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1.
Int J Mol Sci ; 22(14)2021 Jul 13.
Artigo em Inglês | MEDLINE | ID: mdl-34299118

RESUMO

The aryl hydrocarbon receptor (AhR), a ligand-activated transcription factor expressed in all skin cell types, plays a key role in physiological and pathological processes. Several studies have shown that this receptor is involved in the prevention of inflammatory skin diseases, e.g., psoriasis, atopic dermatitis, representing a potential therapeutic target. We tested the safety profile and the biological activity of NPD-0614-13 and NPD-0614-24, two new synthetic AhR ligands structurally related to the natural agonist FICZ, known to be effective in psoriasis. NPD-0614-13 and NPD-0614-24 did not alter per se the physiological functions of the different skin cell populations involved in the pathogenesis of inflammatory skin diseases. In human primary keratinocytes stimulated with tumor necrosis factor-α or lipopolysaccharide the compounds were able to counteract the altered proliferation and to dampen inflammatory signaling by reducing the activation of p38MAPK, c-Jun, NF-kBp65, and the release of cytokines. Furthermore, the molecules were tested for their beneficial effects in human epidermal and full-thickness reconstituted skin models of psoriasis. NPD-0614-13 and NPD-0614-24 recovered the psoriasis skin phenotype exerting pro-differentiating activity and reducing the expression of pro-inflammatory cytokines and antimicrobial peptides. These data provide a rationale for considering NPD-0614-13 and NPD-0614-24 in the management of psoriasis.


Assuntos
Anti-Inflamatórios/farmacologia , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Catecóis/farmacologia , Diferenciação Celular , Inflamação/tratamento farmacológico , Compostos Organometálicos/farmacologia , Psoríase/tratamento farmacológico , Receptores de Hidrocarboneto Arílico/metabolismo , Pele/efeitos dos fármacos , Humanos , Inflamação/metabolismo , Inflamação/patologia , Queratinócitos/efeitos dos fármacos , Queratinócitos/metabolismo , Queratinócitos/patologia , Ligantes , Psoríase/metabolismo , Psoríase/patologia , Pele/metabolismo , Pele/patologia
2.
Pigment Cell Melanoma Res ; 34(1): 72-88, 2021 01.
Artigo em Inglês | MEDLINE | ID: mdl-32608114

RESUMO

The melanocortin-1 receptor (MC1R) belongs to the family of the G protein-coupled receptor (GPCR). Activated GPCRs can promote the phosphoinositide 3-kinase (PI3K) pathway. Few studies deal with the role of the PI3K pathway activation in response to αMSH. On B16-F10 cell line, we investigated the αMSH-dependent modulation of pAKT/AKT, as a key element of the PI3K pathway after rapid and prolonged stimulation. We demonstrated that αMSH triggers a rapid modulation of AKT which culminates in an increase in its phosphorylation. We highlighted a comparable upregulation of pAKT after exposure to αMSH on primary cultures of normal human melanocytes (NHMs) expressing a wild-type MC1R. On B16-F10 cells, NHMs, and an ex vivo model of human skin biopsies, we explored the influence of PI3K/AKT signaling triggered by αMSH, focusing on the control of melanogenesis and pigment release. We showed that the αMSH-dependent PI3K/AKT pathway exerts a negative feedback on melanogenesis and promotes the extracellular release of pigment. We strengthened the role of the PI3K/AKT pathway triggered by αMSH in preserving redox equilibrium and genome integrity, highlighting its role in affecting cell survival.

3.
Exp Dermatol ; 29(9): 833-839, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32779245

RESUMO

Acne is the most common skin disease in adolescent Westernized populations. Several data support the involvement of the mammalian target of rapamycin complex 1 (mTORC1) signalling in the interplay between androgens, insulin, insulin-like growth factor (IGF1) and high-glycaemic index diet in acne. The peroxisome proliferator-activated receptor γ (PPARγ) is involved in both differentiation and anti-inflammatory response. Low differentiated sebocytes showed decreased expression of PPARγ and increased level of insulin and IGF-1 receptors, resulting in the production of acne-like sebum and inflammatory mediators. In this viewpoint, we discuss how in acne the dysregulation of proliferation and differentiation processes in sebocytes and keratinocytes may be associated with altered response to androgens and other hormones, such as insulin or IGF-1. Moreover, we propose PPARγ modulation as an innovative therapeutic approach to normalize sebocyte differentiation process, interfering with the different mechanisms involved in altered pilosebaceous unit.

4.
FASEB J ; 34(5): 6302-6321, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-32157742

RESUMO

Bovine colostrum, the first milk secreted by the mammary glands of cows shortly after they have given birth, provides a natural source of bioactive substances helpful to promote tissue development and repair, and to maintain a healthy immune system. Owing to its properties, the use of colostrum in the treatment of human diseases is under investigation. We evaluated the biological activity of colostrum on human primary keratinocytes, focusing on its effects with regard to a proliferation/differentiation balance. Using cellular and molecular approaches, we showed that colostrum favors a cell cycle withdrawal by increasing the expression of p21/WAF1 and p27/KIP1. It also promotes the transition of keratinocytes from a proliferating to a differentiating state, as assessed by a decrease in keratin 5 and an increase in keratin 16. We demonstrated the ability of colostrum to induce the expression of early and late differentiation markers (keratin 1, involucrin, and filaggrin) and the synthesis of caspase 14 and bleomycin hydrolase, the two main enzymes involved in filaggrin maturation. Moreover, we showed that bovine colostrum is able to promote keratinocyte stratification and terminal differentiation not only in two-dimensional (2D), but also in a more physiological system of three-dimensional (3D) skin equivalents. Finally, we demonstrated that colostrum stimulates cell differentiation through the PI3K/PLC-γ1/PKCα pathways mainly associated to tyrosine kinase receptors. These results suggest the possibility to benefit from colostrum properties for the treatment of skin diseases characterized by altered differentiation and perturbed barrier function.


