DMBA-induced Skin Carcinogenesis Research Articles

Nobiletin-loaded composite penetration enhancer vesicles restore the normal miRNA expression and the chief defence antioxidant levels in skin cancer

Skin cancer is one of the most dangerous diseases, leading to massive losses and high death rates worldwide. Topical delivery of nutraceuticals is considered a suitable approach for efficient and safe treatment of skin cancer. Nobiletin; a flavone occurring in citrus fruits has been reported to inhibit proliferation of carcinogenesis since 1990s, is a promising candidate in this regard. Nobiletin was loaded in various vesicular systems to improve its cytotoxicity against skin cancer. Vesicles were prepared using the thin film hydration method, and characterized for particle size, zeta potential, entrapment efficiency, TEM, ex-vivo skin deposition and physical stability. Nobiletin-loaded composite penetration enhancer vesicles (PEVs) and composite transfersomes exhibited particle size 126.70 ± 11.80 nm, 110.10 ± 0.90 nm, zeta potential + 6.10 ± 0.40 mV, + 9.80 ± 2.60 mV, entrapment efficiency 93.50% ± 3.60, 95.60% ± 1.50 and total skin deposition 95.30% ± 3.40, 100.00% ± 2.80, respectively. These formulations were selected for cytotoxicity study on epidermoid carcinoma cell line (A431). Nobiletin-loaded composite PEVs displayed the lowest IC50 value, thus was selected for the in vivo study, where it restored skin condition in DMBA induced skin carcinogenesis mice, as delineated by histological and immuno-histochemical analysis, biochemical assessment of skin oxidative stress biomarkers, in addition to miRNA21 and miRNA29A. The outcomes confirmed that nobiletin- loaded composite PEVs is an efficient delivery system combating skin cancer.

Effect of nisin and doxorubicin on DMBA-induced skin carcinogenesis--a possible adjunct therapy.

In view of the emergence of multidrug-resistant cancer cells, there is a need for therapeutic alternatives. Keeping this in mind, the present study was aimed at evaluating the synergism between nisin (an antimicrobial peptide) and doxorubicin (DOX) against DMBA-induced skin carcinogenesis. The possible tumoricidal activity of the combination was evaluated in terms of animal bioassay observations, changes in hisotological architecture of skin tissues, in situ apoptosis assay (TUNEL assay) and in terms of oxidant and antioxidant status of the skin tissues. In vivo additive effect of the combination was evidenced by larger decreases in mean tumour burden and tumour volume in mice treated with the combination than those treated with the drugs alone. Histological observations indicated that nisin-DOX therapy causes chromatin condensation and marginalisation of nuclear material in skin tissues of treated mice which correlated well with the results of TUNEL assay wherein a marked increase in the rate of apoptosis was revealed in tissues treated with the combination. A slightly increased oxidative stress in response to the adjunct therapy as compared to dox-alone-treated group was revealed by levels of lipid peroxidation (LPO) and nitrite generation in skin tissue-treated mice. An almost similar marginal enhancement in superoxide dismutase levels corresponding with a decrease in catalase activity could also be observed in nisin + DOX-treated groups as compared to nisin and dox-alone-treated groups. These results point towards the possible use of nisin as an adjunct to doxorubicin may help in developing alternate strategies to combat currently developing drug resistance in cancer cells.

In Vivo Suppression of Heat Shock Protein (HSP)27 and HSP70 Accelerates DMBA-Induced Skin Carcinogenesis by Inducing Antigenic Unresponsiveness to the Initiating Carcinogenic Chemical.

Heat shock proteins (HSPs) are constitutively expressed in murine skin. HSP27 is present in the epidermis, and HSP70 can be found in both the epidermis and dermis. The purpose of this study was to investigate the role of these proteins in cutaneous chemical carcinogenesis and to determine whether their effects on cell-mediated immune function were a contributing factor. In vivo inhibition of HSP27 and HSP70 produced a reduction in the T cell-mediated immune response to 7,12-dimethylbenz(a)anthracene (DMBA) and benzo(a)pyrene in C3H/HeN mice and resulted in a state of Ag-specific tolerance. When mice were pretreated with anti-HSP27 and anti-HSP70 Abs in vivo prior to subjecting them to a standard two-stage DMBA/12-O-tetradecanoylphorbol-13-acetate cutaneous carcinogenesis protocol, the percentage of mice with tumors was much greater (p < 0.05) in anti-HSP27- and HSP70-pretreated animals compared with mice pretreated with control Ab. Similar results were obtained when the data were evaluated as the cumulative number of tumors per group. Mice pretreated with HSP27 and HSP70 Abs developed more H-ras mutations and fewer DMBA-specific cytotoxic T lymphocytes. These findings indicate that in mice HSP27 and HSP70 play a key role in the induction of cell-mediated immunity to carcinogenic polyaromatic hydrocarbons. Bolstering the immune response to carcinogenic polyaromatic hydrocarbons may be an effective method for prevention of the tumors that they produce.

