Differential p53-mediated responses to solar-simulated radiation in human papillomavirus type 16-infected keratinocytes

In immunocompromised patients, cooperative effects of human papillomavirus (HPV) and ultraviolet (UV) radiation have been postulated in the development of non-melanoma skin cancers. The tumor suppressor p53 is a key component of the cellular response to genotoxic agents, such as UV radiation. We have previously demonstrated that in HPV16-infected cells, a higher E6* level was associated with a higher resistance to UV and oxidative stress. Using the two same SKv cell lines, the aim of the present study was to investigate p53 and p21 expression and cell death in HPV-infected keratinocytes in response to UV irradiation and to determine the role of HPV oncoprotein levels on the p53-mediated cellular response. We demonstrated that the weakly E6*-expressing level SKv-e cell line presented both higher cytotoxicity and apoptosis to UV. This high sensitivity was associated with both p53 and p21 nuclear accumulation, while a high E6* level and resistance were associated with no p53 accumulation and a p21 nuclear down-regulation after UV. Moreover, in SKv-e cell line, p21 promoter activation was p53 dependent. Our results suggest that an alteration and/or a modulation of the p53-p21 pathway in response to UV could be determinant for HPV-infected keratinocyte survival and HPV-associated carcinogenic process.     

Ultraviolet-B-induced oxidative DNA base damage in primary normal human epidermal keratinocytes and inhibition by a hydroxyl radical scavenger

To evaluate the effects of ultraviolet-induced environmental trauma on human skin cells, primary normal human epidermal keratinocytes were exposed to ultraviolet-B radiation (290-320 nm). We found that relatively low doses of ultraviolet-B (62.5-500 mJ per cm2) caused dose-dependent increases in 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxo-dG), a biomarker of oxidative DNA damage. Unirradiated normal human epidermal keratinocytes contained 1.49 (+/- 0.11) 8-oxo-dG per 10(6) 2'-deoxyguanosine (dG) residues in cellular DNA, which increased linearly to as high as 6.24 (+/- 0.85) 8-oxo-dG per 10(6) dG after irradiation with 500 mJ per cm2. Further, this oxidative damage was reduced by 60.7% when the cells were pretreated with 1 mM mannitol. As hydrogen peroxide (H2O2) is known to be generated during oxidative stress, its accumulation in ultraviolet-B-irradiated normal human epidermal keratinocytes was also assessed and correlated to 8-oxo-dG formation. An ultraviolet-B-induced increase in H2O2 was observed in normal human epidermal keratinocytes and its production was inhibited by the addition of catalase. Based on the ability of a neutral molecule like H2O2 to permeate membranes, our data indicate that, after ultraviolet-B irradiation, H2O2 migrates from the cytosol to the nucleus where it participates in a Fenton-like reaction that results in the production of hydroxyl radicals (OH*), which may then cause 8-oxo-dG formation in cellular DNA. This conclusion is supported by our data showing that OH* scavengers, such as mannitol, are effective inhibitors of oxidative DNA base damage. Although increased levels of 8-oxo-dG were previously found in immortalized mouse keratinocytes exposed to ultraviolet-B radiation, we now report the induction of 8-oxo-dG in normal human skin keratinocytes at ultraviolet-B doses relevant to human skin exposure.     

Effects of selenium compounds on induction of DNA damage by broadband ultraviolet radiation in human keratinocytes

