA stability and affects levels of protein expression. This study aimed to evaluate whether these polymorphisms are associated with relapse-free survival (RFS) and overall survival (OS) of CM patients. Methods: We analyzed 232 CM patients diagnosed from 1989 to 2013 at a University Hospital. Genomic DNA of patients was analyzed by polymerase chain reaction, and enzymatic digestion, for discrimination of pertinent genotypes. RFS and OS were calculated using the univariate Kaplan-Meier estimate probabilities, and differences between survival curves were analyzed by the log-rank test. OS was calculated from the date of diagnosis until the date of death or last follow-up. Results: The median period of observation of patients in the study was 57 months (range: 1-285). At 60 months of follow-up, we observed that RFS of patients with XPF TT + TC genotypes was higher than that found in patients with CC genotype (73% vs 30%, P = 0.007). At this time, RFS was also higher in patients with XPF TT + TC plus P53 ArgPro + ProPro genotypes than in those with XPF CC plus P53 ArgArg genotypes (74% vs 19%, P = 0.004). At 120 months of follow-up, we observed that OS of patients with XPF TT + TC genotypes was higher than that observed in patients with CC genotype (86% vs 63%, P = 0.05). OS of patients with XPF TT + TC plus P53 ArgPro + ProPro genotypes was also higher than that found in patients with XPF CC plus P53 ArgArg genotypes (89% vs 68%, P = 0.07) at this time. In contrast, RFS and OS were worse in patients with Breslow thickness ≥1.5 mm (87% vs 52%, P = 0.0001; 88% vs 79%, P = 0.008), Clark level IV + V (79% vs 60%, P = 0.004; 86% vs 78%, P = 0.02) and advantage stage III + IV (77% vs 48% P = 0.0001; 88% vs 66% P = 0.0001), respectively. Conclusions: The data found herein provide evidence, for the first time, that inherited abnormality in DNA repair pathway related to XPF and P53 polymorphisms influence RFS and OS in CM patients. Disclosure: All authors have declared no conflicts of interest.

Loss of XRCC1 confers a metastatic phenotype to melanoma cells and is associated with poor survival in patients with melanoma

Ultraviolet (UV) radiation-induced DNA damage and genomic instability is one of the leading causes for melanoma. X-ray repair cross-complementary protein 1, XRCC1, plays a critically important role in base excision repair pathway. This study was therefore performed to analyze the correlation between XRCC1 expression, melanoma progression, and patient survival. Using a tissue microarray with a total of 119 patients with melanoma, we demonstrate that loss of XRCC1 expression is associated with the progression of disease from dysplastic nevi to primary melanoma and to metastatic melanoma. We found that the loss of XRCC1 was correlated with the progression of melanoma from AJCC stage II to stage III and with worse overall and disease-specific 5-yr and 10-yr survival of patients with melanoma. Furthermore, we also illustrate the inhibitory effect of XRCC1 on melanoma cell invasion and migration, which are the regulatory events in melanoma metastasis.