Inflammation-driven carcinogenesis is mediated through STING.

Chronic stimulation of innate immune pathways by microbial agents or damaged tissue is known to promote inflammation-driven tumorigenesis by mechanisms that are not well understood. Here we demonstrate that mutagenic 7,12-dimethylbenz(a)anthracene (DMBA), cisplatin and etoposide induce nuclear DNA leakage into the cytosol that intrinsically activates stimulator of interferon genes (STING)-dependent cytokine production. Inflammatory cytokine levels are subsequently augmented in a STING-dependent extrinsic manner by infiltrating phagocytes purging dying cells. Consequently, STING(-/-) mice, or wild-type mice adoptively transferred with STING(-/-) bone marrow, are almost completely resistant to DMBA-induced skin carcinogenesis compared with their wild-type counterparts. Our data establish a role for STING in the control of cancer, shed significant insight into the causes of inflammation-driven carcinogenesis and may provide a basis for therapeutic strategies to help prevent malignant disease.

Chemopreventive potential of Apium leptophyllum (Pers.) against DMBA induced skin carcinogenesis model by modulatory influence on biochemical and antioxidant biomarkers in Swiss mice.

Aim:The study was designed to investigate the chemopreventive potential of flavonoidal fractions of Apium leptophyllum fruits (FFALF) on Swiss mice.Materials and Methods:Skin tumor or papilloma was developed by topical application of DMBA (25 μg in 0.1 ml acetone) on intrascapular region of mice, twice weekly for 8 weeks. The animals were divided into six groups: Group I (vehicle control); group II (FFALF control, 5 mg/kg); group III (carcinogenic control, DMBA treated initially for 8 weeks); and group IV, V and VI as pre-treated group (FFALF 5, 10 and 20 mg/kg respectively for 16 weeks along with DMBA treatment). After the 16th week of treatment; the tumor morphology, skin histopathology, and biochemical and antioxidant biomarkers were measured and compared with carcinogenic control as well as vehicle control.Results:The co-administration of FFALF with DMBA-treated groups showed significant (P ≤ 0.001) prevention against skin papilloma and normalized the status of lipid peroxidation with antioxidant biomarkers in a dose-dependent manner as compared to carcinogenic control.Conclusions:Thus, the present study suggests that the FFALF is non-carcinogenic and has chemopreventive potential on DMBA-induced carcinogenesis in mouse, which may be due to the modulation of cutaneous lipid peroxidation or enhancement of total antioxidant capacity.

Abstract 5588: Szygium aromaticum (clove): A novel natural breast cancer inhibitor

Abstract The dried flower buds of clove (Syzygium aromaticum syn Eugenia aromaticum or caryophyllata) belonging to family Myrtaceae are used as a spice worldwide. Clove has been reported to exhibit antimicrobial, anti-viral, anesthetic, insecticidal, anti-convulsant, anti-oxidative and free radical scavenging activities. Eugenol, an active component of essential oil isolated from clove has anti-mutagenic, anti-genotoxic and anti-inflammatory properties. It also restricts DMBA induced skin carcinogenesis in mice. Aqueous extract of clove was effective in reducing the incidence of hyperplasia, dysplasia, and carcinoma in situ in benzo[a]pyrene induced lung carcinoma in mice. However, there are no reports on the effects of clove on human cancers. In this study we have analyzed the inhibitory effects of the aqueous extract of clove (CAE) on breast cancer using in vitro studies like cell growth, cell migration, three-dimensional capillary tube formation / morphogenesis and apoptosis; and in vivo studies like breast tumor growth in nude mice. Our results indicate that CAE inhibited the growth and proliferation of breast cancer cell lines MDA-MB-231, MCF10ADCIS.com, and MCF10AEIII 8 cells in a dose dependent manner. In addition, it also reduced cell migration towards Matrigel. CAE also inhibited the enzymatic activity of the Matrix Metalloproteinase-2 (MMP-2), which was analyzed by using two substrates, namely galectin-3 and a fluorescence-quenched substrate MOCAcPLGLA2pr(Dnp)AR-NH2. Cell cycle analysis of MCF10ADCIS.com cells using flow cytometry after propidium iodide (PI) staining indicated an arrest mainly in S-phase. We also observed an inhibition of DNA synthesis in CAE-treated MCF10ADCIS.com cells by bromodeoxyuridine (BrdU) incorporation. Expressions of apoptosis-related proteins, poly(ADP-ribose)polymerase (PARP), caspase-3 and caspase-8 remained unchanged after CAE treatment, but CDK2, CDK4, cyclin A and cyclin D2 were down-regulated in treated cells indicating that CAE might affect tumor growth by inducing cell cycle arrest but not apoptosis. There was a 50% reduction in tumor volume in nude mice injected with MCF10ADCIS.com cells that received CAE extract in their drinking water compared to the control mice. The expression of proliferation, apoptosis, angiogenesis and tumor progression markers (PCNA, TUNEL, p63, CD31, smooth muscle actin, collagen IV, MMP-2 are being investigated in the xenografts. The results suggest that CAE may be explored further as a novel inhibitor of breast cancer. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5588. doi:10.1158/1538-7445.AM2011-5588