Assuntos
Diferenciação Celular , Colostro/metabolismo , Proteínas de Filamentos Intermediários/metabolismo , Queratinócitos/citologia , Pele/citologia , Animais , Bovinos , Células Cultivadas , Feminino , Humanos , Queratinócitos/metabolismo , Gravidez , Pele/metabolismo
5.
J Exp Clin Cancer Res ; 36(1): 142, 2017 10 11.
Artigo em Inglês | MEDLINE | ID: mdl-29020973

RESUMO

BACKGROUND: The α-Melanocyte Stimulating Hormone (αMSH)/Melanocortin-1 receptor (MC1R) interaction promotes melanogenesis through the cAMP/PKA pathway. The direct induction of this pathway by Forskolin (FSK) is also known to enhance melanocyte proliferation. αMSH acts as a mitogenic agent in melanocytes and its effect on proliferation of melanoma cells is less known. We previously identified the αMSH/Peroxisome Proliferator Activated Receptor (PPARγ) pathway as a new pathway on the B16-F10 mouse melanoma cell line. αMSH induced the translocation of PPARγ into the nucleus as an active transcription factor. This effect was independent of the cAMP/PKA pathway and was mediated by the activation of the PI(4,5)P2/PLC pathway, a pathway which we have described to be triggered by the αMSH-dependent MC1R stimulation. Moreover, in the same study, preliminary experiments showed that mouse melanoma cells responded to αMSH by reducing proliferation and that PPARγ was involved in this effect. Due to its key role in the control of cell proliferation, PPARγ agonists are used in therapeutic models for different forms of cancer, including melanoma. The purpose of this study was: (a) to confirm the different proliferative behavior in response to αMSH in healthy and in melanoma condition; (b) to verify whether the cAMP/PKA pathway and the PLC/PPARγ pathway could exert an antagonistic function in the control of proliferation; (c) to deepen the knowledge of the molecular basis responsible for the down-proliferative response of melanoma cells after exposure to αMSH. METHODS: We employed B16-F10 cell line, a human melanoma cell line (Mel 13) and two primary cultures of human melanocytes (NHM 1 and NHM 2, respectively), all expressing a wild type MC1R and responding to the αMSH in terms of pigmentation. We evaluated cell proliferation through: a) cell counting, b) cell cycle analysis c) protein expression of proliferation modulators (p27, p21, cyclin D1 and cyclin E). RESULTS: The αMSH acted as a mitogenic agent in primary cultures of human melanocytes, whereas it determined a slow down of proliferation in melanoma cell lines. FSK, as an inducer of the cAMP/PKA pathway, reproduced the αMSH mediated effect on proliferation in NHMs but it did not mimic the αMSH effect on proliferation in B16-F10 and Mel 13 melanoma cell lines. Meanwhile, 3 M3-FBS (3 M3), as an inducer of PI(4,5)P2/PLC pathway, reproduced the αMSH proliferative effect. Further experiments, treating melanoma cell lines with αMSH in the presence/absence of GW9662, as an inhibitor of PPARγ, confirmed the key role of this transcription factor in decreasing cell proliferation in response to the hormone exposure. CONCLUSIONS: In both melanoma cell lines, αMSH determined the reduction of proliferation through the PI(4,5)P2/PLC pathway, employing PPARγ as an effector element. These evidence could offer perspectives for new therapeutic approaches for melanoma.


Assuntos
Melanoma/metabolismo , PPAR gama/metabolismo , Transdução de Sinais , alfa-MSH/metabolismo , Animais , Linhagem Celular Tumoral , Proliferação de Células , AMP Cíclico/metabolismo , Proteínas Quinases Dependentes de AMP Cíclico/metabolismo , Expressão Gênica , Genes Reporter , Humanos , Melanoma/genética , Melanoma/patologia , Melanoma Experimental , Camundongos , PPAR gama/genética , Ativação Transcricional
6.
Sci Rep ; 7(1): 9241, 2017 08 23.
Artigo em Inglês | MEDLINE | ID: mdl-28835664

RESUMO

Increasing attention is addressed to identify products able to enhance skin photoprotection and to prevent skin carcinogenesis. Several studies have demonstrated that the α-melanocyte stimulating hormone (αMSH), acting on a functional MC1R, provides a photoprotective effect by inducing pigmentation, antioxidants and DNA repair. We discovered a link between αMSH and the nuclear receptor Peroxisome Proliferator-Activated Receptor-γ (PPARγ), suggesting that some of the αMSH protective effects may be dependent on PPARγ transcriptional activity. Moreover, we demonstrated that the activation of PPARγ by the parrodiene 2,4,6-octatrienoic acid (Octa) induces melanogenesis and antioxidant defence in human melanocytes and counteracts senescence-like phenotype in human fibroblasts. In this study, we demonstrate that the activation of PPARγ by Octa exerts a protective effect against UVA- and UVB-induced damage on normal human keratinocytes (NHKs), the major target cells of UV radiation. Octa promotes the antioxidant defence, augments DNA repair and reduces the induction of proteins involved in UV-induced DNA damage response. Our results contribute to deepen the analysis of the αMSH/PPARγ connection and suggest perspectives for the development of new molecules and formulations able to prevent cutaneous UV damage by acting on the different skin cell populations through PPARγ activation.