Once Upon a Mine: The Legacy of Uranium on the Navajo Nation

On a low, windswept rise at the southeastern edge of the Navajo Nation, Jackie Bell-Jefferson prepares to move her family from their home for a temporary stay that could last up to seven years. A mound of uranium-laden waste the size of several football fields, covered with a thin veneer of gravel, dominates the view from her front door. After many years of living next to the contamination and a litany of health problems she believes it caused, Bell-Jefferson and several other local families will have to vacate their homes for a third round of cleanup efforts by the U.S. Environmental Protection Agency (EPA). Decades of uranium mining have dotted the landscape across the Navajo Nation with piles of contaminated mine waste. The EPA has mapped 521 abandoned uranium mines on the reservation, ranging from small holes dug by a single prospector into the side of a mesa to large commercial mining operations.1 The Navajo people did not have a word for “radioactivity” when mining outfits looking for vanadium2 and uranium3 began moving onto their land in the 1940s, and they did not understand that radiation could be dangerous. They were not told that the men who worked in the mines were breathing carcinogenic radon gas and showering in radioactive water, nor that the women washing their husbands’ work clothes could spread radionuclides to the rest of the family’s laundry. Bell-Jefferson and her brother Peterson Bell played in and around the mines, splashing and swimming in pools of radioactive water that had been pumped out of the mines and then collected on their property. The contaminated water looked and tasted perfectly clean. Families used it for cooking, drinking, and cleaning. Hogans and corrals were built with mine wastes, as were roads. All that changed on 16 July 1979. Just about a mile and a half from Bell-Jefferson’s home, a dam broke at the United Nuclear Corporation mill, where workers processed ore from the nearby Northeast Church Rock uranium mine. The spill dumped 94 million gallons of mill process effluent and 1,100 tons of tailings—an acidic, radioactive sludge—into a large arroyo that emptied into the Puerco River.4 The Church Rock spill occurred less than four months after the partial meltdown of the Three Mile Island nuclear reactor, and it released three times as much radiation, making it the biggest nuclear spill in U.S. history, yet it received only a tiny fraction of the news coverage.5 Declared a Superfund site in 1983, the heaps of waste around the mill still cause radiation survey instruments to squeal from the invisible uranium atoms that remain active 30 years later.6 “This area used to be my playground,” Bell-Jefferson says. “Now it’s just a huge wound.” For the Bells and other Dine (the term by which many Navajo people refer to themselves), the Church Rock spill was a turning point. When corporate and government officials appeared in the spill’s aftermath and began inquiring into exposure to the slurry and potential health problems, the Navajo people finally learned the truth—far from being harmless, these uranium mines were poisoning people, and researchers say they will continue to do so for decades to come.

Arsenite binding-induced zinc loss from PARP-1 is equivalent to zinc deficiency in reducing PARP-1 activity, leading to inhibition of DNA repair

Inhibition of DNA repair is a recognized mechanism for arsenic enhancement of ultraviolet radiation-induced DNA damage and carcinogenesis. Poly(ADP-ribose) polymerase-1 (PARP-1), a zinc finger DNA repair protein, has been identified as a sensitive molecular target for arsenic. The zinc finger domains of PARP-1 protein function as a critical structure in DNA recognition and binding. Since cellular poly(ADP-ribosyl)ation capacity has been positively correlated with zinc status in cells, we hypothesize that arsenite binding-induced zinc loss from PARP-1 is equivalent to zinc deficiency in reducing PARP-1 activity, leading to inhibition of DNA repair. To test this hypothesis, we compared the effects of arsenite exposure with zinc deficiency, created by using the membrane-permeable zinc chelator TPEN, on 8-OHdG formation, PARP-1 activity and zinc binding to PARP-1 in HaCat cells. Our results show that arsenite exposure and zinc deficiency had similar effects on PARP-1 protein, whereas supplemental zinc reversed these effects. To investigate the molecular mechanism of zinc loss induced by arsenite, ICP-AES, near UV spectroscopy, fluorescence, and circular dichroism spectroscopy were utilized to examine arsenite binding and occupation of a peptide representing the first zinc finger of PARP-1. We found that arsenite binding as well as zinc loss altered the conformation of zinc finger structure which functionally leads to PARP-1 inhibition. These findings suggest that arsenite binding to PARP-1 protein created similar adverse biological effects as zinc deficiency, which establishes the molecular mechanism for zinc supplementation as a potentially effective treatment to reverse the detrimental outcomes of arsenic exposure.

The photoprotective and antioxidative properties of the flavonoid luteolin are synergistically augmented by tocopherol and ubiquinone

Ultraviolet radiation induces DNA damage and oxidative stress which can result in skin inflammation, photoaging and photocarcinogenesis. The flavonoid luteolin that is present in high amounts in the dyers weld, Reseda luteola, is one of the most potent antioxidative plant polyphenols that has also ultraviolet-absorbing properties.

