UVB-induced Cell Research Articles

Naproxen Inhibits UVB‐induced Basal Cell and Squamous Cell Carcinoma Development in Ptch1+/−/SKH‐1 Hairless Mice

Naproxen possesses anti-proliferative and pro-apoptotic effects besides its known anti-inflammatory functions. Here, we demonstrate the anticancer effects of naproxen against UVB-induced basal cell carcinoma (BCCs) and squamous cell carcinoma (SCCs) in a highly susceptible murine model of UVB carcinogenesis. Naproxen significantly inhibited UVB-induced BCCs and SCCs in this model. Tumor number and volume were significantly decreased (P < 0.005 and P < 0.05, respectively). Inhibition in UVB-induced SCCs and BCCs was 77% and 86%, respectively, which was associated with reduced PCNA and cyclin D1 and increased apoptosis. As expected, inflammation-related iNOS, COX-2 and nuclear NFκBp65 were also diminished by naproxen treatment. Residual tumors excised from naproxen-treated animal were less invasive and showed reduced expression of epithelial-mesenchymal transition (EMT) markers N-cadherin, Vimentin, Snail and Twist with increased expression of E-cadherin. In BCC and SCC cells, naproxen-induced apoptosis and activated unfolded protein response (UPR) signaling with increased expression of ATF4, p-eIF2α and CHOP. Employing iRNA-based approaches, we found that naproxen-induced apoptosis was regulated by CHOP as sensitivity of these cutaneous neoplastic cells for apoptosis was significantly diminished by ablating CHOP. In summary, these data show that naproxen is a potent inhibitor of UVB-induced skin carcinogenesis. ER stress pathway protein CHOP may play an important role in inducing apoptosis in cancer cells.

Anti-aging effects of casuarictin on UVB-induced CCD-986sk cell

본 연구는 새로운 기능성 화장품 소재 개발을 위해 복분자 씨앗의 구성성분인 casuarictin을 이용하여 섬유아세포내에서의 항노화 효과를 확인 하고자 하였다. Casuarictin의 항노화 효과를 CCD-986sk 세포를 이용하여 측정한 결과, casuarictin은 UVB에 의해 증가된 ROS 발현을 억제하였으며, pro-collagen의 발현을 증가시키고 MMP-1의 발현을 억제하는 것으로 나타났다. 이는 노화 인자인 활성산소종의 생성 억제와 주름과 밀접한 관련이 있는 콜라겐의 합성 및 분해억제를 통해 항산화 및 주름개선 효과가 있는 것으로 판단되며, 따라서 casuarictin은 화장품 분야에서 항노화 소재로서의 활용가치가 있을 것으로 사료된다.

Effect of hemp fiber on UVB-induced epidermal cell proliferation and PCNA expression

Ultraviolet radiation commonly causes serious skin diseases, and skin cell death. The UVB-blocking effect of hemp fabric which known to be a powerful agent against UVB was evaluated using mouse auricle skin. Based on UVB irradiation and the use of different fabrics, four mouse groups were evaluated in this study: two experimental groups, Group 1 (UVB exposed hemp fabric-shield site, EHFS), Group 2 (UVB exposed polyester fabric-shield site, EPFS), and two control groups, Group 3 (UVB exposed non-fabric-shield site, ENFS), and Group 4 (UVB unexposed non-fabric-shield site, UNFS). Except for UNFS all samples were exposed to UVB for 28 h and showed clear histologic changes in epidermis and dermis. After 45 h chronic irradiation, epidermal thickness was doubled in EHFS, roughly tripled in EPFS, and more than quadrupled in ENFS over that of UNFS. Based on the thickness of the altered epidermis, the blocking effect of hemp fabric was 50% higher than that of polyester fabric. Additionally, immunohistochemical analysis revealed expression of proliferating cell nuclear antigen (PCNA) in ENFS and EPFS throughout the hyperplasia of keratinocytes and sebocytes. After 45 h irradiation, the sebocytes of sebaceous glands, observed in sectioned images, increased on average to 14 cells in ENFS, 9 cells in EPFS, and 7 cells in EHFS, as compared to 8 cells in UNFS. In contrast, the cell area of adipose tissue was decreased by half in EHFS, one-fourth in EPFS, and one-tenth in ENFS, and mostly replaced with fibroblasts and other supporting cells. These results suggest that UVB irradiation directly affects epidermal and dermal tissues, and induces abnormal proliferation of keratinocytes and hyperplasia of sebocytes consuming fats in adipose tissue. For skin health, hemp fabric is a better material for protecting the skin against UVB than polyester fabric.