Assuntos
Ácidos Graxos Insaturados/farmacologia , Queratinócitos/efeitos dos fármacos , Queratinócitos/enzimologia , Queratinócitos/efeitos da radiação , PPAR gama/agonistas , Protetores contra Radiação/farmacologia , Raios Ultravioleta/efeitos adversos , Antioxidantes/metabolismo , Células Cultivadas , Dano ao DNA/efeitos dos fármacos , Dano ao DNA/efeitos da radiação , Ativação Enzimática/efeitos dos fármacos , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Queratinócitos/patologia , PPAR gama/genética , PPAR gama/metabolismo , Receptor Tipo 1 de Melanocortina/genética , Receptor Tipo 1 de Melanocortina/metabolismo , alfa-MSH/metabolismo
7.
Pigment Cell Melanoma Res ; 28(4): 378-89, 2015 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-25786343

RESUMO

Cutaneous phototype is considered mainly related to cutaneous pigmentation and to the eumelanin/pheomelanin ratio, which is mostly genetically determined by the melanocortin 1 receptor (MC1R) polymorphisms. However, data in literature indicate that, in addition to stimulation of eumelanin synthesis, the MC1R signalling activates antioxidant, DNA repair and survival pathways. New emerging aspects regarding photoprotection and skin phototypes are going beyond those features connected to the melanin content in the skin. Important new findings link the MC1R to nuclear receptors activation, shedding light on new extra-melanogenic effects dependent on the α-melanocyte-stimulating hormone (α-MSH) activity and new ways through which such functions are modulated. These evidences indicate that several factors including melanin play a part in defining the basis for individual sun sensitivity, suggesting that the cutaneous phototype represents a 'biochemical fingerprint'.


Assuntos
Pele/patologia , Pele/efeitos da radiação , Raios Ultravioleta , Humanos , Melaninas/biossíntese , Estresse Oxidativo/efeitos da radiação , Receptor Tipo 1 de Melanocortina/metabolismo , Transdução de Sinais/efeitos da radiação , Pigmentação da Pele/efeitos da radiação
8.
PLoS One ; 9(8): e104045, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-25101957

RESUMO

Peroxisome proliferator-activated receptor gamma (PPARγ) may be involved in a key mechanism of the skin aging process, influencing several aspects related to the age-related degeneration of skin cells, including antioxidant unbalance. Therefore, we investigated whether the up-modulation of this nuclear receptor exerts a protective effect in a stress-induced premature senescence (SIPS) model based on a single exposure of human dermal fibroblasts to 8-methoxypsoralen plus + ultraviolet-A-irradiation (PUVA). Among possible PPARγ modulators, we selected 2,4,6-octatrienoic acid (Octa), a member of the parrodiene family, previously reported to promote melanogenesis and antioxidant defense in normal human melanocytes through a mechanism involving PPARγ activation. Exposure to PUVA induced an early and significant decrease in PPARγ expression and activity. PPARγ up-modulation counteracted the antioxidant imbalance induced by PUVA and reduced the expression of stress response genes with a synergistic increase of different components of the cell antioxidant network, such as catalase and reduced glutathione. PUVA-treated fibroblasts grown in the presence of Octa are partially but significantly rescued from the features of the cellular senescence-like phenotype, such as cytoplasmic enlargement, the expression of senescence-associated-ß-galactosidase, matrix-metalloproteinase-1, and cell cycle proteins. Moreover, the alterations in the cell membrane lipids, such as the decrease in the polyunsaturated fatty acid content of phospholipids and the increase in cholesterol levels, which are typical features of cell aging, were prevented. Our data suggest that PPARγ is one of the targets of PUVA-SIPS and that its pharmacological up-modulation may represent a novel therapeutic approach for the photooxidative skin damage.


Assuntos
Senescência Celular , Fibroblastos/efeitos dos fármacos , PPAR gama/fisiologia , Estresse Fisiológico , Membrana Celular/química , Membrana Celular/metabolismo , Colesterol/metabolismo , Ácidos Graxos Insaturados/metabolismo , Ácidos Graxos Insaturados/farmacologia , Fibroblastos/efeitos da radiação , Humanos , Peroxidação de Lipídeos , Metoxaleno/farmacologia , PPAR gama/genética , PPAR gama/metabolismo , Fosfolipídeos/metabolismo , Substâncias Protetoras/farmacologia , Espécies Reativas de Oxigênio/metabolismo , Raios Ultravioleta , Regulação para Cima
9.
Exp Dermatol ; 22(1): 41-7, 2013 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-23278893