Elimination of Deleterious Effects of DMBA-Induced Skin Carcinogenesis in Mice by Syzygium cumini Seed Extract

The inhibition of tumor incidence by hydro-alcoholic extract of S.cumini seed was evaluated in mice on two stage process of skin carcinogenesis induced by single application of 7, 12-dimethyl benz(a)anthracene (100 µg/100µl of acetone), and 2 weeks later promoted by repeated application of croton oil (1% acetone/thrice in a week) till the end of the experiment (i.e. 16 weeks). Oral administration of extract at a dose of 250mg/kg b.wt./day at the peri-initiational stage (i.e. 7 days before and 7 days after DMBA application), promotional stage (i.e. from the time of croton oil application) and at both the stages (i.e. 7 days prior to DMBA application & continued till the end of experiment) to the mice, recorded a significant reduction in tumor incidence to 37.5, 50 & 25% respectively in comparison to the carcinogen treated control, where tumor incidence was found as 100%. Tumor yield and Tumor burden were also significantly reduced by SCE. Similarly, the cumulative number of papillomas after 16 weeks was 68 in the control group, which was reduced to 15, 21 & 8 in the animals treated with the SCE continuously at peri-, post- and peri- & post- initiation stage respectively. A significant impairment was noticed in the levels of reduced glutathione, superoxide dismutase, catalase & protein and enhancement in LPO in liver and skin of carcinogen treated control mice as compared with vehicle treated mice. All such parameters were returned to near normal value by administration of SCE to DMBA treated mice. These results suggest a possible chemopreventive property of S.cumini against DMBA induced skin carcinogenesis in mice.

Abstract 2880: Spices suppress oxidative stress during DMBA induced skin carcinogenesis in mice, mimicking human non-melanoma skin cancer

Abstract Background: Cancer chemoprevention is the use of natural, synthetic or biologic substances to reverse, suppress, or prevent the development of cancer. Spices like cinnamon, cardamom and cumin are the naturally occurring phytoproducts that display an active cancer preventive strategy to inhibit, delay or reverse the human carcinogenesis. Aims and Objectives: Our aim of this study is to decipher the chemopreventive effect of aqueous infusion of the spices as a diet on DMBA induced skin carcinogenesis, which mimics UV radiation mediated non melanoma skin cancer that has high mortality rate if not diagnosed early. Research Design and Methodology: Female Swiss Albino Mice have been divided into 5 groups in each case. Normal control (NC) and DMBA treated control (CC) mice received acetone and DMBA/croton oil on it's dorsally shaven skin for eight weeks, respectively. Aqueous suspensions of the spices were orally administered to three groups of mice either before or after or both before and after DMBA applications on the shaved skin during the entire study period. Treated skins were excised after harvesting the mice and H &amp; E staining, immunohistochemistry assays for lipid peroxides, antioxidation enzymes activities, immunoblotting, cell proliferation and apoptosis were performed. Results: All the spices demonstrate inhibition of the skin papillomagenesis. The best chemopreventive actions of these spices were observed when the spice treatments were performed both before and after the induction of skin carcinogenesis by DMBA. An up-regulation of the phase II detoxification enzymes and induction of non-protein thiol antioxidants were observed in mice treated with the spices. Additionally, cumin and cardamom demonstrate downregulation of COX-2 expression, inhibit cell proliferation and block p53-induced apoptosis. Cardamom stimulates NRF2 and inhibits NF-κB transcription factors to demonstrate its beneficial action against skin carcinogenesis. Conclusion: Spices inhibit DMBA induced skin carcinoma in mice when treated early and it is believed to be due, at least in part, to the induction of cellular defense systems. We are in a process of identifying unique chemical components of these spices to maximize the benefits.