Background Ultraviolet radiation (UVR), a ubiquitous environmental genotoxin for the skin, produces DNA damage. The trace element selenium induces synthesis of the glutathione peroxidase and thioredoxin reductase enzyme families. These selenoenzymes detoxify a range of toxic compounds generated by free radicals. Objectives To assess the effects of pretreatment of primary human keratinocytes with selenium on UVR-induced DNA damage. Methods Cells were irradiated with UVR from FS-20 lamps and were subjected to comet assay. Results Comet tail length due to UVR-induced T4 endonuclease V-sensitive sites (caused by cyclopyrimidine dimers, CPDs) increased to 35 +/- 4.5 microm (mean +/- SD) immediately after irradiation (time 0 h, 100%). After 4 h, 68% of the damage remained and after 24 h, 23% of the damage was still present. Treatment with up to 200 nmol L-1 selenomethionine or 50 nmol L-1 sodium selenite had no effect on CPD formation or rates of repair, or on the number of excision repair sites as measured by cytosine arabino furanoside and hydroxyurea treatment. However, selenite and selenomethionine protected against oxidative damage to DNA as measured by formation of formamidopyrimidine (FaPy) glycosylase-sensitive sites, which are indicative of 8-hydroxy-2-deoxyguanosine photoproduct formation. In this assay, irradiation of keratinocytes increased mean +/- SD glycosylase-specific comet tail length from 5 +/- 1.5 microm to 19 +/- 3.3 microm. Preincubation for 18 h with 50 nmol L-1 selenite abolished the UVR-induced increase in comet length. Preincubation with 200 nmol L-1 selenomethionine was similarly protective. Conclusions Selenite and selenomethionine protect keratinocytes from UVR-induced oxidative damage, but not from formation of UVR-induced excision repair sites.     

人类乳头瘤病毒16全基因在体外培养的正常人角质形成细胞 …

目的 利用含有人类乳头瘤病毒（ＨＰＶ）１６全基因的质粒转染正常人角质形成细胞，观察ＨＰＶ１６ｍＲＮＡ在转染细胞的表达。方法 用ＦｕＧＥＮＥ＾ＴＭ６转染试剂，将携带ＨＰＶ１６全基因的质粒ｐＳＶ２－ｎｅｏ／１６转染体外培养的正常人角质形成细胞，在转染后２４ｈ提取细胞总ＲＮＡ和ＤＮＡ，进行ＲＴ－ＰＣＲ和Ｓｏｕｔｈｅｒｎ印迹分析，结果 ２４ｈ后，ＲＴ－ＰＣＲ成功地扩增出１１０ｂｐ的片段，转染细胞中已出现Ｈ     

人类乳头瘤病毒16全基因在体外培养的正常人角质形成细胞中的瞬时表达

目的 利用含有人类乳头瘤病毒(HPV)16全基因的质粒转染正常人角质形成细胞,观察HPV16 mRNA在转染细胞的表达。方法 用FuGENE^TM6转染试剂,将携带HPV16全基因的质粒pSV2-neo/16转染体外培养的正常人角质形成细胞,在转染后24h提取细胞总RNA和DNA,进行RT-PCR和Southern印迹分析。结果 24h后,RT-PCR成功地扩增出110bp的片段,转染细胞中已出现HPV16 mRNA的表达,Southern印迹证实转染细胞中含有HPV16全基因7.9kb。结论 pSV2-neo/16转染正常人表皮角质形成细胞,24h内即可检测到HPV16mRNA表达。     

Disparate antioxidant enzyme activities in cultured human cutaneous fibroblasts, keratinocytes, and melanocytes

Antioxidant enzyme activities of cultured human foreskin fibroblasts, keratinocytes, and melanocytes from healthy black and Caucasian donors were measured and compared. Fibroblasts had more (p less than 0.05) peroxidase, catalase, glutathione peroxidase, and superoxide dismutase activity than keratinocytes. Keratinocytes had more (p less than 0.05) peroxidase, catalase, glutathione peroxidase, and superoxide dismutase activity than melanocytes. No differences in antioxidant enzyme activities were observed between the cells of any type taken from black or Caucasian people. Antioxidant enzyme activities may affect resistance to damage by oxidants induced by ultraviolet radiation and inflammation.     