RESUMO

Azelaic acid (AzA) has been used for the treatment for inflammatory skin diseases, such as acne and rosacea. Interestingly, an improvement in skin texture has been observed after long-time treatment with AzA. We previously unrevealed that anti-inflammatory activity of AzA involves a specific activation of PPARγ, a nuclear receptor that plays a relevant role in inflammation and even in ageing processes. As rosacea has been considered as a photo-aggravated disease, we investigated the ability of AzA to counteract stress-induced premature cell senescence (SIPS). We employed a SIPS model based on single exposure of human dermal fibroblasts (HDFs) to UVA and 8-methoxypsoralen (PUVA), previously reported to activate a senescence-like phenotype, including long-term growth arrest, flattened morphology and increased synthesis of matrix metalloproteinases (MMPs) and senescence-associated ß-galactosidase (SA-ß-gal). We found that PUVA-treated HDFs grown in the presence of AzA maintained their morphology and reduced MMP-1 release and SA-ß-galactosidase-positive cells. Moreover, AzA induced a reduction in ROS generation, an up-modulation of antioxidant enzymes and a decrease in cell membrane lipid damages in PUVA-treated HDFs. Further evidences of AzA anti-senescence effect were repression of p53 and p21, increase in type I pro-collagen and abrogation of the enhanced expression of growth factors, such as HGF and SCF. Interestingly, PUVA-SIPS showed a decreased activation of PPARγ and AzA counteracted this effect, suggesting that AzA effect involves PPARγ modulation. All together these data showed that AzA interferes with PUVA-induced senescence-like phenotype and its ability to activate PPAR-γ provides relevant insights into the anti-senescence mechanism.


Assuntos
Senescência Celular/efeitos dos fármacos , Fármacos Dermatológicos/farmacologia , Ácidos Dicarboxílicos/farmacologia , Fibroblastos/efeitos dos fármacos , Fibroblastos/metabolismo , PPAR gama/metabolismo , Membrana Celular/efeitos dos fármacos , Membrana Celular/metabolismo , Células Cultivadas , Senescência Celular/efeitos da radiação , Colágeno Tipo I/metabolismo , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Fibroblastos/citologia , Fator de Crescimento de Hepatócito/metabolismo , Humanos , Metaloproteinase 1 da Matriz/metabolismo , Metoxaleno/farmacologia , Terapia PUVA , Fenótipo , Fosfolipídeos/metabolismo , Fármacos Fotossensibilizantes/farmacologia , Pró-Colágeno/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Fator de Células-Tronco/metabolismo , Proteína Supressora de Tumor p53/metabolismo , Raios Ultravioleta , beta-Galactosidase/metabolismo
10.
Pigment Cell Melanoma Res ; 26(1): 113-27, 2013 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-22863076

RESUMO

We have discovered a new α-melanocyte stimulating hormone (α-MSH)/peroxisome proliferator activated receptor-γ (PPAR-γ) connection in B16-F10 cells. Both PPAR-γ up-regulation and its induction as an active transcription factor were observed in response to α-MSH. The α-MSH/PPAR-γ connection influenced both pigmentation and proliferation. The forskolin-stimulated cAMP/PKA pathway was not able to induce either PPAR-γ translocation into the nucleus or PPAR-γ transcriptional activity. As the melanocortin-1 receptor, the specific receptor for the α-MSH, is a G-protein coupled receptor, we wondered whether the phosphatidylinositol [PI(4,5)P(2) /PLC(ß) ] signal pathway was involved in mediating the α-MSH-dependent PPAR-γ activation. Employing inhibitors of PI(4,5)P(2) /PLC(ß) pathway, the results of our experiments suggested that this pathway was promoted by α-MSH and that α-MSH played a role in mediating PPAR-γ activation. We have demonstrated, for the first time, that α-MSH induces the PI(4,5)P(2) /PLC(ß) pathway, through analysis of the basic steps of the pathway. The α-MSH effect on PPAR-γ was independent of animal species and was not correlated with the physio-pathological status.


Assuntos
Melanoma Experimental/metabolismo , PPAR gama/metabolismo , Neoplasias Cutâneas/metabolismo , alfa-MSH/farmacologia , Animais , Ácido Araquidônico/metabolismo , Cálcio/metabolismo , Membrana Celular/efeitos dos fármacos , Membrana Celular/metabolismo , Núcleo Celular/efeitos dos fármacos , Núcleo Celular/metabolismo , Proliferação de Células/efeitos dos fármacos , AMP Cíclico/metabolismo , Proteínas Quinases Dependentes de AMP Cíclico/metabolismo , Diglicerídeos/metabolismo , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Células HeLa , Humanos , Hidrólise/efeitos dos fármacos , Fosfatos de Inositol/metabolismo , Melanoma Experimental/genética , Melanoma Experimental/patologia , Camundongos , PPAR gama/genética , Fosfolipase C beta/metabolismo , Transporte Proteico/efeitos dos fármacos , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genética , Neoplasias Cutâneas/genética , Neoplasias Cutâneas/patologia
11.
FASEB J ; 26(9): 3779-89, 2012 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-22649030