Abstract A64: Chemopreventive potential of Bauhinia variegata against DMBA-induced skin papillomagenesis in Swiss albino mice

Abstract The present study is an attempt to evaluate the cancer chemopreventive potential of Bauhinia variegata bark against DMBA-induced skin papillomagenesis in mice. We investigated chemopreventive effect of three different doses (2%, 4% and 6% w/w) of test diets of bark of B. variegata during DMBA-induced skin papillomagenesis in mice and tumor cell proliferation and apoptosis. Its effect was also studied in a short-term animal protocol on carcinogen metabolizing enzymes in liver. DMBA and croton oil was used to induce and promote skin papillomagenesis, respectively. Skin papillomagenesis study was done by using three different doses of B. variegata before, during and after the DMBA treatment to mice till termination of the experiment (120 days). In addition, the expression of proliferation [proliferating cell nuclear antigen (PCNA), glutathione-S-transferse (GST-π) and apoptosis were analyzed in the skin tumors by immunohistochemistry. For biochemical studies, mice were given with the same doses of B. variegata in the test diet for two weeks. The specific activities/levels of phase-I and phase-II enzymes, antioxidant enzymes, lactate dehydrogenase, glutathione content, and peroxidative damage were determined in the liver of the mice. Results showed a significant reduction in the skin tumors in B. variegata treated animals, as compared to the control group of animals. The tumor incidence in case of control group of animals was 100% whereas it was 90%, 70% and 99% in groups treated with 2%, 4% and 6% test diets, respectively. Tumor multiplicity in control group was 12±3.9 whereas it was reduced to 8±3.7, 3±2.9 and 4±2.4 by the lower, intermediate and higher dose of B. variegata treatment. The dietary administration of three different doses of B. variegata significantly decreased the expression of PCNA, GST-π, Bcl-2 and VEGF and on the other hand significantly increased the expression of Bax and total Caspase 3 as compared to DMBA and croton oil treated group of animals. The effect of 2 weeks of dietary feeding of B. variegata inhibited phase I, activated phase II and antioxidant enzymes in the mouse liver. The content of reduced glutathione was significantly elevated whereas the peroxidative damage and specific activity of lactate dehydrogenase exhibited a significant reduction with all the doses of test diets, respectively. All these findings were indicative of that Bauhinia variegata has chemopreventive potential against DMBA-induced skin carcinogenesis. It would be of interest to investigate for active cancer chemopreventive agents in Bauhinia variegata for its future implication in cancer prevention and control. Citation Information: Cancer Prev Res 2010;3(1 Suppl):A64.

Chemopreventive potential of piperine in 7,12-dimethylbenz[a]anthracene-induced skin carcinogenesis in Swiss albino mice

The chemopreventive potential of orally administered piperine was studied in Swiss albino mice against 7,12-dimethylbenz[a]anthracene (DMBA)-induced skin carcinogenesis. The mechanistic pathway for the chemopreventive potential of piperine was evaluated by analysing the status of phase I and phase II detoxification agents, lipid peroxidation by-products and antioxidants during DMBA-induced skin carcinogenesis. Skin squamous cell carcinoma was induced in the shaved back of mice, by painting with DMBA (25 μg in 0.1 ml acetone/mouse) two times weekly for 8 weeks. We observed severe hyperplasia, dysplasia, and well-differentiated squamous cell carcinoma in the 8th, 10th and 15th week of experimental period respectively in mice treated with DMBA alone. Marked alterations in the status of phase I and phase II detoxification agents, lipid peroxidation by-products and antioxidants were observed in tumor bearing mice. Oral administration of piperine (50 mg kg −1 body weight) by gastric gavage significantly prevented the formation of skin tumors during DMBA-induced mouse skin carcinogenesis. Also, piperine administration brought back the status of phase I and phase II detoxification agents, lipid peroxidation by-products and antioxidants to near normal range in DMBA treated mice. The present study thus demonstrates that piperine has significant suppressing effect on cell proliferation during DMBA-induced mouse skin carcinogenesis. The chemopreventive potential of piperine is probably due to its modulating effect on the status of lipid peroxidation, antioxidants and detoxification agents during DMBA-induced skin carcinogenesis.