表达人乳头瘤病毒6b和11型E6/E7基因的小鼠肿瘤细胞株的建立

目的 构建含有人乳头瘤病毒(HPV)6b和11型E6/E7基因的表达质粒和建立这些基因的转染细胞株。方法 分别将HPV6bE6、HPV6bE7、HPV11E6和HPV11E7克隆于含绿色荧光蛋白基因的真核表达载体pcDNA3.1,经脂质体介导转染B16细胞,以G418筛选阳性克隆,荧光显微镜观察,RT-PCR鉴定mRNA转录。结果 通过酶切鉴定和基因测序证实,构建质粒中目的基因片段序列完全正确。转染后荧光显微镜下可观测到B16细胞发出绿色荧光。以G418筛选的抗性细胞克隆提取RNA并经RT-PCR扩增出与目的基因大小一致的基因片段。结论 建立的小鼠肿瘤细胞B16表达HPV6bE6、HPV6bE7、HPV11E6和HPV11E7基因。提示转染的细胞株可移植于小鼠,利于体内研究这些基因的生物学作用和免疫学机制。     

16型人乳头瘤病毒E6/E7诱导角质形成细胞表达热休克蛋白70

目的 了解１６型人乳头瘤病毒（ＨＰＶ１６）Ｅ６／Ｅ７、ＨＳＰ７０与角质形成细胞永生化及转化之间的关系。方法 Ｌｉｐｏｆｅｃｔａｍｉｎｅ介导法进行转染,以Ｇ４１８筛选阳性克隆,免疫印迹分析及免疫荧光双标记染色结合激光扫描共聚焦显微镜对ＨＳＰ７０和ＨＰＶ１６Ｅ６／Ｅ７在转染然质形成细胞中的表达进行了观察。结果 免疫印迹分析显示正常负擀形成细胞和转染角质形成细胞均在相对分子质量７００００处出现一条染色带     

Functional regulation of tyrosinase and LAMP gene family of melanogenesis and cell death in immortal murine melanocytes after repeated exposure to ultraviolet B

The aim of this study was to investigate the effects of topical α-tocopherol application on epidermal and dermal antioxidants and its ability to prevent ultraviolet (UV)-induced oxidative damage. Hairless mice received topical applications of α-tocopherol 24 h before a single, acute UV irradiation (10 × minimal erythemal dose). The four major antioxidant enzymes (catalase, superoxide dismutase, glutathione reductase and glutathione peroxidase), hydrophilic and lipophilic antioxidants, and lipid hydroperoxides, markers of oxidative damage, were assayed in both epidermis and dermis of hairless mice. Topical α-tocopherol treatment increased dermal superoxide dismutase activity by 30% ( P  < 0.01) and protected epidermal glutathione peroxidase and superoxide dismutase from depletion after UV irradiation. Total and reduced glutathione levels in the epidermis increased by 50% after the topical treatment ( P  < 0.05), as did dermal ascorbate levels (by 40%; P  < 0.01). The topical treatment increased α-tocopherol levels both in the epidermis (62-fold) and the dermis (22-fold; P  < 0.001 in each layer). Furthermore, α-tocopherol treatment significantly reduced the formation of epidermal lipid hydroperoxides after UV irradiation ( P  < 0.05). These results demonstrate that topical administration of α-tocopherol protects cutaneous tissues against oxidative damage induced by UV irradiation in vivo , and suggest that the underlying mechanism of this effect involves the up-regulation of a network of enzymatic and non-enzymatic antioxidants.     

