Toshinori Bito

UV-induced skin damage

Solar radiation induces acute and chronic reactions in human and animal skin. Chronic repeated exposures are the primary cause of benign and malignant skin tumors, including malignant melanoma. Among types of solar radiation, ultraviolet B (290-320 nm) radiation is highly mutagenic and carcinogenic in animal experiments compared to ultraviolet A (320-400 nm) radiation. Epidemiological studies suggest that solar UV radiation is responsible for skin tumor development via gene mutations and immunosuppression, and possibly for photoaging. In this review, recent understanding of DNA damage caused by direct UV radiation and by indirect stress via reactive oxygen species (ROS) and DNA repair mechanisms, particularly nucleotide excision repair of human cells, are discussed. In addition, mutations induced by solar UV radiation in p53, ras and patched genes of non-melanoma skin cancer cells, and the role of ROS as both a promoter in UV-carcinogenesis and an inducer of UV-apoptosis, are described based primarily on the findings reported during the last decade. Furthermore, the effect of UV on immunological reaction in the skin is discussed. Finally, possible prevention of UV-induced skin cancer by feeding or topical use of antioxidants, such as polyphenols, vitamin C, and vitamin E, is discussed.

Preventive effect of antioxidant on ultraviolet-induced skin cancer in mice

Reactive oxygen species (ROS) have been shown to be responsible for inducing DNA damage after ultraviolet radiation (UV). Antioxidant, vitamin E and epigallocatechin gallate extracted from green tea, applied topically to the skin, delayed the onset of UV-induced skin cancer in mice. Since olive oil is reported to have a potent antioxidative effect in in vitro system, we asked whether, topical use of olive oil reduces the number and delays the onset of UV-induced skin cancer in mice. We found that super virgin olive oil painted immediately after UVB radiation significantly delayed the onset and reduced the number of skin cancer, but pretreatment of super virgin olive oil and pre- and/or post treatment by regular olive oil neither retarded nor reduced skin cancer formation in UV-irradiated mice. Further, 8-hydroxy-deoxyguanosine (8-OHdG) formation in mice epidermis was apparently reduced by super virgin olive oil painted immediately after UV radiation, although cyclobutane pyrimidine dimers and (6-4) photoproducts were not reduced by olive oil treatment. Our results suggest that daily topical use of super virgin olive oil after sun bathing may delay and reduce UV-induced skin cancer development in human skin, possibly by decreasing ROS-induced 8-OHdG which is responsible for gene mutation.

A group of atopic dermatitis without IgE elevation or barrier impairment shows a high Th1 frequency: possible immunological state of the intrinsic type.

BACKGROUND Atopic dermatitis (AD) can be classified into the major extrinsic type with high serum IgE levels and impaired barrier, and the minor intrinsic type with normal IgE levels and unimpaired barrier. OBJECTIVE To characterize the intrinsic type of Japanese AD patients in the T helper cell polarization in relation to the barrier condition. METHODS Enrolled in this study were 21 AD patients with IgE<200kU/L (IgE-low group; 82.5±59.6kU/L) having unimpaired barrier, and 48 AD patients with IgE>500kU/L (IgE-high group; 8.050±10.400kU/L). We investigated filaggrin gene (FLG) mutations evaluated in the eight loci common to Japanese patients, circulating Th1, Th2 and Th17 cells by intracellular cytokine staining and flow cytometry, and blood levels of CCL17/TARC, IL-18, and substance P by ELISA. RESULTS The incidence of FLG mutations was significantly lower in the IgE-low group (10.5%) than the IgE-high group (44.4%) (normal individuals, 3.7%). The percentage of IFN-γ-producing Th1, but not Th2 or Th17, was significantly higher in the IgE-low than IgE-high group. Accordingly, Th2-attracting chemokine CCL17/TARC, was significantly lower in the IgE-low than the IgE-high group. There were no differences between them in serum IL-18 levels, or the plasma substance P levels or its correlation with pruritus. CONCLUSION The IgE-low group differed from the IgE-high group in that it had much less FLG mutations, increased frequency of Th1 cells, and lower levels of CCL17. In the intrinsic type, non-protein antigens capable of penetrating the unimpaired barrier may induce a Th1 eczematous response.