Resveratrol enhances cell‐mediated immune response to DMBA through TLR4 and prevents DMBA induced cutaneous carcinogenesis

Toll-like receptors (TLRs) activate signals that are critically involved in innate immune responses and that contribute to the initiation of adaptive immune responses. Resveratrol (trans-3,5,4-trihydroxystilbene), a polyphenol found in red grapes and in several other plant sources, is an effective chemopreventive agent in cutaneous chemical carcinogenesis. In this study, we investigated whether TLR4 was required for the chemopreventive action of resveratrol in DMBA skin carcinogenesis. For this purpose, mice with normal and deficient TLR4 function were compared when pretreated with resveratrol and then subjected to a DMBA-induced skin carcinogenesis protocol. There were fewer tumors/group (P < 0.001) in resveratrol treated TLR4 competent C3H/HeN mice than in TLR4 deficient C3H/HeJ mice. In addition, the size of tumors in C3H/HeN mice was reduced in vivo and their survival in vitro was inhibited by resveratrol to a significantly greater extent than in C3H/HeJ mice. Resveratrol inhibited angiogenesis to a much greater extent in the TLR4 competent mice than in TLR4 deficient mice. IFN-gamma and IL-12 levels were also increased in TLR4 competent mice compared to TLR4 deficient mice, and TLR4 competent C3H/HeN mice exhibited a greater increase in the cell-mediated immune response to DMBA. The results of this study indicate that TLR4 is an important mediator of resveratrol chemoprevention in DMBA skin tumorigenesis.

Protective effect of ferulic acid on 7,12-dimethylbenz[a]anthracene-induced skin carcinogenesis in Swiss albino mice

Our aim was to evaluate and compare the chemopreventive potential of topically applied and orally administered ferulic acid in 7,12-dimethylbenz[a]anthracene (DMBA)-induced skin carcinogenesis. Estimating the status of phase I and phase II detoxication agents, lipid peroxidation byproducts and antioxidants during DMBA-induced skin carcinogenesis assessed the mechanistic pathway for its chemopreventive efficacy. Skin squamous cell carcinoma was induced in the shaved back of mice, by painting with DMBA (25 μg in 0.1 mL −1 acetone) twice weekly for 8 weeks. We have observed 100% tumor formation in the 15th week of experimental period in mice treated with DMBA alone. Marked alterations in the status of phase I and phase II detoxication agents, lipid peroxidaton byproducts and antioxidants were observed in tumor bearing mice. Oral administration of ferulic acid completely prevented the formation of skin tumors, whereas topically applied ferulic acid did not show significant chemopreventive activity during DMBA-induced mouse skin carcinogenesis. Also, oral administration of ferulic acid reverted the status of phase I and phase II detoxication agents, lipid peroxidaton byproducts and antioxidants to near-normal range in DMBA-treated mice. Our results thus demonstrate that orally administered ferulic acid has potent suppressing effect on cell proliferation during DMBA-induced skin carcinogenesis. This is probably due to its modulating effect on the status of lipid peroxidation, antioxidants and detoxication agents during DMBA-induced skin carcinogenesis.

Chemopreventive and Antilipidperoxidative Potential of Clerodendron inerme (L) Gaertn in 7,12-dimethylbenz(a)anthracene Indcued Skin Carcinogenesis in Swiss Albino Mic

The present study has investigated the chemopreventive and antilipidperoxidative effects of the ethanolic extract of Clerodendron inerme leaves (CiELet) in DMBA induced skin carcinogenesis in Swiss albino mice. The skin squamous cell carcinoma was induced in the shaved back of mice, by painting with DMBA (25 microg 0.1 mL(-1) acetone) twice weekly for 8 weeks. We have observed 100% tumor formation in the fifteenth week of experimental period. Elevated lipid peroxidation and decline in enzymatic and non-enzymatic antioxidants status was observed in tumor bearing mice. Oral administration of CiELet (300 mg kg(-1) bw) for 25 weeks significantly prevented the tumor incidence, volume and burden of the tumor. The CiELet also showed potent antilipidperoxidative effect as well as enhanced the antioxidant defense mechanisms in DMBA painted mice. The present study thus demonstrated the chemopreventive and antilipidperoxidative efficacy of CiELet in DMBA induced mouse skin carcinogenesis.