The Photoprotective and Antioxidative Properties of Luteolin are Synergistically Augmented by Tocopherol and Ubiquinone

Ultraviolet radiation induces DNA damage and oxidative stress which can result in skin inflammation, photoaging, and photocarcinogenesis. The flavonoid luteolin that is present in high amounts in the dyers weld, Reseda luteola, is one of the most potent antioxidative plant metabolites and also has ultraviolet-absorbing properties.The aim of this study was to determine whether tocopherol and ubiquinone add synergistic antioxidative values to luteolin. None of the substances showed cytotoxic effects in concentrations from 0.25 to 4 µg/mL. The photoprotective and antioxidant effect of equivalent concentrations of luteolin, tocopherol, and ubiquinone and their combination in a ratio of 4 : 4 : 1 were studied in solar simulator irradiated human skin fibroblasts. Luteolin had a half-maximal radical scavenging concentration of 2 µg/mL, whereas tocopherol and ubiquinone were only effective at higher concentrations. None of the substances showed a phototoxic effect, and only luteolin had a moderate photoprotective effect at 2 µg/mL. The combination of luteolin, tocopherol, and ubiquinone exerted a synergistic radical scavenging effect already at a concentration of 0.25 µg/mL and a complete photoprotection at 2 µg/mL.In summary, our findings suggest that the potent antioxidant and photoprotective effect of flavonoids like luteolin may be further increased by the addition of low concentrations of other antioxidants such as tocopherol and ubiquinone.

Reduction of arsenite-enhanced ultraviolet radiation-induced DNA damage by supplemental zinc

Arsenic is a recognized human carcinogen and there is evidence that arsenic augments the carcinogenicity of DNA damaging agents such as ultraviolet radiation (UVR) thereby acting as a co-carcinogen. Inhibition of DNA repair is one proposed mechanism to account for the co-carcinogenic actions of arsenic. We and others find that arsenite interferes with the function of certain zinc finger DNA repair proteins. Furthermore, we reported that zinc reverses the effects of arsenite in cultured cells and a DNA repair target protein, poly (ADP-ribose) polymerase-1. In order to determine whether zinc ameliorates the effects of arsenite on UVR-induced DNA damage in human keratinocytes and in an in vivo model, normal human epidermal keratinocytes and SKH-1 hairless mice were exposed to arsenite, zinc or both before solar-simulated (ss) UVR exposure. Poly (ADP-ribose) polymerase activity, DNA damage and mutation frequencies at the Hprt locus were measured in each treatment group in normal human keratinocytes. DNA damage was assessed in vivo by immunohistochemical staining of skin sections isolated from SKH-1 hairless mice. Cell-based findings demonstrate that ssUVR-induced DNA damage and mutagenesis are enhanced by arsenite, and supplemental zinc partially reverses the arsenite effect. In vivo studies confirm that zinc supplementation decreases arsenite-enhanced DNA damage in response to ssUVR exposure. From these data we can conclude that zinc offsets the impact of arsenic on ssUVR-stimulated DNA damage in cells and in vivo suggesting that zinc supplementation may provide a strategy to improve DNA repair capacity in arsenic exposed human populations.

Nicotinamide enhances repair of ultraviolet radiation-induced DNA damage in human keratinocytes and ex vivo skin

Nicotinamide (vitamin B3) protects from ultraviolet (UV) radiation-induced carcinogenesis in mice and from UV-induced immunosuppression in mice and humans. Recent double-blinded randomized controlled Phase 2 studies in heavily sun-damaged individuals have shown that oral nicotinamide significantly reduces premalignant actinic keratoses, and may reduce new non-melanoma skin cancers. Nicotinamide is a precursor of nicotinamide adenine dinucleotide (NAD(+)), an essential coenzyme in adenosine triphosphate (ATP) production. Previously, we showed that nicotinamide prevents UV-induced ATP decline in HaCaT keratinocytes. Energy-dependent DNA repair is a key determinant of cellular survival after exposure to DNA-damaging agents such as UV radiation. Hence, in this study we investigated whether nicotinamide protection from cellular energy loss influences DNA repair. We treated HaCaT keratinocytes with nicotinamide and exposed them to low-dose solar-simulated UV (ssUV). Excision repair was quantified using an assay of unscheduled DNA synthesis. Nicotinamide increased both the proportion of cells undergoing excision repair and the repair rate in each cell. We then investigated ssUV-induced cyclobutane pyrimidine dimers (CPDs) and 8-oxo-7,8-dihydro-2'-deoxyguanosine (8oxoG) formation and repair by comet assay in keratinocytes and with immunohistochemistry in human skin. Nicotinamide reduced CPDs and 8oxoG in both models and the reduction appeared to be due to enhancement of DNA repair. These results show that nicotinamide enhances two different pathways for repair of UV-induced photolesions, supporting nicotinamide's potential as an inexpensive, convenient and non-toxic agent for skin cancer chemoprevention.