Involvement of EGF receptor activation in the induction of cyclooxygenase‐2 in HaCaT keratinocytes after UVB

Abstract:  Because selective inhibition of cyclooxygenase‐2 (COX‐2) suppressed the induction of skin tumors in mice by UV and as UV has been shown to induce expression of COX‐2 in skin and cells, COX‐2 may be crucial for photocarcinogenesis of the skin. We studied the mechanism of UVB‐induced expression of COX‐2 focusing on the signal transduction pathway involved. Hydrogen peroxide (H2O2) treatment of HaCaT cells induced expression of COX‐2 and pretreatment with the antioxidant N‐acetylcysteine (NAC) partly inhibited the UVB‐induced expression of COX‐2 protein in HaCaT cells, suggesting that oxidative stress contributes to COX‐2 induction. To examine the signaling pathways involved in the UVB‐induced expression of COX‐2 in HaCaT cells, we analysed the expression of COX‐2 protein after treatment with various inhibitors of signaling molecules. Inhibition of EGFR by a specific inhibitor and by a neutralizing antibody suppressed the induction of COX‐2 expression by UV. Although a neutralizing antibody to transforming growth factor‐α (TGF‐α) suppressed COX‐2 expression induced by TGF‐α, it did not suppress COX‐2 expression by UV, indicating that a direct activation of EGFR is involved. Treatment of cells at low temperature (4°C) inhibited UVB‐induced JNK activation, but it did not inhibit COX‐2 expression by UV. Inhibitors of MEK, p38 MAP kinase and PI3‐kinase, suppressed the induction of COX‐2 expression by UV. In contrast, an erbB‐2 inhibitor augmented the UVB‐induced increase of COX‐2 protein. These data indicate that oxidative stress in association with activation of EGFR, ERK, p38 MAP kinase, and PI3‐kinase plays crucial roles in the UVB induction of expression of COX‐2.

Cyclin D and retinoblastoma gene product expression in actinic keratosis and cutaneous squamous cell carcinoma in relation to p53 expression

Masato Ueda, Abnormality of the molecules regulating the cell cycle has been shown to lead cells to transformation. Recently, overexpression of cyclin D protein, one of the G1 cyclins, and the abnormality of the retinoblastoma gene have been found in various human cancers. We analyzed the expression of cyclin D, retinoblastoma gene product (pRB) and p53 in actinic keratoses (AKs) and cutaneous squamous cell carcinomas (SCCs) by immunohistochemistry to elucidate the role of these molecules in keratinocyte carcinogenesis. In the normal epidermis, a few cyclin D positive cells were seen mainly at die basal layer. In 11 seborrheic keratoses, no overexpression of cyclin D was observed. Twelve of 26 AKs (4.5%) and 27 of 45 SCCs (60%) showed cyclin D overexpression. A few pRB positive cells were seen in the basal layer and in the supra‐basal spinous layer of the normal epidermis. An abnormality of pRB, loss of expression, was seen in 2 of 26 AKs (8%) and 7 of 45 SCCs (16%). p53 protein was positive in 12 of 26 AKs (46%) and 24 of 45 SCCs (53%). Forty‐five SCCs examined were divided into 22 ultraviolet (UV)‐related SCCs and 23 UV‐unrelated SCCs. Though UV‐related SCCs showed a significantly higher incidence of p53 positivity, as previously reported by us, no significant difference in cyclin D overexpression and loss of the pRB expression was observed between UV‐related and UV‐unrelated SCCs. These results suggest that cyclin D overexpression is frequently involved in keratinocyte carcinogenesis and that this is an early event, as well as p53 abnormality. In addition, abnormality of the retinoblastoma gene is also related to epidermal cell carcinogenesis, though die frequency is relatively low.

Impact of reactive oxygen species on keratinocyte signaling pathways

Human epidermal keratinocytes are located on the body surface, which is a specialized location for component cells of human skin tissue. Those cells are always exposed to external stimuli, which constantly generate reactive oxygen species (ROS) in the cells. Regulation of the redox state is a key for maintaining intracellular homeostasis. Originally, each cell type has defensive systems against oxidative stress, thus keratinocytes may have a unique system for regulating ROS levels. Intracellular signaling appropriately reacts to changes of ROS in cooperation with intra- and extra-cellular antioxidant agents, and is sometimes affected by excessive ROS generated by various stresses. We review in this paper the impact of ROS on keratinocytes based on published data and focus on related signaling pathways involved in inflammation and oncogenesis.

Comparison of skin barrier function and sensory nerve electric current perception threshold between IgE‐high extrinsic and IgE‐normal intrinsic types of atopic dermatitis

Background  Two types of atopic dermatitis (AD) have been proposed, with different pathophysiological mechanisms underlying this seemingly heterogeneous disorder. The extrinsic type shows high IgE levels presumably as a consequence of skin barrier damage and feasible allergen permeation, whereas the intrinsic type exhibits normal IgE levels and is not mediated by allergen‐specific IgE.

