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Redox Biol ; 24: 101206, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-31039479


We tested whether novel CYP11A1-derived vitamin D3- and lumisterol-hydroxyderivatives, including 1,25(OH)2D3, 20(OH)D3, 1,20(OH)2D3, 20,23(OH)2D3, 1,20,23(OH)3D3, lumisterol, 20(OH)L3, 22(OH)L3, 20,22(OH)2L3, and 24(OH)L3, can protect against UVB-induced damage in human epidermal keratinocytes. Cells were treated with above compounds for 24 h, then subjected to UVB irradiation at UVB doses of 25, 50, 75, or 200 mJ/cm2, and then examined for oxidant formation, proliferation, DNA damage, and the expression of genes at the mRNA and protein levels. Oxidant formation and proliferation were determined by the DCFA-DA and MTS assays, respectively. DNA damage was assessed using the comet assay. Expression of antioxidative genes was evaluated by real-time RT-PCR analysis. Nuclear expression of CPD, phospho-p53, and Nrf2 as well as its target proteins including HO-1, CAT, and MnSOD, were assayed by immunofluorescence and western blotting. Treatment of cells with the above compounds at concentrations of 1 or 100 nM showed a dose-dependent reduction in oxidant formation. At 100 nM they inhibited the proliferation of cultured keratinocytes. When keratinocytes were irradiated with 50-200 mJ/cm2 of UVB they also protected against DNA damage, and/or induced DNA repair by enhancing the repair of 6-4PP and attenuating CPD levels and the tail moment of comets. Treatment with test compounds increased expression of Nrf2-target genes involved in the antioxidant response including GR, HO-1, CAT, SOD1, and SOD2, with increased protein expression for HO-1, CAT, and MnSOD. The treatment also stimulated the phosphorylation of p53 at Ser-15, increased its concentration in the nucleus and enhanced Nrf2 translocation into the nucleus. In conclusion, pretreatment of keratinocytes with 1,25(OH)2D3 or CYP11A1-derived vitamin D3- or lumisterol hydroxy-derivatives, protected them against UVB-induced damage via activation of the Nrf2-dependent antioxidant response and p53-phosphorylation, as well as by the induction of the DNA repair system. Thus, the new vitamin D3 and lumisterol hydroxy-derivatives represent promising anti-photodamaging agents.

J Pharmacol Exp Ther ; 360(3): 388-398, 2017 03.
Artigo em Inglês | MEDLINE | ID: mdl-28011874


UVA irradiation plays a role in premature aging of the skin through triggering oxidative stress-associated stimulation of matrix metalloproteinase-1 (MMP-1) responsible for collagen degradation, a hallmark of photoaged skin. Compounds that can activate nuclear factor E2-related factor 2 (Nrf2), a transcription factor regulating antioxidant gene expression, should therefore serve as effective antiphotoaging agents. We investigated whether genetic silencing of Nrf2 could relieve UVA-mediated MMP-1 upregulation via activation of mitogen-activated protein kinase (MAPK)/activator protein 1 (AP-1) signaling using human keratinocyte cell line (HaCaT). Antiphotoaging effects of hispidulin (HPD) and sulforaphane (SFN) were assessed on their abilities to activate Nrf2 in controlling MMP-1 and collagen expressions in association with phosphorylation of MAPKs (extracellular signal-regulated kinase, c-Jun N-terminal kinase, and p38), c-Jun, and c-Fos, using the skin of BALB/c mice subjected to repetitive UVA irradiation. Our findings suggested that depletion of Nrf2 promoted both mRNA expression and activity of MMP-1 in the UVA-irradiated HaCaT cells. Treatment of Nrf2 knocked-down HaCaT cells with MAPK inhibitors significantly suppressed UVA-induced MMP-1 and AP-1 activities. Moreover, pretreatment of the mouse skin with HPD and SFN, which could activate Nrf2, provided protective effects against UVA-mediated MMP-1 induction and collagen depletion in correlation with the decreased levels of phosphorylated MAPKs, c-Jun, and c-Fos in the mouse skin. In conclusion, Nrf2 could influence UVA-mediated MMP-1 upregulation through the MAPK/AP-1 signaling cascades. HPD and SFN may therefore represent promising antiphotoaging candidates.