Visualization of UV-induced damage on single DNA molecules

Ultraviolet radiation induced DNA damage such as double stranded breaks and single stranded breaks is visualized at the level of single DNA molecules. Molecular observations provide a map of DNA radiation-mediated breakages, revealing the sequence dependency of DNA damage rather than random occurrences.

Non-melanoma skin cancer: The hygiene hypothesis

Protection against ultra violet radiation-induced DNA-damage in the skin is not only provided by the pigmentary system. The epidermal barrier consisting of stratum corneum keratinocytes, filaggrin and other proteins is an additional component of the UV-shield. Disruption of the epidermal barrier through frequent body cleansing with soaps and cosmetics may increase the risk of non-melanoma skin cancer.

Quantification of ultraviolet radiation-induced DNA damage in the urine of Swedish adults and children following exposure to sunlight

Context: DNA damage following exposure to ultraviolet radiation (UVR) is important in skin cancer development. The predominant photoproduct, cyclobutane thymine dimer (T=T), is repaired and excreted in the urine, where it provides a biomarker of exposure.Objective: To quantify urinary T=T levels after recreational sunlight exposure in adults and children.Methods: Average UVR doses were measured with personal dosimeters. Urinary T=T was analysed with 32P-postlabelling.Results: Background levels of T=T increased significantly following exposure to sunlight. Amounts of T=T in urine of children and adults were not significantly different after adjusting for area of skin exposed and physiological differences. UVR dose and amounts of T=T correlated for both adults and children.Conclusion: Recreational exposure to sunlight in Sweden induces levels of DNA damage, clearly detectable in urine.

Abstract 1599: Grape seed proanthocyanidins inhibit UV-induced tolerogenic T-cell response through DNA repair-dependent functional activation of Langerhans cells

Abstract It is well documented that ultraviolet (UV) radiation-induced DNA damage in the form of cyclobutane pyrimidine dimers (CPD) diminishes the antigen presenting capacity of epidermal Langerhans cells (LCs) and this leads to the suppression of immune system. Previously, we have shown that dietary grape seed proanthocyanidins (GSPs) inhibit UVB-induced immunosuppression in mice via activation of CD8+ T cells. However, it is not understood whether (1) GSPs target LCs while exerting its immunomodulatory effects on T cells in response to the application of cutaneous contact sensitizer, 2,4-dinitrofluorobenzene (DNFB) onto the UV-exposed skin, and whether (2) the ability of GSPs to activate dendritic cells/LCs correlates with their DNA repair properties. To understand this mechanism, we first determined DNA repair ability of GSPs in UV-exposed dendritic cells for which CD11c+ cells were prepared from bone marrow of xeroderma pigmentosum complementation group A-knockout (XPA-KO) mice which are deficient in DNA repair, and their wild-type (WT) counterparts. CD11c+ cells were exposed to UVB radiation with or without pretreatment with GSPs. Treatment of GSPs resulted in significant reduction in the number of CPD-positive cells in CD11c+ cells from WT mice, but this DNA repair effect of GSPs was not seen in the CD11c+ cells from XPA-KO mice. We then analyzed effects of dietary GSPs (0.5%, w/w) on secretion of cytokines by CD11c+ cells that were isolated from lymph nodes and spleens of mice (WT and XPA-KO) that were exposed to UVB [150 mJ/cm2 (WT), 20 mJ/cm2 (XPA-KO)]. Our results showed that in comparison to CD11c+ cells isolated from WT control mice (GSP-untreated), CD11c+ cells isolated from mice that received GSPs in diet secreted a significantly higher level of Th1 cytokines (IL-12, IFNα), while the levels of Th2 cytokines (IL-4, IL-10) were significantly lower, but this effect of GSPs was not observed in CD11c+ cells isolated from XPA-KO mice. Finally, we used an adoptive transfer approach, wherein draining lymph nodes were harvested from mice that were exposed to UVB with and without GSPs supplementation in diet and sensitized by DNFB onto the UVB-exposed skin. CD11c+ cells were positively selected and transferred into naïve mice that were subsequently challenged by application of DNFB on the ear skin. Naïve recipients that received CD11c+ cells from WT, GSPs-treated, UVB-irradiated donors exhibited full contact hypersensitivity (CHS) response, whereas no significant CHS was observed in XPA-KO mice. These data suggest that dietary GSPs inhibit UVB-induced immunosuppression in mice by improving the ability of LCs to mount an immune response via activation of T cells in a manner which correlates with its capacity to enhance repair of UVB-induced DNA damage. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1599. doi:1538-7445.AM2012-1599