Abstract B12: Expression of Integrated Stress Response proteins during the progression of UVB-induced squamous cell carcinoma

Abstract The major environmental risk factor for developing non-melanoma skin cancers (NMSCs) is exposure to the ultraviolet-B (UVB) wavelengths found in sunlight. Unlike other types of cancer where the occurrence each year is precisely known, so many NMSCs occur each year in the United States that the incidence of NMSC is not accurately reported. Treatment of NMSC costs the U.S. healthcare system over 1 billion dollars annually. Therefore while NMSC is rarely lethal, the expense of NMSC treatment is the fifth highest among all cancer treatments. In response to a myriad of environmental stresses, including UVB irradiation, eukaryotic cells rapidly modulate protein synthesis. An important mechanism for translation regulation involves phosphorylation of the α subunit of eukaryotic initiation factor 2 (eIF2α), which results in a prompt decrease in global protein synthesis, concurrent with preferential translation of cytoprotective gene transcripts. Because a range of different stresses can activate one of four eIF2 kinases, this pathway is referred to as the Integrated Stress Response (ISR). Although the ISR pathway has been implicated in carcinogenesis in a variety of tissues, little is known about whether the ISR can modulate the development of NMSC in human skin. We previously defined the cytoprotective role of the ISR in response to UVB-irradiation of human keratinocytes in vitro. Following UVB exposure, keratinocytes lacking an ISR response have a shortened G1 arrest, a decreased ability to repair UVB-damaged DNA, a diminished senescence response, and an increased susceptibility to apoptosis. Collectively, these functions attributed to the ISR response in keratinocytes following UVB-irradiation are tumor suppressive. To address how the ISR is involved during photocarcinogenesis, we analyzed the expression of ISR proteins (eIF2α-phosphorylation, ATF4, CHOP, GADD34) in the progression of a subtype of NMSC, squamous cell carcinoma (SCC). The progression ranges from normal skin, to a precursor lesion of SCC (actinic keratosis, AK), and finally to SCC tumors. ISR marker proteins were detected using immunofluorescence on formalin-fixed, paraffin-embedded tissue from at least four distinct specimens in each category. In normal skin, expression of ISR proteins were found in differentiating layers of the epidermis. eIF2α phosphorylation was predominantly observed in both the nucleus and cytoplasm of keratinocytes in the squamous and granular layers of the skin. Similarly, GADD34, ATF4, and CHOP were expressed in normal differentiating keratinocytes. In contrast, using a human skin model of AK, ATF4 expression was greatly increased in all but the basal cell layer of the epidermis. CHOP and GADD34 expression, as well as eIF2α phosphorylation, in human AKs followed a similar pattern. Progression to SCC resulted in a striking decrease in ISR protein expression. ATF4 and CHOP expression were absent, no eIF2α phosphorylation was detected, and GADD34 expression was greatly diminished. Therefore, the standard ISR response in differentiating keratinocytes is enhanced in AK lesions but silenced as the tumor progresses to SCC. These data suggest that sustaining the ISR response in UVB-damaged epidermis could be a valuable therapeutic target for the prevention of SCC development. Citation Format: Dan F. Spandau, Ann Collier, Ron Wek. Expression of Integrated Stress Response proteins during the progression of UVB-induced squamous cell carcinoma. [abstract]. In: Proceedings of the AACR Special Conference on Translational Control of Cancer: A New Frontier in Cancer Biology and Therapy; 2016 Oct 27-30; San Francisco, CA. Philadelphia (PA): AACR; Cancer Res 2017;77(6 Suppl):Abstract nr B12.

Protective effects of Polypodium leucotomos extract against UVB-induced damage in a model of reconstructed human epidermis.

Polypodium leucotomos (PL) exerts potent antioxidant, photo-protective, and immune-modulatory activities. A reconstructed human epidermis (RHE) (Episkin) is a suitable model for the evaluation of acute UV-induced cell damage. No data regarding the photo-protective action of PL in this model are available. We evaluated the effects of PL on the prevention of UVB-induced cell damage assessing sunburn cells, CPD formation, p53, Ki-67, p21 expression, and epidermal growth factor (EGF) production. RHE was incubated in standard conditions. PL was topically applied at the concentration of 2 mg/cm2 , immediately before UVB exposition. UVB exposition (300 mJ/cm2 ) was performed using a dedicated UVB lamp. Irradiated samples without PL and non-irradiated samples were used as positive and negative controls. Expression of p53, p21, and Ki-67 was evaluated with immune-histochemical methods. CPD were measured using a monoclonal antibody. PL significantly reduced sunburned cells (-80%) in comparison with positive control. PL significantly prevented the increase in EGF production at tested times. PL significantly reduced the p53 (-80%), p21 (-84%), and Ki-67 (-48%) positive cells. Finally, PL prevented the formation of CPD (0% vs. 20% positive cells). In this model, PL has shown to prevent UVB cell damage, the upregulation of proliferating proteins, and fully blocking the formation of CPD.