The Mandatory Role of IL-10–Producing and OX40 Ligand-Expressing Mature Langerhans Cells in Local UVB-Induced Immunosuppression

The mechanism underlying the local UVB-induced immunosuppression is a central issue to be clarified in photoimmunology. There have been reported a considerable number of cells and factors that participate in the sensitization phase-dependent suppression, including Langerhans cells (LCs), regulatory T cells, IL-10, and TNF-α. The recent important finding that LC-depleted mice rather exhibit enhanced contact hypersensitivity responses urged us to re-evaluate the role of LCs along with dermal dendritic cells (dDCs) in the mechanism of UVB-induced immunosuppression. We studied the surface expression of OX40 ligand (OX40L) and the intracellular expression of IL-10 in LCs and dDCs from UVB-irradiated (300 mJ/cm2) skin of BALB/c mice and those migrating to the regional lymph nodes from UVB-irradiated, hapten-painted mice. In epidermal and dermal cell suspensions prepared from the UVB-irradiated skin, LCs expressed OX40L as well as CD86 and produced IL-10 at a higher level than Langerin‒ dDCs. The UVB-induced immunosuppression was attenuated by the administration of IL-10–neutralizing or OX40L-blocking Abs. In mice whose UVB-irradiated, hapten-painted skin was dissected 1 d after hapten application, the contact hypersensitivity response was restored, because this treatment allowed dDCs but not LCs to migrate to the draining lymph nodes. Moreover, LC-depleted mice by using Langerin-diphtheria toxin receptor–knocked-in mice showed impaired UVB-induced immunosuppression. These results suggest that IL-10–producing and OX40L-expressing LCs in the UVB-exposed skin are mandatory for the induction of Ag-specific regulatory T cells.

Etodolac, a Selective Cyclooxygenase-2 Inhibitor, Induces Upregulation of E-Cadherin and Has Antitumor Effect on Human Bladder Cancer Cells In Vitro and In Vivo

OBJECTIVES Cyclooxygenase-2 (COX-2) is highly expressed in several human cancers, including bladder cancer. Thus, a selective COX-2 inhibitor could be useful as an antitumor agent for a range of cancers. In the present study, we investigated the antitumor effect and E-cadherin induction of etodolac, a highly selective COX-2 inhibitor, on human bladder cancer cells in vitro and in vivo. METHODS We examined the cytotoxicity of etodolac against three human bladder cancer cell lines, T24, 5637, and KK47, and performed quantitative reverse transcriptase-polymerase chain reaction to measure the mRNA expression of COX-2, and E-cadherin. RESULTS Etodolac showed significant cytotoxicity only to T24 cells, which expressed the greatest level of COX-2 mRNA and the lowest level of E-cadherin mRNA among the three cell lines. Etodolac also increased the E-cadherin mRNA expression in T24 cells in vitro. We also found that etodolac suppressed tumor growth and induced E-cadherin expression and cell apoptosis in a T24 tumor xenograft mouse model. CONCLUSIONS Etodolac exhibited antitumor activity and induced E-cadherin expression in bladder cancer cells and might be useful for the clinical treatment and prevention of bladder cancer, especially in poorly differentiated bladder cancer with high COX-2 and low E-cadherin expression.

Cows Milk-Dependent Exercise- Induced Anaphylaxis under the Condition of a Premenstrual or Ovulatory Phase Following Skin Sensitization

BACKGROUND A 24 year-old woman with atopic dermatitis occasionally developed symptoms, including dyspnea and generalized urticaria, following ingestion of food containing cows milk. Similar episodes had continued, and had been treated empirically since the age of 16 years. CASE SUMMARY Although a skin test and IgE RAST showed positive reactions to cows milk, a provocation test with cows milk alone did not induce any symptoms. Therefore, food-dependent exercise-induced anaphylaxis (FDEIA) was suspected, but examination using various combinations of cows milk, aspirin and exercise failed to elicit any symptoms. Finally, a provocation test during the ovulatory phase with cows milk followed by aspirin and exercise evoked systemic urticaria, dyspnea and hypotension. DISCUSSION The symptoms against cows milk began when she took baths with bath salts containing cows milk as its main ingredient for one year at the age 15 years. Sensitization to cows milk through eczematous skin is indicated from this history. Hormonal change during a premenstrual or ovulatory phase is also an important factor for the development of FDEIA in this case.BACKGROUND A 24 year-old woman with atopic dermatitis occasionally developed symptoms, including dyspnea and generalized urticaria, following ingestion of food containing cows milk. Similar episodes had continued, and had been treated empirically since the age of 16 years. CASE SUMMARY Although a skin test and IgE RAST showed positive reactions to cows milk, a provocation test with cows milk alone did not induce any symptoms. Therefore, food-dependent exercise-induced anaphylaxis (FDEIA) was suspected, but examination using various combinations of cows milk, aspirin and exercise failed to elicit any symptoms. Finally, a provocation test during the ovulatory phase with cows milk followed by aspirin and exercise evoked systemic urticaria, dyspnea and hypotension. DISCUSSION The symptoms against cows milk began when she took baths with bath salts containing cows milk as its main ingredient for one year at the age 15 years. Sensitization to cows milk through eczematous skin is indicated from this history. Hormonal change during a premenstrual or ovulatory phase is also an important factor for the development of FDEIA in this case.