Abstract 525: Toll-like receptor-4 mediated cutaneous immune responses augment ultraviolet radiation-induced DNA damage and tumor development

Abstract UV (ultraviolet) B induced DNA damage plays a critical role in development of skin cancer. The molecular basis for this biologic activity resides at least in part from the demonstrated ability of this form of radiant energy to damage DNA, predominantly in the form of cyclobutane pyrimidine dimers (CPD). The purpose of this study was to determine whether toll-like receptor-4 (TLR4) contributed to UV-induced DNA damage responses and cutaneous tumor development. TLR4 deficient and wild type (WT) mice were subjected to a local UVB induced DNA damage regimen consisting of 90 mJ/cm2 UVB radiation. Wild type mice exhibited significant (p<0.05) DNA damage in the form of cyclobutane pyrimidine dimers (CPD), whereas TLR4 deficient mice developed significantly fewer CPD (p<0.05) in their skin and bone marrow dendritic cells (BMDC). The expression of DNA repair gene XPA was significantly less (p<0.05) in skin and BMDC from WT mice than TLR4 deficient mice after UVB exposure. When cytokine levels were compared in these two strains after UVB exposure, BMDC from UV-irradiated TLR4 deficient mice produced significantly more IL-12 and IL-23 cytokines (p<0.05) than BMDC from WT mice. Furthermore, we found that Toll like receptor-4 (TLR4) promoted UVB induced tumor development. We observed that UVB induced skin carcinogenesis was retarded in terms of tumor incidence, and tumor latency, in mice deficient in TLR4 compared to WT mice, whereas significantly greater (p<0.05) numbers of tumors occurred in WT mice. Further, we found that tumor-bearing WT mice produced more IL-17 and IL-10 (p<0.05), whereas TLR4 deficient mice produced more IFN-γ (p<0.05). CD4+CD25+ regulatory T-cells from WT mice produced more IL-10 (p<0.05) than regulatory T-cells from TLR4 deficient mice. There was a significant increase (p<0.05) in Foxp3 expression in skin of WT mice than TLR4 deficient mice. Thus, strategies to inhibit TLR4 may allow us to develop immunopreventive and immunotherapeutic approaches for management of UVB induced cutaneous DNA damage and skin cancer. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 525. doi:1538-7445.AM2012-525

Ultraviolet radiation-induced DNA damage in bryophytes: what is already known and new results on rapid repair in a liverwort in an artificial diel cycle.