RESUMO

Cystinosis is a rare autosomal recessive disease characterized by cystine crystal accumulation leading to multiorgan dysfunctions and caused by mutation in CTNS. CTNS encodes cystinosin, a cystine/H(+) symporter that exports cystine out of the lysosomes. Patients with cystinosis frequently exhibit blond hair and fair complexion, suggesting an alteration in melanogenesis. However, the pigmentation singularities of these patients have not been studied, and the role of cystinosin in melanogenesis has remained unknown. In our study, a clinical evaluation of 27 patients with cystinosis showed that 44% had a cutaneous pigmentation dilution compared to their relatives. Analysis of the hair melanin content in these patients by HPLC demonstrated a 50% decrease in eumelanin (4360 vs. 9360 ng/mg), and a 2-fold increase in pheomelanin (53 vs. 20 ng/mg), the yellow/red pigments. Cystinosin-deficient mice also showed a 4-fold increase in hair pheomelanin content. In vitro studies showed that cystinosin was located at melanosomes. CTNS silencing led to a 75% reduction of melanin synthesis that was caused by a degradation of tyrosinase by lysosomal proteases. Our results objectify the pigmentation defect in patients with cystinosis. We also identify the role of CTNS in melanogenesis and add a new gene to the list of the genes involved in the control of skin and hair pigmentation.


Assuntos
Sistemas de Transporte de Aminoácidos Neutros/fisiologia , Melaninas/biossíntese , Melanossomas/metabolismo , Adolescente , Adulto , Sistemas de Transporte de Aminoácidos Neutros/genética , Sistemas de Transporte de Aminoácidos Neutros/metabolismo , Animais , Linhagem Celular Tumoral , Criança , Pré-Escolar , Cistinose/metabolismo , Feminino , Humanos , Masculino , Camundongos , Camundongos Knockout , Mutação , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Pigmentação da Pele/genética , Adulto Jovem
12.
J Invest Dermatol ; 132(4): 1196-205, 2012 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-22297637

RESUMO

Interest in colorless intermediates of melanocyte metabolism has traditionally been related to their role as melanin precursors, though several lines of evidence scattered in the literature suggested that these compounds may exert an antioxidant and protective function per se unrelated to pigment synthesis. Herein, we disclose the remarkable protective and differentiating effects of 5,6-dihydroxyindole-2-carboxylic acid (DHICA), a diffusible dopachrome tautomerase (DCT)-dependent eumelanin intermediate, on primary cultures of human keratinocytes. At micromolar concentrations, DHICA induced: (a) time- and dose-dependent reduction of cell proliferation without concomitant toxicity; (b) enhanced expression of early (spinous keratins K1 and K10 and envelope protein involucrin) and late (loricrin and filaggrin) differentiation markers; (c) increased activities and expression of antioxidant enzymes; and (d) decreased cell damage and apoptosis following UVA exposure. The hitherto unrecognized role of DHICA as an antiproliferative, protective, and antiapoptotic endogenous cell messenger points to a reappraisal of the biological functions of melanocytes and DCT in skin homeostasis and photoprotection beyond the mere provision of melanin pigments, and provides, to our knowledge, a previously unreported possible explanation to the higher resistance of the dark-skinned eumelanic phenotypes to sunburn and skin cancer.


Assuntos
Comunicação Celular/efeitos dos fármacos , Diferenciação Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Células Epidérmicas , Epiderme/efeitos dos fármacos , Indóis/farmacologia , Melaninas/metabolismo , Apoptose/efeitos dos fármacos , Apoptose/efeitos da radiação , Comunicação Celular/fisiologia , Células Cultivadas , Relação Dose-Resposta a Droga , Epiderme/metabolismo , Humanos , Proteínas de Filamentos Intermediários/metabolismo , Queratinócitos/citologia , Queratinócitos/efeitos dos fármacos , Queratinócitos/metabolismo , Queratinas/metabolismo , Melanócitos/citologia , Melanócitos/efeitos dos fármacos , Melanócitos/metabolismo , Proteínas de Membrana/metabolismo , Receptores Ativados por Proliferador de Peroxissomo/metabolismo , Precursores de Proteínas/metabolismo , Fatores de Tempo
13.
Pigment Cell Melanoma Res ; 24(4): 618-30, 2011 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-21762468

RESUMO

Given the importance of the tanning response in protecting human skin from the harmful effects of UV radiation, one important research priority is to identify novel molecules that are capable of promoting pigmentation and/or antioxidant defence. Parrodienes share some structural features with carotenoids and retinoids, stimulate cell antioxidant defence and counteract senescence-like phenotype in fibroblasts. We selected the parrodiene-derivative 2,4,6-octatrienoic acid (Octa) to study its impact on key parameters of melanogenesis and antioxidant defence in organ-cultured human skin and in normal human melanocytes. Octa promoted melanogenesis by up-regulating tyrosinase and microphthalmia-associated transcription factor expression. This correlated with an increase of melanin content in both human epidermis in situ and cultured human epidermal melanocytes. Moreover, Octa increased the biological antioxidant potential content and the expression and activity of catalase. Activation of peroxisome proliferator-activated receptor (PPAR)-γ was necessary to evoke these effects. These data strongly encourage the systematic study of Octa as a novel candidate promoter of human skin photoprotection.