Photoaging protective effects of BIOGF1K, a compound-K-rich fraction prepared from Panax ginseng

BackgroundBIOGF1K, a compound-K-rich fraction, has been shown to display anti-inflammatory activity. Although Panax ginseng is widely used for the prevention of photoaging events induced by UVB irradiation, the effect of BIOGF1K on photoaging has not yet been examined. In this study, we investigated the effects of BIOGF1K on UVB-induced photoaging events. MethodsWe analyzed the ability of BIOGF1K to prevent UVB-induced apoptosis, enhance matrix metalloproteinase (MMP) expression, upregulate anti-inflammatory activity, reduce sirtuin 1 expression, and melanin production using reverse transcription-polymerase chain reaction, melanin content assay, tyrosinase assay, and flow cytometry. We also evaluated the effects of BIOGF1K on the activator protein-1 signaling pathway, which plays an important role in photoaging, by immunoblot analysis and luciferase reporter gene assays. ResultsTreatment of UVB-irradiated NIH3T3 fibroblasts with BIOGF1K prevented UVB-induced cell death, inhibited apoptosis, suppressed morphological changes, reduced melanin secretion, restored the levels of type I procollagen and sirtuin 1, and prevented mRNA upregulation of MMP-1, MMP-2, and cyclo-oxygenase-2; these effects all occurred in a dose-dependent manner. In addition, BIOGF1K markedly reduced activator-protein-1-mediated luciferase activity and decreased the activity of mitogen-activated protein kinases (extracellular response kinase, p38, and C-Jun N-terminal kinase). ConclusionOur results strongly suggest that BIOGF1K has anti-photoaging activity and that BIOGF1K could be used in anti-aging cosmeceutical preparations.

Dual Protective Effects of Flavonoids from Petasites japonicus against UVB-Induced Apoptosis Mediated via HSF-1 Activated Heat Shock Proteins and Nrf2-Activated Heme Oxygenase-1 Pathways.

The leaves of Petasites japonicus are used for their anti-allergic properties in traditional Korean, Japanese, and Chinese medicine. This study aimed to identify bioactive compounds isolated from P. japonicus leaves. All compounds were assessed for their ability of transcriptional activation, induction of phase 2 enzymes and heat shock proteins (HSPs), as well as protection against the UVB-induced apoptotic cell death. Bioactive compounds were isolated from P. japonicus leaves. All compounds were evaluated for their protective effect using human dermal fibroblasts (HDF) and human epidermal keratinocyte cells (HEKC) treated with UVB radiation. Four flavonoids were isolated from the leaves of P. japonicus and identified as kaempferol-3-O-(6″-acetyl)-β-D-glucoside (1), quercetin-3-O-(6″-acetyl)-β-D-glucoside (2), kaempferol-3-O-β-D-glucoside (3), and quercetin-3-O-β-D-glucoside (4). These compounds activated nuclear factor (erythroid-derived 2)-like 2 (Nrf2) and heat-shock response transcription elements (HSE) that resulted in the induction of heme oxygenase-1 (HO-1) and HSP70, respectively. Activation of these pathways provided protection to the skin cells against UVB radiation. The isolated compounds activated the Nrf2 and HSE pathways and could protect against UVB-induced apoptosis.

Lippia graveolens photochemopreventive effect against UVB radiation-induced skin carcinogenesis

Lippia graveolens HBK (Mexican oregano) is a species that is regularly used as a condiment in Mexican cuisine. In traditional medicine, it is used for the treatment of respiratory and digestive illnesses, headaches, rheumatism and inflammation-related disorders. The main chemical components reported in this species include the following: terpenoids, iridoids and flavonoids. The aim of this study was to determine the potential photochemopreventive effect of the methanolic extract of Lippia graveolens (MELG) against ultraviolet B (UVB)-induced skin cancer in SKH-1 mice. The phenolic content, radical scavenger activity, penetration and genotoxicity of the MELG were also evaluated. The MELG exhibited scavenging activity against 1,1-diphenyl-2-picrylhydrazyl, superoxide and hydroxyl radicals, and it did not exhibit genotoxic activity in the micronucleus test. In addition, the MELG absorbed UVB (280nm) electromagnetic radiation. The main components detected in the plant extract were naringenin and galangin, and pinocembrin was also isolated and identified through spectroscopic analysis. The MELG demonstrated a photoprotective effect against UVB-induced cell death in Escherichia coli. In chronic challenge experiments, the MELG protected against UVB-induced skin cancer in SKH-1 mice. The MELG penetrated the skin of mice. Topical administration of the MELG protected against chronic UVB-induced damage in mouse SKH-1 skin. Our results suggest that the MELG has photochemopreventive activity and may potentially prevent photo-tumorigenesis.