We studied, under laboratory conditions, the diel variations of DNA damage in response to changes in ultraviolet (UV) radiation in the liverwort Jungermannia exsertifolia subsp. cordifolia. The samples were exposed to three radiation regimes: P (only photosynthetically active radiation, PAR), PA (PAR + UV-A), and PAB (PAR + UV-A + UV-B). The day was divided in four periods: darkness, a first low-PAR period, a high-PAR plus UV period, and a second low-PAR period. After 15 days of culture, we measured the amount of thymine dimers in the four periods of the day on two consecutive days. DNA damage was significantly stronger in PAB than in P and PA samples, which corroborates the fact that it is a good and considerably specific biomarker of artificially enhanced UV-B. The diel changes in DNA damage consisted of a significant increase in thymine dimers at the end of the period of high-PAR plus UV, followed by a quick and complete repair after UV cessation and the instauration of low-PAR. These diel changes occurred much more clearly in PAB samples than in PA and P samples, and thus the differences in DNA damage were mainly due to UV-B. The low amount of thymine dimers produced and the rapid repair indicated that the UV-B levels applied caused little DNA damage in this liverwort, which is well adapted to those UV-B levels. This is the first time in which diel variations in DNA damage in response to artificial changes in UV-B are detected in a liverwort. Se han estudiado, en condiciones de laboratorio, los daños producidos en el ADN de la hepática Jungermannia exsertifolia subsp. cordifolia por exposición a la radiación ultravioleta (UV) a lo largo de ciclos diarios artificiales de 24 horas. Las muestras se expusieron a tres regímenes de radiación: P (con sólo radiación fotosintéticamente activa, PAR), PA (PAR + UV-A), y PAB (PAR + UV-A + UV-B). El día se dividió en cuatro periodos: oscuridad, un primer periodo de baja PAR, un periodo de alta PAR más radiación UV, y un segundo periodo de baja PAR. Después de 15 días de cultivo, se midió la cantidad de dímeros de timina en los cuatro periodos del día en dos días consecutivos. Los daños en el ADN fueron significativamente mayores en las muestras PAB que en las P y PA, lo que confirma que es un biomarcador bueno y considerablemente específico del aumento artificial de la radiación UV-B. Con respecto a los cambios en el ciclo diario, el número de dímeros de timina aumentó significativamente al final del periodo de alta PAR más radiación UV, y posteriormente se produjo una reparación rápida y completa de los daños tras el apagado de la radiación UV y la instauración del segundo periodo de baja PAR. Estos cambios fueron mucho más claros en las muestras PAB que en las P y PA, por lo que las diferencias en los daños al ADN se debieron principalmente a la radiación UV-B. La baja cantidad de dímeros de timina producidos y la rapidez de la reparación indican que los niveles de radiación UV-B aplicados provocaban pocos daños en el ADN de esta especie, que por lo tanto está bien adaptada a dichos niveles. Ésta es la primera vez que se detectan variaciones en los daños al ADN de una hepática en respuesta a cambios artificiales de UV-B a lo largo del ciclo diario.

Silymarin Protects Epidermal Keratinocytes from Ultraviolet Radiation-Induced Apoptosis and DNA Damage by Nucleotide Excision Repair Mechanism

Solar ultraviolet (UV) radiation is a well recognized epidemiologic risk factor for melanoma and non-melanoma skin cancers. This observation has been linked to the accumulation of UVB radiation-induced DNA lesions in cells, and that finally lead to the development of skin cancers. Earlier, we have shown that topical treatment of skin with silymarin, a plant flavanoid from milk thistle (Silybum marianum), inhibits photocarcinogenesis in mice; however it is less understood whether chemopreventive effect of silymarin is mediated through the repair of DNA lesions in skin cells and that protect the cells from apoptosis. Here, we show that treatment of normal human epidermal keratinocytes (NHEK) with silymarin blocks UVB-induced apoptosis of NHEK in vitro. Silymarin reduces the amount of UVB radiation-induced DNA damage as demonstrated by reduced amounts of cyclobutane pyrimidine dimers (CPDs) and as measured by comet assay, and that ultimately may lead to reduced apoptosis of NHEK. The reduction of UV radiation-induced DNA damage by silymarin appears to be related with induction of nucleotide excision repair (NER) genes, because UV radiation-induced apoptosis was not blocked by silymarin in NER-deficient human fibroblasts. Cytostaining and dot-blot analysis revealed that silymarin repaired UV-induced CPDs in NER-proficient fibroblasts from a healthy individual but did not repair UV-induced CPD-positive cells in NER-deficient fibroblasts from patients suffering from xeroderma pigmentosum complementation-A disease. Similarly, immunohistochemical analysis revealed that silymarin did not reduce the number of UVB-induced sunburn/apoptotic cells in the skin of NER-deficient mice, but reduced the number of sunburn cells in their wild-type counterparts. Together, these results suggest that silymarin exert the capacity to reduce UV radiation-induced DNA damage and, thus, prevent the harmful effects of UV radiation on the genomic stability of epidermal cells.