Assuntos
Antioxidantes/metabolismo , Ácidos Graxos Insaturados/farmacologia , Melaninas/biossíntese , Melanócitos/efeitos dos fármacos , Melanócitos/imunologia , PPAR gama/metabolismo , Catalase/metabolismo , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Indução Enzimática/efeitos dos fármacos , Epiderme/efeitos dos fármacos , Epiderme/fisiologia , Regulação da Expressão Gênica/efeitos dos fármacos , Inativação Gênica/efeitos dos fármacos , Humanos , Masculino , Melanócitos/citologia , Melanócitos/enzimologia , Fator de Transcrição Associado à Microftalmia/genética , Fator de Transcrição Associado à Microftalmia/metabolismo , Monofenol Mono-Oxigenase/biossíntese , Monofenol Mono-Oxigenase/genética , Técnicas de Cultura de Órgãos , PPAR gama/antagonistas & inibidores , PPAR gama/genética , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Couro Cabeludo/efeitos dos fármacos , Couro Cabeludo/fisiologia , Pigmentação da Pele/efeitos dos fármacos
14.
J Biol Chem ; 285(10): 7288-99, 2010 Mar 05.
Artigo em Inglês | MEDLINE | ID: mdl-20053998

RESUMO

The synthesis of melanin pigments, or melanogenesis, is regulated by the balance of a variety of signal transduction pathways. Among these pathways, p38 MAPK signaling was found to be involved in stress-induced melanogenesis and to be activated by alpha-melanocyte-stimulating hormone (alpha-MSH) and ultraviolet irradiation. Previous studies have shown that alpha-MSH-stimulated melanogenesis can be inhibited by blocking p38 MAPK activity with SB203580, a pyridinyl imidazole compound. Consistent with this, we observed that pyridinyl imidazoles (SB203580 and SB202190) inhibited both basal and alpha-MSH-induced melanogenesis in B16 melanoma cells. However, SB202474, which has no ability to inhibit p38 MAPK activity and is usually used as a negative control compound in p38 MAPK studies, also suppressed melanin synthesis induction. Furthermore, the independence of the p38 kinase pathway from the repression of melanogenesis by pyridinyl imidazole compounds was also confirmed by small interfering RNA experiments. Interfering with p38 MAPK expression surprisingly stimulated melanogenesis and tyrosinase family protein expression. Although the molecular mechanism(s) by which p38 promotes the degradation of melanogenic enzymes remain to be determined, the involvement of the ubiquitin-proteasome pathway was demonstrated by co-treatment with the proteasome-specific inhibitor MG132 and the relative decrease in the ubiquitination of tyrosinase in cells transfected with p38-specific small interfering RNA.


Assuntos
Sistema de Sinalização das MAP Quinases/fisiologia , Monofenol Mono-Oxigenase/metabolismo , Pigmentação/fisiologia , Complexo de Endopeptidases do Proteassoma/metabolismo , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo , Animais , Inibidores Enzimáticos/metabolismo , Estabilidade Enzimática , Regulação da Expressão Gênica/efeitos da radiação , Humanos , Imidazóis/metabolismo , Melaninas/biossíntese , Melaninas/genética , Melanoma Experimental/metabolismo , Camundongos , Fator de Transcrição Associado à Microftalmia/genética , Fator de Transcrição Associado à Microftalmia/metabolismo , Monofenol Mono-Oxigenase/genética , Piridinas/metabolismo , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Proteínas Ubiquitinadas/metabolismo , Raios Ultravioleta , alfa-MSH/metabolismo , Proteínas Quinases p38 Ativadas por Mitógeno/antagonistas & inibidores , Proteínas Quinases p38 Ativadas por Mitógeno/genética
15.
Pigment Cell Melanoma Res ; 23(2): 263-75, 2010 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-20067588

RESUMO

We demonstrated a direct correlation between melanogenic and catalase activities on in vitro and ex vivo models. Here, we investigated whether the stimulation of Melanocortin-1 Receptor (MC1R) could influence catalase expression, activity and cellular localization. For this purpose, we treated B16-F0 melanoma cells with alpha-Melanocyte Stimulating Hormone (alpha-MSH) and we showed a rapid induction of catalase through a cAMP/PKA-dependent, microphthalmia-associated transcription factor (MITF) independent mechanism, acting at post-transcriptional level. Moreover, alpha-MSH promoted a partial re-distribution of catalase to the cell periphery and dendrites. This work strengthens the correlation between melanogenesis and anti-oxidants, demonstrating the induction of catalase in response to a melanogenic stimulation, such as alpha-MSH-dependent MC1R activation. Moreover, this study highlights catalase regulatory mechanisms poorly known, and attributes to alpha-MSH a protective role in defending melanocytes, and possibly keratinocytes, not only on the basis of its pigmentary action, but also for its capacity to stimulate a quick anti-oxidant defence.