Flavonolignans from Silybum marianum moderate UVA-induced oxidative damage to HaCaT keratinocytes

BACKGROUND: UV radiation from sunlight is a very potent environmental risk factor in the pathogenesis of skin cancer. Exposure to UV light, especially the UVA part, provokes the generation of reactive oxygen species (ROS), which induce oxidative stress in exposed cells. Topical application of antioxidants is a successful strategy for protecting the skin against UV-caused oxidative damage. OBJECTIVE: In this study, silybin (SB) and 2,3-dehydrosilybin (DS) (1-50 micromol/l), flavonolignan components of Silybum marianum, were tested for their ability to moderate UVA-induced damage. METHODS: Human keratinocytes HaCaT were used as an appropriate experimental in vitro model, to monitor the effects of SB and DS on cell viability, proliferation, intracellular ATP and GSH level, ROS generation, membrane lipid peroxidation, caspase-3 activation and DNA damage. RESULTS: Application of the flavonolignans (1-50 micromol/l) led to an increase in cell viability of irradiated (20 J/cm(2)) HaCaT keratinocytes. SB and DS also suppressed intracellular ATP and GSH depletion, ROS production and peroxidation of membrane lipids. UVA-induced caspases-3 activity/activation was suppressed by treatment with SB and DS. Lower concentrations of both compounds (10 micromol/l) significantly reduced cellular DNA single strand break formation. CONCLUSION: Taken together, the results suggest that these flavonolignans suppress UVA-caused oxidative stress and may be useful in the treatment of UVA-induced skin damage.     

Sesamol inhibits UVB-induced ROS generation and subsequent oxidative damage in cultured human skin dermal fibroblasts

The exposure of cells to ultraviolet B radiation (UVB) can induce the production of reactive oxygen species (ROS) which damage cellular components. Free radical scavengers and antioxidants can interfere with the production of ROS. We studied cytotoxicity, intracellular ROS levels, lipid peroxidation, antioxidant status and oxidative DNA damage in cultured human skin dermal fibroblast adult cells (HDFa) exposed to UVB in the presence of sesamol, a natural phenolic compound. The levels of cytotoxicity, intracellular ROS, lipid peroxidation, oxidative DNA damage and apoptotic morphological changes were significantly increased in UVB irradiated HDFa cells. We also observed that the activities of enzymatic antioxidants (superoxide dismutase, catalase and glutathione peroxidase) and the levels of non-enzymatic antioxidant status (GSH) were significantly decreased in UVB irradiated cells. On the other hand, sesamol pretreatment significantly decreased cytotoxicity, intracellular ROS, lipid peroxidation, oxidative DNA damage and apoptotic morphological changes in sesamol-pretreated and UVB-irradiated HDFa cells. We have also observed increased enzymatic and non-enzymatic antioxidants status in sesamol plus UVB-irradiated cells. Among the different doses tested, 80 μM of sesamol shows maximum protection for UVB-induced oxidative damage. In conclusion, UVB-induced ROS formation, cell fatality, lipid peroxidation, antioxidant depletion and oxidative DNA damage in HDFa cells is inhibited by sesamol, which, probably through its ROS scavenging activity.     

Attenuation of UVA-induced damage to human keratinocytes by silymarin

BACKGROUND: UV radiation from sunlight is a potent environmental risk factor in skin cancer pathogenesis. UVA is the major portion of UV light reaching the earth surface ( approximately 95%) and it is reported to lead to benign and malignant tumor formation. UVA-mediated cellular damage occurs primarily through the release of reactive oxygen species (ROS) and it is responsible for inflammation, immunosuppression, photoaging and photocarcinogenesis. OBJECTIVE: The aim of our study was to investigate the potency of silymarin, the polyphenol fraction from the seeds of Silybum marianum, to modulate UVA-induced oxidative damage to human keratinocytes. METHODS: Skin epidermal cell line HaCaT, extensively used for studying the influence of UV radiation, was chosen as an experimental model. Silymarin's effect on UVA-disrupted cell viability, proliferation, mitochondrial function, and intracellular ATP and GSH level was measured. Furthermore, silymarin's potency to reduce UVA-induced ROS generation, membrane lipid peroxidation, caspase-3 activation and DNA damage was monitored. RESULTS: Treatment of irradiated HaCaT (20 J/cm(2)) with silymarin (0.7-34 mg/l; 4h) resulted in concentration-dependent diminution of UVA-caused oxidative stress on all studied parameters. Silymarin application extensively reduced GSH depletion and ROS production as well as lipid peroxidation in irradiated cells. Formation of UVA-induced DNA single strand breaks and caspase-3 activity was also significantly decreased by silymarin. CONCLUSION: The results suggest that silymarin may be beneficial in the treatment of UVA-induced skin oxidative injury and inflammation. However, further studies especially whose using human systems are needed to determine efficacy of silymarin in vivo.     