Colágeno/metabolismo , Flavonas/farmacologia , Isotiocianatos/farmacologia , Fator 2 Relacionado a NF-E2/metabolismo , Envelhecimento da Pele , Raios Ultravioleta/efeitos adversos , Senilidade Prematura/etiologia , Senilidade Prematura/metabolismo , Animais , Antimutagênicos/farmacologia , Linhagem Celular , Ativação Enzimática/efeitos da radiação , Humanos , Queratinócitos , Sistema de Sinalização das MAP Quinases/efeitos da radiação , Metaloproteinase 1 da Matriz/metabolismo , Camundongos , Estresse Oxidativo , Pele/metabolismo , Envelhecimento da Pele/efeitos dos fármacos , Envelhecimento da Pele/efeitos da radiação , Fator de Transcrição AP-1/metabolismo , Regulação para Cima/efeitos da radiação
Redox Biol ; 8: 79-90, 2016 08.
Artigo em Inglês | MEDLINE | ID: mdl-26765101


Dietary phenolics may play a protective role in UV-mediated skin pigmentation through their antioxidant and UV-absorbing actions. In this study, we investigated whether genetic silencing of Nrf2, regulating the transcription of antioxidant genes, affected melanogenesis in primary human epidermal melanocytes (HEMn) and B16F10 melanoma cells subjected to UVA (8J/cm(2)) exposure. Then, we explored the antimelanogenic actions of phenolics; caffeic acid (CA) and ferulic acid (FA) providing partial UVA protection; quercetin (QU) and rutin (RU) providing strong UVA protection and; avobenzone (AV), an efficient UVA filter, in association with modulation of Nrf2-mediated antioxidant defenses in response to UVA insults in B16F10 cells. Upon oxidative insults, Nrf2 silencing promoted melanogenesis in both HEMn and B16F10 cells irradiated with UVA. Stimulation of melanogenesis by UVA correlated with increased ROS and oxidative DNA damage (8-OHdG), GSH depletion as well as a transient downregulation of Nrf2 nuclear translocation and of Nrf2-ARE signaling in B16F10 cells. All test compounds exerted antimelanogenic effects with respect to their abilities to reverse UVA-mediated oxidative damage as well as downregulation of Nrf2 activity and its target antioxidants (GCLC, GST and NQO1) in B16F10 cells. In conclusion, defective Nrf2 may promote melanogenesis under UVA irradiation through oxidative stress mechanisms. Compounds with antioxidant and/or UVA absorption properties could protect against UVA-induced melanogenesis through indirect regulatory effect on Nrf2-ARE pathway.

Antioxidantes/farmacologia , Suplementos Nutricionais , Melaninas/biossíntese , Fator 2 Relacionado a NF-E2/metabolismo , Fenóis/farmacologia , Substâncias Protetoras/farmacologia , Raios Ultravioleta , Animais , Elementos de Resposta Antioxidante , Vias Biossintéticas/efeitos dos fármacos , Vias Biossintéticas/efeitos da radiação , Dano ao DNA/efeitos dos fármacos , Desoxiguanosina/análogos & derivados , Desoxiguanosina/farmacologia , Técnicas de Silenciamento de Genes , Glutationa/metabolismo , Humanos , Melanócitos/efeitos dos fármacos , Melanócitos/metabolismo , Melanócitos/efeitos da radiação , Melanoma/genética , Melanoma/metabolismo , Melanoma Experimental , Camundongos , Monofenol Mono-Oxigenase/metabolismo , Fator 2 Relacionado a NF-E2/genética , Interferência de RNA , Espécies Reativas de Oxigênio/metabolismo , Transcrição Genética