Irradiance-dependent UVB Photocarcinogenesis.

Ultraviolet B (UVB) radiation from the sun may lead to photocarcinogenesis of the skin. Sunscreens were used to protect the skin by reducing UVB irradiance, but sunscreen use did not reduce sunburn episodes. It was shown that UVB-induced erythema depends on surface exposure but not irradiance of UVB. We previously showed that irradiance plays a critical role in UVB-induced cell differentiation. This study investigated the impact of irradiance on UVB-induced photocarcinogenesis. For hairless mice receiving equivalent exposure of UVB radiation, the low irradiance (LI) UVB treated mice showed more rapid tumor development, larger tumor burden, and more keratinocytes harboring mutant p53 in the epidermis as compared to their high irradiance (HI) UVB treated counterpart. Mechanistically, using cell models, we demonstrated that LI UVB radiation allowed more keratinocytes harboring DNA damages to enter cell cycle via ERK-related signaling as compared to its HI UVB counterpart. These results indicated that at equivalent exposure, UVB radiation at LI has higher photocarcinogenic potential as compared to its HI counterpart. Since erythema is the observed sunburn at moderate doses and use of sunscreen was not found to associate with reduced sunburn episodes, the biological significance of sunburn with or without sunscreen use warrants further investigation.

Faster DNA Repair of Ultraviolet-Induced Cyclobutane Pyrimidine Dimers and Lower Sensitivity to Apoptosis in Human Corneal Epithelial Cells than in Epidermal Keratinocytes.

Absorption of UV rays by DNA generates the formation of mutagenic cyclobutane pyrimidine dimers (CPD) and pyrimidine (6–4) pyrimidone photoproducts (6-4PP). These damages are the major cause of skin cancer because in turn, they can lead to signature UV mutations. The eye is exposed to UV light, but the cornea is orders of magnitude less prone to UV-induced cancer. In an attempt to shed light on this paradox, we compared cells of the corneal epithelium and the epidermis for UVB-induced DNA damage frequency, repair and cell death sensitivity. We found similar CPD levels but a 4-time faster UVB-induced CPD, but not 6-4PP, repair and lower UV-induced apoptosis sensitivity in corneal epithelial cells than epidermal. We then investigated levels of DDB2, a UV-induced DNA damage recognition protein mostly impacting CPD repair, XPC, essential for the repair of both CPD and 6-4PP and p53 a protein upstream of the genotoxic stress response. We found more DDB2, XPC and p53 in corneal epithelial cells than in epidermal cells. According to our results analyzing the protein stability of DDB2 and XPC, the higher level of DDB2 and XPC in corneal epithelial cells is most likely due to an increased stability of the protein. Taken together, our results show that corneal epithelial cells have a better efficiency to repair UV-induced mutagenic CPD. On the other hand, they are less prone to UV-induced apoptosis, which could be related to the fact that since the repair is more efficient in the HCEC, the need to eliminate highly damaged cells by apoptosis is reduced.

Protective effect of fermented Cyclopia intermedia against UVB-induced damage in HaCaT human keratinocytes.