Autocytotoxic T-cell clones in lichen planus

We examined the in vitro cytotoxic activity of cutaneous T-cell lines and clones from lichen planus (LP) patients against autologous epidermal keratinocytes. T cells were cultured from LP lesions and adjacent clinically normal skin and cloned by limiting dilution. Keratinocytes were cultured from LP lesions and adjacent clinically normal skin and immortalized by transfection with the E6 and E7 genes from human papillomavirus 16 (HPV16). The lesional T-cell line from one LP patient contained 27% gammadelta+ T cells and was significantly more cytotoxic against autologous lesional keratinocytes than the T-cell line from clinically normal skin. Clones isolated from the lesional T-cell line were significantly more cytotoxic against autologous lesional keratinocytes than clones isolated from the non-lesional T-cell line. Most cytotoxic clones from LP lesions were CD8+ and most non-cytotoxic clones from LP lesions were CD4+. One cytotoxic clone was CD4- and CD8- and expressed the gammadelta T-cell receptor. Two CD8+ LP lesional T-cell clones showed dose-dependent killing of HPV16 E6/E7-immortalized autologous lesional and normal keratinocytes, but no cytotoxic activity against Epstein-Barr virus-transformed autologous B-cell blasts. The cytotoxic activity of CD8+ lesional T-cell clones against autologous lesional keratinocytes was partially blocked with anti-major histocompatibility complex (MHC) class I monoclonal antibodies. These data support the hypothesis that CD8+ lesional T cells recognize an antigen associated with MHC class I on lesional keratinocytes and that CD8+ cytotoxic T cells lyse keratinocytes in LP lesions.     

Role of solar conditioning in DNA repair response and survival of human epidermal keratinocytes following UV irradiation

We have investigated the cumulative effects of sunlight exposure upon the excision-repair of UV radiation damage to DNA in epidermal keratinocytes from human donors of different ages as well as the possible effect on DNA repair of periodic conditioning of the cultured keratinocytes with sublethal UV radiation exposures. We have also compared the growth properties of UV-irradiated keratinocytes derived from habitually sun-exposed and nonexposed areas from the bodies of young and aged donors. DNA repair replication in keratinocytes from habitually sun-exposed facial skin and the less sun-exposed abdominal skin of middle-aged adults was found to be similar, with respect to both the UV dose response and the time course of repair after 20 J/m2, 254 nm. Growth and survival (after exposure up to 50 J/m2, 254 nm) were greater for keratinocytes from protected areas of the upper arm of young donors (under 18 years) than for cells from their own sun-exposed areas. Growth and survival were markedly reduced for all keratinocyte cultures from aged donors, especially those cultures developed from sun-exposed areas. Nevertheless, the DNA repair response to UV radiation was similar in all cases. The evident uncoupling of UV sensitivity from DNA repair capacity remains to be understood. Our studies confirm that the cumulative effect of sunlight exposure indeed contributes to some skin aging processes. However, we have found no indication that an overall reduction in capacity for excision-repair of UV photoproducts in keratinocyte DNA accompanies senescence in human skin.     