Assuntos
Catalase/metabolismo , Membrana Celular/efeitos dos fármacos , Dendritos/efeitos dos fármacos , Melanoma/metabolismo , Receptor Tipo 1 de Melanocortina/metabolismo , alfa-MSH/farmacologia , Animais , Catalase/biossíntese , Membrana Celular/enzimologia , AMP Cíclico/metabolismo , Proteína Quinase Tipo II Dependente de AMP Cíclico/metabolismo , Dendritos/enzimologia , Melanoma/patologia , Camundongos , Transporte Proteico/efeitos dos fármacos , Receptor Tipo 1 de Melanocortina/agonistas , Relação Estrutura-Atividade , Células Tumorais Cultivadas
16.
Melanoma Res ; 19(6): 372-8, 2009 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-19741552

RESUMO

Malignant melanoma incidence is increasing rapidly in Western countries. Its prevention requires a deep knowledge of the biological basis of the neoplasm leading to the identification of new biological risk markers. In in-vitro and ex-vivo models we demonstrated that catalase was modified not only in its activity but also in its charge properties after ultraviolet A irradiation through pheomelanin. Here we focus on the electrophoretic behaviour of catalase in the human skin in vivo, in association with cutaneous phototype. Zymographic analysis of the enzyme on skin biopsies from Caucasian population (phototype I-IV), collected from the trunk in autumn-winter, to exclude possible influences of an acute photoexposure, evidenced a protein doublet, representing the coexistence of two active isoforms of catalase with different charge properties. In the skin from low-phototype subjects, the percent contribution of the more acidic component of the doublet was prevalent, inversely correlated with total melanin concentration in hair, and associated with a high number of melanocytic nevi. In summary, this study shows for the first time the existence of an acidic catalase in association with clinically defined risk characteristics in low phototype skin in vivo, contributing to the knowledge of a new biochemical marker of cutaneous photosusceptibility.


Assuntos
Catalase/metabolismo , Cabelo/enzimologia , Melanoma/enzimologia , Melanoma/metabolismo , Pele/enzimologia , Adolescente , Adulto , Idoso , Biomarcadores Tumorais , Feminino , Cabelo/metabolismo , Humanos , Concentração de Íons de Hidrogênio , Masculino , Melaninas , Pessoa de Meia-Idade , Nevo Pigmentado , Isoformas de Proteínas , Pele/metabolismo , Neoplasias Cutâneas/enzimologia , Neoplasias Cutâneas/metabolismo , Raios Ultravioleta , Adulto Jovem
17.
J Dermatol Sci ; 54(2): 106-13, 2009 May.
Artigo em Inglês | MEDLINE | ID: mdl-19250802

RESUMO

BACKGROUND: Exposure to ultraviolet (UV) radiation causes a complex cellular response, mostly mediated by the production of reactive oxygen species (ROS), which can be counteracted by exogenous treatments and endogenous mechanisms with anti-oxidant and scavenger properties. Keratinocyte growth factor (KGF/FGF7), a member of the fibroblast growth factor family, promotes epithelial growth and differentiation and is involved in cell survival after oxidant injuries. OBJECTIVE: We analyzed the role of KGF in the control of intracellular ROS production and oxidative stress after UVB exposure on KGF receptor (KGFR) transfected cells and human immortalized and primary keratinocytes. METHODS: We assessed the intracellular ROS production measuring the intensity of the oxidation-sensitive fluorescent probe 2',7'-dichlorofluorescein diacetate (DCFH-DA) by confocal microscopy, as well as the catalase activity by spectrophotometric assay. Moreover, morphological and biochemical analysis of actin cytoskeleton reorganization was evaluated as a further marker of oxidative damage. RESULTS: Our data show that KGF significantly reduces intracellular ROS generation in response to UVB, preserves the decrease of catalase activity and prevents actin cytoskeleton rearrangement. CONCLUSION: Our results provide a further evidence that KGF may be crucial for an efficient skin photoprotection, demonstrating a direct role for KGF in the reduction of intracellular ROS content following UVB exposure.


Assuntos
Actinas/metabolismo , Catalase/metabolismo , Fator 7 de Crescimento de Fibroblastos/fisiologia , Queratinócitos/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Receptor Tipo 2 de Fator de Crescimento de Fibroblastos/metabolismo , Actinas/efeitos dos fármacos , Actinas/efeitos da radiação , Animais , Catalase/efeitos dos fármacos , Catalase/efeitos da radiação , Linhagem Celular , Citoesqueleto/efeitos dos fármacos , Citoesqueleto/metabolismo , Citoesqueleto/efeitos da radiação , Regulação para Baixo/efeitos dos fármacos , Fator 7 de Crescimento de Fibroblastos/farmacologia , Humanos , Queratinócitos/efeitos dos fármacos , Queratinócitos/efeitos da radiação , Camundongos , Células NIH 3T3 , Estresse Oxidativo/efeitos dos fármacos , Estresse Oxidativo/efeitos da radiação , Receptor Tipo 2 de Fator de Crescimento de Fibroblastos/agonistas , Transfecção , Raios Ultravioleta
18.
Cell Signal ; 20(10): 1750-61, 2008 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-18602000

RESUMO

Glycogen synthase kinase 3beta (GSK3beta) is implicated in many biological events, including embryonic development, cell differentiation, apoptosis, and the insulin response. GSK3beta also plays a key role in the Wnt/beta-catenin pathway. The master regulator of the pigmentation microphthalmia-associated transcription factor (MITF) is a target for the Wnt pathway, however, to date, the regulatory role of GSK3beta in the control of melanogenesis has not been elucidated. In this study, we evaluated the effect of inhibiting GSK3beta activity on the regulation of melanocyte differentiation. Exposure of the murine melanoma cell line B16 and normal human melanocytes to GSK3beta specific inhibitors (SB216763, SB415286, BIO, and LiCl) resulted in a dose-dependent accumulation of beta-catenin. This is associated with the induction of melanocyte differentiation-associated markers such as melanin synthesis, tyrosinase activity, and expression of tyrosinase and the microphthalmia-associated transcription factor. Attenuation of GSK3beta activity has an inhibitory effect on cell growth, and this was accompanied by morphological changes. Moreover, treatment of B16 cells with a siRNA targeted against beta-catenin completely abolished the promelanogenic effect of GSK3beta inhibition, however, the overexpression of a constitutively active mutant form of beta-catenin (pCS2beta-cat-mut) only slightly increased the degree of pigmentation. These results demonstrated that GSK3beta is implicated in the regulation of melanogenesis and that pharmacological inhibition of its activity could increase melanin synthesis through mechanisms probably not restricted to Wnt/beta-catenin pathway activation.