BackgroundThe fermented leaves and stems of Cyclopia intermedia are used to brew honeybush tea, a herbal tea indigenous to South Africa. The aim of this study was to evaluate the protective effect of fermented honeybush extracts (FH ex) and scale-up fermented honeybush extracts (SFH ex) against ultraviolet B (UVB)-induced damage in HaCaT keratinocytes. To this end, we examined UVB-induced cell viability, antioxidant enzymes, and inflammatory mediators in HaCaT cells.MethodsUVB significantly decreased HaCaT cell viability, whereas FH ex and SFH ex did not exhibit cytotoxic effects and increased the viability of the HaCaT cells. To further investigate the protective effects of FH ex on UVB-induced oxidative stress in HaCaT cells, the activities of superoxide dismutase (SOD), catalase (CAT), matrix metalloproteinases (MMPs), pro-inflammatory cytokines and skin barrier function in terms of involucrin, filaggrin, and loricrin were analyzed.ResultsUVB-induced treatment reduced the activity of antioxidant enzymes and skin barrier function, while FH ex or SFH ex increased their activity. These results suggest that FH ex exerted cytoprotective activity against UVB-induced oxidative stress in HaCaT cells through stimulation of antioxidant enzymes activities. Furthermore, FH ex and SFH ex suppressed the UVB-induced expression of inflammatory mediators, such as IL-1β, IL-6, and IL-8, at mRNA level together with down regulation of matrix metalloproteinase (MMPs). In addition, FH ex and SFH ex reversed the phosphorylation of mitogen-activated protein kinase (MAPK) induced by UVB-irradiation. Notably, FH ex and SFH ex markedly inhibited UVB-induced activation of ERK, p38, and JNK. Thus, this agent exhibits anti-oxidative and -inflammatory effects via lowering ROS production, suppressing p38, ERK, and JNK activation, and down-regulating expression of MMPs.ConclusionsThese findings suggest that FH ex and SFH ex can be used as a skin anti-photoaging agent.

Sclareol isolated from Salvia officinalis improves facial wrinkles via an antiphotoaging mechanism.

Ultraviolet (UV) irradiation triggers skin photoaging processes, which disrupt the normal three-dimensional integrity of skin. UV-induced oxidative stress, both directly and indirectly, stimulates complex signaling pathways. UV radiation activates skin cell surface receptors on a molecular level and triggers severe changes in extracellular matrix (ECM) proteins, resulting in skin photoaging. Sclareol isolated from Salvia officinalis is widely used as a fragrance material. Sclareol is known to exert various biological activities, but its antiphotoaging effect has not been elucidated to date. Therefore, we evaluated wrinkle improvement efficacy of sclareol. Human dermal fibroblast cell line (Hs68) and a reconstructed human epidermis (RHE) model were used to evaluate the antiphotoaging effect of sclareol in vitro. A clinical study treated with 0.02% sclareol-containing cream was conducted to identify the ability of sclareol to improve wrinkles. First, sclareol enhanced cellular proliferation and blocked UVB-induced cell death. Sclareol inhibited the UVB-induced mRNA expression of matrix metalloproteinases (MMPs) by regulating the protein expression of AP-1 constituents. In RHE model, sclareol recovered the UVB-induced decrease in epidermal thickness and the expression of proliferating cell nuclear antigen (PCNA). In clinical trial, visually assessed changes and several wrinkle parameters were considered to be statistically different between the test and control groups at 12 weeks. In this study, sclareol inhibited various photoaging phenomena in human fibroblasts and RHE model. In addition, sclareol-containing cream improved wrinkles in a clinical trial. Taken together, sclareol alleviates facial wrinkle formation via an antiphotoaging mechanism and may be an effective candidate ingredient.

Abstract 2022: Twist1 regulates UVB-induced epidermal cell proliferation in non-melanoma skin cancer

Abstract Nonmelanoma skin cancer (NMSC) is the most common cancer and consists of basal cell carcinoma (BCC) and squamous cell carcinoma (SCC). SCC metastases account for up to 20% of all deaths from skin cancer. The major risk factor for the development of NMSC is solar ultraviolet radiation (UV). UVA and UVB both function as initiators and promoters in carcinogenesis and possess immunosuppressive activity. Given the prevalence of this disease, there remains a pressing need for the development of novel prevention and treatment strategies for NMSC. In a recent study (Srivastava et al. 2015. Mol. Carcinog. doi:10.1002/mc.22335), we evaluated the effect of Twist1, a transcription factor that is known to play a role in tumor progression, during epithelial carcinogenesis. Using the two-stage chemical carcinogenesis model, we identified that BK5.Cre x Twist1flox/flox mice, which have a keratinocyte-specific Twist1 knockout (Twist1 KO), developed significantly reduced numbers of papillomas (70% reduction) and SCCs (75% reduction) as well as delayed tumor latency compared to wild-type (WT) mice. Twist1 KO mice also exhibited significantly reduced epidermal proliferation in response to chemical promotion that correlated with reduced levels of cell cycle regulators and increased levels of p53 and p21. In further studies, we have begun analyzing the effects of UVB irradiation in Twist1 KO mice. Specifically, Twist1 KO mice exhibited heightened sensitivity to UVB-induced apoptosis and decreased epidermal proliferation as compared to similarly exposed WT mice. In addition, the accumulation of mast cells in the skin was significantly decreased in Twist1 KO mice as compared to WT mice. Further preliminary experiments showed that the protein expression levels of c-Myc, phospho-p65 (S536) and phospho-p50 (S337) decreased following UVB irradiation whereas levels of the cell cycle inhibitors p21 and p27 exhibited an increased expression in Twist1 KO mice as compared to WT mice. Further experiments have suggested that Twist1 may play a pivotal role in the proliferation/migration of bulge region keratinocyte stem cells (KSCs) following exposure to UVB. These and other results from ongoing experiments will be presented. Collectively, these findings suggest that Twist1 may have a novel role in UVB-induced epithelial carcinogenesis by regulating proliferation and migration of keratinocytes and KSCs. Research supported by CPRIT RP150346, CPRIT RP140108, and NIH T32 ES007247. Citation Format: Jaya Srivastava, Okkyung Rho, John DiGiovanni. Twist1 regulates UVB-induced epidermal cell proliferation in non-melanoma skin cancer. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2022.