HPV16 activates the AIM2 inflammasome in keratinocytes

Human papillomaviruses (HPV) are double-stranded DNA viruses, which selectively infect keratinocytes in stratified epithelia. After an initial infection, many patients clear HPV. In some patients, however, HPV persist, and dysfunctional innate immune responses to HPV infection could be involved in the ineffective clearing of these viruses. In this study, the mechanisms of HPV-induced immune responses in keratinocytes were investigated. Binding of viral DNA leads to AIM2 inflammasome activation and IL-1β release, while IFI16 activation results in IFN-β release. Using immunohistochemistry, AIM2 and IFI16—two recently identified sensors for cytosolic DNA—were also detected in HPV positive skin lesions. CISH stainings further confirmed the presence of cytosolic HPV16 DNA in biopsy samples. Moreover, active IL-1β and cleaved caspase-1 were detected in HPV infected skin, suggesting inflammasome activation by viral DNA. In subsequent functional studies, HPV16 DNA triggered IL-1β and IL-18 release via the AIM2 inflammasome in normal human keratinocytes. Although HPV DNA did not induce IFN-β in keratinocytes, IFN-β secretion was observed when AIM2 was blocked. Meanwhile, blocking of IFI16 increased HPV16 DNA-induced IL-1β, but not IL-18, secretion. These findings suggest crosstalk between IFI16 and AIM2 in the immune response to HPV DNA. In sum, novel aspects concerning HPV-induced innate immune responses were identified. Eventually, understanding the mechanisms of HPV-induced inflammasome activation could lead to the development of novel strategies for the prevention and treatment of HPV infections.     

人乳头状瘤病毒16 E6/E7基因稳定表达的永生化角质形成细胞的建立及鉴定

【摘要】 目的 构建人乳头状瘤病毒（HPV）16 E6/E7基因稳定表达的人永生化角质形成细胞（KC），为研究HPV16 E6/E7诱导的细胞永生化及恶性转化机制提供细胞模型。方法 两步消化法分离培养原代人包皮角质形成细胞（HFK），利用慢病毒感染技术对细胞稳定转染HPV16 E6/E7基因，连续培养30代以上，筛选出永生化KC，分为3组：①空白对照组：传代2次的原代HFK；②实验组：传代2次的原代HFK感染LV5-HPV16 E6/E7，感染细胞记录为A0代，感染后以传代次数记录（A1、A2……）；③阳性对照组：HPV16阳性宫颈癌细胞SiHa。应用实时荧光定量PCR（qRT-PCR）、Western印迹实验分别检测空白对照组、实验组、阳性对照组HPV16 E6/E7 mRNA、蛋白及CK14蛋白的表达，CCK-8及Transwell Insert方法检测细胞的增殖及侵袭能力。裸鼠致瘤实验检测实验组A30、阳性对照组SiHa细胞的致瘤能力。结果 成功分离原代HFK。LV5-HPV16 E6/E7重组质粒感染原代HFK后，空白对照组细胞无荧光表达，连续传代后出现衰老表现，实验组A30细胞体积、形态较原代HFK无明显变化，且荧光表达率为100%。与空白对照组相比，实验组A1、A10、A20、A30细胞HPV16 E6 mRNA表达水平显著升高（t值分别为7.12、8.07、6.53、5.66；P值分别 < 0.001、 < 0.001、 = 0.001、 = 0.005）；与空白对照组相比，实验组A1、A10、A20、A30细胞HPV16 E7 mRNA表达水平也显著升高（t值分别为3.20、4.29、3.75、4.22；P值分别为0.024、0.008、0.013、0.014）。空白对照组未见HPV16 E6/E7蛋白的表达，而A30及SiHa细胞可见HPV16 E6/E7蛋白的表达。CCK-8实验显示，实验组A10、A20、A30细胞增殖水平显著高于空白对照组（t值分别为6.49、7.55、9.43；P值分别为0.003、0.002、0.001），而A1的增殖水平与空白对照组比较，差异无统计学意义（t = 2.40，P = 0.074）。Transwell Insert侵袭实验显示，A30不能穿过基底膜，SiHa细胞可穿过基底膜被染成蓝色。裸鼠接种A30细胞2个月后无肉眼可见肿瘤，组织学亦显示无肿瘤形成，裸鼠接种SiHa细胞后可于皮下形成肿瘤。结论 通过采用慢病毒技术转染HPV16 E6/E7基因成功建立人永生化角质形成细胞，可作为HPV相关研究中的理想细胞模型。     