PRAK Is Essential for ras-Induced Senescence and Tumor Suppression

Like apoptosis, oncogene-induced senescence is a barrier to tumor development. However, relatively little is known about the signaling pathways mediating the senescence response. p38-regulated/activated protein kinase (PRAK) is a p38 MAPK substrate whose physiological functions are poorly understood. Here we describe a role for PRAK in tumor suppression by demonstrating that PRAK mediates senescence upon activation by p38 in response to oncogenic ras. PRAK deficiency in mice enhances DMBA-induced skin carcinogenesis, coinciding with compromised senescence induction. In primary cells, inactivation of PRAK prevents senescence and promotes oncogenic transformation. Furthermore, we show that PRAK activates p53 by direct phosphorylation. We propose that phosphorylation of p53 by PRAK following activation of p38 MAPK by ras plays an important role in ras-induced senescence and tumor suppression.

Inhibitory effects of Azadirachta indica on DMBA-induced skin carcinogenesis in Balb/c mice

Male Balb/c mice were divided into four groups on the basis of their respective treatments wherein mice of Group I served as controls. For induction of skin tumors, mice of Group II and IV were injected sub-cutaneously with 7,12-dimethylbenz(a)anthracene (DMBA). Mice of Group III and IV were administered aqueous Azadirachta indica leaf extract (AAILE) thrice a week throughout the experiment. After 14 weeks of the first DMBA injection, Group II and IV mice developed tumors. In the tumor-bearing mice that received AAILE (Group IV), a significant reduction in mean tumor burden and tumor volume was observed. The tumors were confirmed to be papillomas and interestingly, the extent of hyper-chromatia was observed to be much more in skin tumors of Group II mice vis a vis the mice receiving AAILE. An increase in the extent of lipid peroxidation was observed in tumorous tissue of Group IV when compared to that of Group II mice. Glutathione (GSH) content and the activities of GSH-based antioxidant enzymes viz. glutathione peroxidase (GPx) and glutathione reductase (GR) increased significantly in the skin tissues of all the groups of mice when compared to control counterparts. Catalase activity was found to decrease significantly in the skin of mice, which received AAILE treatment only (Group III). Activity of super-oxide dismutase (SOD) decreased significantly in all the tumorous tissues (Group II and IV mice). In light of the above observations, the role of AAILE in inhibition of DMBA-induced skin carcinogenesis is discussed in the present study.

Effect of radiofrequency radiation exposure on mouse skin tumorigenesis initiated by 7,12-dimethybenz[α]anthracene

Purpose: Although radiofrequency (RF) radiation is not considered mutagenic, it has been suggested as a promoter of tumorigenesis. To study if RF radiation has a tumor promoting effect, we exposed mice with skin tumorigenesis initiated by 7,12-dimethybenz[α]anthracene (DMBA) to RF radiation.Materials and methods: Eighty male ICR mice were subjected to a single DMBA application (100 μg/100 μl acetone/mouse) on shaved dorsal skin at the age of 7 weeks. After one week, the mice were randomized into four equal groups of 20 mice each: i.e., sham-, 849 MHz-, 1,763 MHz-exposed, and 12-O-tetradecanoylphorbol-13-acetate (TPA)-treated groups. The RF exposure was conducted at a whole body average specific absorption rate (SAR) of 0.4 W/Kg, for 2 cycles of 45 min exposure with a 15 min interval each day, 5 days a week for 19 weeks. The TPA-treated group served as a positive control for skin tumorigenesis and were administered TPA (4 μg/100 μl acetone/mouse) twice weekly without RF exposure.Results: All mice were examined weekly at a macroscopic level. No skin tumors were observed in any groups except in the TPA-treated positive control group. TPA is known tumor promoter in DMBA-induced skin carcinogenesis and tumor incidence in the TPA treated group was 95%. At week 20 after DMBA initiation, skin tissues were analyzed immunohistochemically using anti-proliferating cell nuclear antigen (PCNA) antibody. No differences were observed by pathological examination or by PCNA staining between the sham- and the RF-exposed groups. The expression of cyclin D1 and c-fos were detected only in the tumorous skin tissues of the TPA-treated group.Conclusion: No evidence was found that RF radiation serves as a tumor promoter for skin tumors. Our data suggests that 849 MHz and 1,763 MHz RF radiations, similar to those emitted from mobile phones, do not have any promoting effect on skin tumor development in DMBA-initiated mice.