Heat-mediated reduction of apoptosis in UVB-damaged keratinocytes in vitro and in human skin ex vivo.

BackgroundUV radiation induces significant DNA damage in keratinocytes and is a known risk factor for skin carcinogenesis. However, it has been reported previously that repeated and simultaneous exposure to UV and heat stress increases the rate of cutaneous tumour formation in mice. Since constant exposure to high temperatures and UV are often experienced in the environment, the effects of exposure to UV and heat needs to be clearly addressed in human epidermal cells.MethodsIn this study, we determined the effects of repeated UVB exposure 1 kJ/m2 followed by heat (39 °C) to human keratinocytes. Normal human ex vivo skin models and primary keratinocytes (NHEK) were exposed once a day to UVB and/or heat stress for four consecutive days. Cells were then assessed for changes in proliferation, apoptosis and gene expression at 2 days post-exposure, to determine the cumulative and persistent effects of UV and/or heat in skin keratinocytes.ResultsUsing ex vivo skin models and primary keratinocytes in vitro, we showed that UVB plus heat treated keratinocytes exhibit persistent DNA damage, as observed with UVB alone. However, we found that apoptosis was significantly reduced in UVB plus heat treated samples. Immunohistochemical and whole genome transcription analysis showed that multiple UVB plus heat exposures induced inactivation of the p53-mediated stress response. Furthermore, we demonstrated that repeated exposure to UV plus heat induced SIRT1 expression and a decrease in acetylated p53 in keratinocytes, which is consistent with the significant downregulation of p53-regulated pro-apoptotic and DNA damage repair genes in these cells.ConclusionOur results suggest that UVB-induced p53-mediated cell cycle arrest and apoptosis are reduced in the presence of heat stress, leading to increased survival of DNA damaged cells. Thus, exposure to UVB and heat stress may act synergistically to allow survival of damaged cells, which could have implications for initiation skin carcinogenesis.Electronic supplementary materialThe online version of this article (doi:10.1186/s12895-016-0043-4) contains supplementary material, which is available to authorized users.

Retinoid X Receptor Agonists Upregulate Genes Responsible for the Biosynthesis of All-Trans-Retinoic Acid in Human Epidermis.

UAB30 is an RXR selective agonist that has been shown to have potential cancer chemopreventive properties. Due to high efficacy and low toxicity, it is currently being evaluated in human Phase I clinical trials by the National Cancer Institute. While UAB30 shows promise as a low toxicity chemopreventive drug, the mechanism of its action is not well understood. In this study, we investigated the effects of UAB30 on gene expression in human organotypic skin raft cultures and mouse epidermis. The results of this study indicate that treatment with UAB30 results in upregulation of genes responsible for the uptake and metabolism of all-trans-retinol to all-trans-retinoic acid (ATRA), the natural agonist of RAR nuclear receptors. Consistent with the increased expression of these genes, the steady-state levels of ATRA are elevated in human skin rafts. In ultraviolet B (UVB) irradiated mouse skin, the expression of ATRA target genes is found to be reduced. A reduced expression of ATRA sensitive genes is also observed in epidermis of mouse models of UVB-induced squamous cell carcinoma and basal cell carcinomas. However, treatment of mouse skin with UAB30 prior to UVB irradiation prevents the UVB-induced decrease in expression of some of the ATRA-responsive genes. Considering its positive effects on ATRA signaling in the epidermis and its low toxicity, UAB30 could be used as a chemoprophylactic agent in the treatment of non-melanoma skin cancer, particularly in organ transplant recipients and other high risk populations.