The role of human papillomavirus in common skin conditions: current viewpoints and therapeutic options

A direct causal relationship between human papillomavirus (HPV) infection and cervical neoplasia is well-accepted, but the specific role of HPV in the pathogenesis of other cutaneous disorders is less clear. This article explores the role of HPV in 2 common disorders associated with considerable morbidity: external genital and perianal warts (EGWs) and actinic keratosis (AK). Because the potential role of HPV in the pathogenesis of EGW and AK may have implications that influence management, the available topical pharmacotherapy for each disorder also is reviewed. External genital and perianal warts represent a possible phenotypic expression of HPV infection and results from hyperkeratosis and hyperplasia of keratinocytes. The cell cycle disruption caused by low-risk anogenital HPV subtypes (eg, HPV-6, HPV-11) is similar to high-risk HPV subtypes, except low-risk HPV E6 and E7 proteins likely bind regulatory proteins with less affinity. Although UV light clearly has a primary causal role in the development of AK, new data suggest that HPV infection, particularly with 3-HPV subtypes, may serve as a cocarcinogen. By impairing normal DNA repair and apoptotic mechanisms, HPV may set the stage for later UV-induced transformation. It also has been suggested that HPV may increase the severity of AK lesions and contribute to their recurrence following therapy.     

E6 and E7 proteins from different beta-papillomaviruses types do not interfere in UVB-induced apoptosis of HaCaT keratinocytes

Beta-papillomaviruses (beta-HPV) have been linked to the development of skin cancer in humans. Because both E6 and E7 proteins from beta-HPV have been involved in the potential carcinogenicity of these viruses, we investigated their role on UVB-induced apoptosis in HaCaT cell line. HaCaT cells have been transduced with both E6/E7 using a retroviral system and treated with PRIMA-1. Apoptosis was assessed by flow cytometry to measure mitochondrial membrane potential and DNA fragmentation. HaCat keratinocytes transduced with both E6 and E7 genes of seven beta-HPV types (HPV5, HPV8, HPV14, HPV24, HPV36, HPV38 and HPV49) did not demonstrate any inhibition of UVB-induced apoptosis, even after p53 reactivation through PRIMA-1. Our data suggest that the expression of E6 and E7 exert different modulatory effects on UVB-induced apoptosis according to beta-HPV types and to the cellular genetic context.     

Voriconazole enhances UV-induced DNA damage by inhibiting catalase and promoting oxidative stress

Cutaneous squamous cell carcinoma (cSCC) is the second most common form of skin cancer and is associated with cumulative UV exposure. Studies have shown that prolonged voriconazole use promotes cSCC formation; however, the biological mechanisms responsible for the increased incidence remain unclear. Here, we show that voriconazole directly increases oxidative stress in human keratinocytes and promotes UV-induced DNA damage as determined by comet assay, 8-oxoguanine immunofluorescence and mass spectrometry. Voriconazole treatment of human keratinocytes potentiates UV-induced apoptosis and activation of the p38 MAP kinase and 53BP1 UV stress response pathways. The p38 MAP kinase activation promoted by voriconazole exposure can be mitigated by pretreating keratinocytes with N-acetylcysteine. Voriconazole increases oxidative stress in keratinocytes by directly inhibiting catalase leading to lower intracellular NADPH levels and the triazole moieties in voriconazole are critical for inhibiting catalase. Furthermore, voriconazole is shown to promote UV-induced dysplasia in an in vivo model. Together, these data demonstrate that voriconazole potentiates oxidative stress in UV-irradiated keratinocytes through catalase inhibition. Use of antioxidants may mitigate the pro-oncogenic effects of voriconazole.