Ultraviolet B (UVB) Induces Down-regulation of Parkin Gene Expression

Ultraviolet (UV) irradiation induces cellular damage. A variety of cellular responses for repairing cellular damage including DNA damage occur after UV irradiation. During the repair processes, expression and activation of various molecules are regulated depending on the types of cellular damage. Parkin is an E3 ligase and act as a tumor suppressor. Recently, it has been reported that Parkin is involved in the DNA repair process. In the current study, we investigated whether UVB irradiation influences expression of Parkin. Parkin expression transiently decreased after UVB irradiation both at the mRNA and protein levels, but returned to normal levels thereafter. Taken together with cell viability data, Parkin expression is down-regulated during UVB-induced suppression of cell growth and is increased again in accordance with recovery of UVB-induced cell growth inhibition. However, Parkin overexpression or knockdown did not influence UVB-induced cell growth inhibition and recovery. We propose that Parkin could be a useful molecular marker for evaluating conditions of cells after UVB irradiation.

Inhibition of mTOR by apigenin in UVB-irradiated keratinocytes: A new implication of skin cancer prevention

Ultraviolet B (UVB) radiation is the major environmental risk factor for developing skin cancer, the most common cancer worldwide, which is characterized by aberrant activation of Akt/mTOR (mammalian target of rapamycin). Importantly, the link between UV irradiation and mTOR signaling has not been fully established. Apigenin is a naturally occurring flavonoid that has been shown to inhibit UV-induced skin cancer. Previously, we have demonstrated that apigenin activates AMP-activated protein kinase (AMPK), which leads to suppression of basal mTOR activity in cultured keratinocytes. Here, we demonstrated that apigenin inhibited UVB-induced mTOR activation, cell proliferation and cell cycle progression in mouse skin and in mouse epidermal keratinocytes. Interestingly, UVB induced mTOR signaling via PI3K/Akt pathway, however, the inhibition of UVB-induced mTOR signaling by apigenin was not Akt-dependent. Instead, it was driven by AMPK activation. In addition, mTOR inhibition by apigenin in keratinocytes enhanced autophagy, which was responsible, at least in part, for the decreased proliferation in keratinocytes. In contrast, apigenin did not alter UVB-induced apoptosis. Taken together, our results indicate the important role of mTOR inhibition in UVB protection by apigenin, and provide a new target and strategy for better prevention of UV-induced skin cancer.

Extract of Ettlia sp. YC001 Exerts Photoprotective Effects against UVB Irradiation in Normal Human Dermal Fibroblasts.

The identification of novel reagents that exert a biological ultraviolet (UV)-protective effect in skin cells represents an important strategy for preventing UV-induced skin aging. To this end, we investigated the potential protective effects of Ettlia sp. YC001 extracts against UV-induced cellular damage in normal human dermal fibroblasts (NHDFs). We generated four different extracts from Ettlia sp. YC001, and found that they exhibit low cytotoxicity in NHDFs. The ethyl acetate extract of Ettlia sp. YC001 markedly decreased UVB-induced cytotoxicity. Additionally, the ethyl acetate extract significantly inhibited the production of hydrogen peroxide-induced reactive oxygen species. Moreover, it inhibited UVB-induced thymine dimers, as confirmed by luciferase assay and thymine dimer dot-blot assay. Thus, the study findings suggest Ettlia sp. YC001 extract as a novel photoprotective reagent on UVB-induced cell dysfunctions in NHDFs.

Abstract 1915: Effects of carvedilol, a β-adrenergic receptor antagonist, on UVB-induced skin cancer development and its possible mechanism of action

Abstract Recently, we proposed that carvedilol, a β-adrenergic receptor antagonist (i.e., β-blocker), can act as a chemopreventive agent against skin cancer (Chang et al., Cancer Prev. Res, 2014). Since approximately 90 percent of diagnosed non-melanoma skin cancers are associated with exposure to ultraviolet (UV) radiation from the sun, the primary form being UVB irradition, we hypothesized that carvedilol will prevent UVB-induced skin carcinogenesis. The transformable JB6 P+ cell line, an established model for studying skin cancer promotion, was used for evaluating the impact of carvedilol on UVB-induced oxidative damage. Exposures of JB6 P+ cells to 50 - 400 mJ/cm2 UVB resulted in a dose-dependent decrease in cell viability. 25 mJ/cm2 UVB induced ∼20% of cell death, and thus was chosen to study the effect of carvedilol. 0.1 to 1.0 μM carvedilol prevented UVB-induced growth inhibition and apoptosis; however, 3 to 5 μM carvedilol, although nontoxic alone, enhanced UVB-induced cell death. Contrarily, all concentrations of carvedilol tested blocked H2O2-dependent cytotoxicity of JB6 P+ cells. Thus, although both UVB and H2O2 are classic inducers of oxidative stress resulting in DNA damage and tumor promotion, carvedilol may modulate UVB and H2O2-mediated effects via different mechanisms. Cell signaling studies revealed that carvedilol inhibited EGF-induced phosphorylation of AKT at S473 and GSK3β at S9 as well as AP-1 transactivation, which are all well-known principal mediators of skin carcinogenesis. A pitfall with experiments involving UV exposure is the potential UV absorption properties of test compounds. Examination of the structure of carvedilol indicates that it will absorb UV due to the presence of a carbazole moiety and benzene rings. If not controlled in the experiments, the observed effects will be a mixture of pharmacological properties and a UV absorption effect. To identify a solution for this problem, 4-hydroxycarbazole, which mimicks the carbazole moiety of carvedilol, was found to have the same UVB absorption properties as carvedilol but does not function as a β-adrenergic receptor antagonist nor inhibit EGF-mediated colony formation of JB6 P+ cells. 4-hydroxycarbazole is currently being used to repeat the UVB experiments mentioned above to act as a sunscreen/absorption control. Additionally, 4-hydroxycarbazole should be used as a control for carvedilol for other on-going studies for prevention of UVB-induced skin cancer. Since 4-hydroxycarbazole only has the UV absorption (i.e., sunscreen) effect, we anticipate that it will have some protective effect, but that carvedilol provides greater preventive activity against UVB-induced skin carcinogenesis. Since carvedilol and other β-blockers are already FDA-approved drugs, these results may be readily translated into clinical trials as a new approach for skin cancer chemoprevention. Citation Format: Kevin M. Huang, Andy Chang, Kristan H. Cleveland, Bradley T. Andresen, Ying Huang. Effects of carvedilol, a β-adrenergic receptor antagonist, on UVB-induced skin cancer development and its possible mechanism of action. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 1915. doi:10.1158/1538-7445.AM2015-1915

Abstract A20: Deregulation of cell cycle and apoptotic mechanisms in UVB-irradiated p16-mutant inducible melanoma cell lines

Abstract Melanoma is the deadliest skin cancer as it progresses to a highly aggressive and metastatic tumor during melanomagenesis. However, its underlying mechanisms regulating melanoma-genesis remain nascent. Our previous studies have demonstrated that the p53 and p16INK4a sub-pathways components of the Rb-signaling pathway, plays a critical role in UVB-induced skin cancer. The aim of this study was to study the molecular changes in the expression/function of the different components of the Rb signaling pathway associated with UVB-induced cell cycle and apoptotic programs. Pursuant to this goal, normal melanocytes, and a panel of p16-induced cell lines (wild-type p16INK4a-inducible clone p16wt-A4, and the N-terminal 8 amino-acid duplication p16INK4a-inducible clone p16Dp1 (+24 base pairs) were used to elucidate the early molecular events especially associated with the p16INK4a/Rb-signaling pathway regulating cell-cycle and apoptotic responses. Our results showed that: 1) in the menlanocytes high UVB dose induced p16 but not p53, and the Bax/Bcl2 ratio was also increased, suggesting that the cells had selected to undergo apoptosis in a p53-independent manner; 2) In the wt-p16 melanoma cells however, while low UVB irradiation upregulated p16, high dose reduced p16 expression and abrogated Cdk6 but not Cdk4; 3) Interestingly, mutant-p16 melanoma cells over-expressed cell cycle proteins Cdk4/6 and pRb proteins upon p16 induction, however, UVB exposure did not affect this increase; and 4) The anti-apoptotic properties of mutant cells were predominant at high dose of exposure associated with a decrease in Bax and an increase in Bcl2 expression. These data put together, suggest that the mutant p16 dictate cell cycle deregulation and causes resistance of melanoma cells to apoptosis. In conclusion, our study shows that UVB radiation-induced deregulation of p16INK4a/Cdk4 sub-pathway is a critical early event in melanomagenesis. Citation Format: Allal Ouhtit, Ishita Gupta, Zakariya Y. Abd Elmageed, Therese M. Becker. Deregulation of cell cycle and apoptotic mechanisms in UVB-irradiated p16-mutant inducible melanoma cell lines. [abstract]. In: Proceedings of the AACR Special Conference on Advances in Melanoma: From Biology to Therapy; Sep 20-23, 2014; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(14 Suppl):Abstract nr A20.