Assuntos
Quinase 3 da Glicogênio Sintase/antagonistas & inibidores , Melaninas/biossíntese , Melanócitos/enzimologia , Melanoma Experimental/enzimologia , Animais , Diferenciação Celular/efeitos dos fármacos , Núcleo Celular/efeitos dos fármacos , Núcleo Celular/metabolismo , Proliferação de Células/efeitos dos fármacos , Relação Dose-Resposta a Droga , Imunofluorescência , Glicogênio Sintase Quinase 3 beta , Humanos , Melanócitos/citologia , Melanócitos/efeitos dos fármacos , Camundongos , Fator de Transcrição Associado à Microftalmia/metabolismo , Monofenol Mono-Oxigenase/metabolismo , Pigmentação/efeitos dos fármacos , Inibidores de Proteínas Quinases/farmacologia , Transporte Proteico/efeitos dos fármacos , RNA Interferente Pequeno/metabolismo , Regulação para Cima/efeitos dos fármacos , beta Catenina/metabolismo
19.
Free Radic Biol Med ; 45(5): 636-44, 2008 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-18538675

RESUMO

Exposure of human fibroblasts to 8-methoxypsoralen plus ultraviolet-A irradiation (PUVA) results in stress-induced cellular senescence in fibroblasts. We here studied the role of the antioxidant defense system in the accumulation of reactive oxygen species (ROS) and the effect of the antioxidants alpha-tocopherol, N-acetylcysteine, and alpha-lipoic acid on PUVA-induced cellular senescence. PUVA treatment induced an immediate and increasing generation of intracellular ROS. Supplementation of PUVA-treated fibroblasts with alpha-tocopherol (alpha-Toc), N-acetylcysteine (NAC), or alpha-lipoic acid (alpha-LA) abrogated the increased ROS generation and rescued fibroblasts from the ROS-dependent changes into the cellular senescence phenotype, such as cytoplasmic enlargement, enhanced expression of senescence-associated-beta-galactosidase and matrix-metalloproteinase-1, hallmarks of photoaging and intrinsic aging. PUVA treatment disrupted the integrity of cellular membranes and impaired homeostasis and function of the cellular antioxidant system with a significant decrease in glutathione and hydrogen peroxide-detoxifying enzymes activities. Supplementation with NAC, alpha-LA, and alpha-Toc counteracted these changes. Our data provide causal evidence that (i) oxidative stress due to an imbalance in the overall cellular antioxidant capacity contributes to the induction and maintenance of the PUVA-induced fibroblast senescence and that (ii) low molecular antioxidants protect effectively against these deleterious alterations.


Assuntos
Antioxidantes/farmacologia , Senescência Celular/efeitos dos fármacos , Senescência Celular/efeitos da radiação , Estresse Oxidativo/efeitos dos fármacos , Estresse Oxidativo/efeitos da radiação , Células Cultivadas , Criança , Pré-Escolar , Fibroblastos , Glutationa/metabolismo , Humanos , Masculino , Fenótipo , Espécies Reativas de Oxigênio/metabolismo
20.
Pigment Cell Melanoma Res ; 21(2): 200-5, 2008 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-18426413

RESUMO

UV-induced DNA damage can lead to melanoma, the most dangerous form of skin cancer. Understanding the mechanisms employed by melanocytes to protect against UV is therefore a key issue. In melanocytes, catalase is the main enzyme responsible for degrading hydrogen peroxide and we have previously shown that that low basal levels of catalase activity are associated with the light phototype in in vitro and ex vivo models. Here we investigate the possible correlation between its activity and melanogenesis in primary cultures of human melanocytes. We show that while the total melanin concentration is directly correlated to the level of pigmentation, the more the degree of pigmentation increased, the lower the proportion of pheomelanin present. Moreover, in human melanocytes in vitro, catalase-specific mRNA, protein and enzymatic activity were all directly correlated with total cellular melanin content. We also observed that immediately after a peroxidative treatment, the increase in reactive oxygen species was inversely associated with pigmentation level. Darkly pigmented melanocytes therefore possess two protective strategies represented by melanins and catalase activity that are likely to act synergistically to counteract the deleterious effects of UV radiation. By contrast, lightly pigmented melanocytes possess lower levels of melanogenic and catalase activity and are therefore more susceptible to accumulate damage after UV exposition.


Assuntos
Catalase/metabolismo , Melaninas/metabolismo , Melanócitos/metabolismo , Monofenol Mono-Oxigenase/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Pigmentação da Pele , Western Blotting , Catalase/genética , Células Cultivadas , Humanos , Melanócitos/efeitos dos fármacos , Melanócitos/enzimologia , Monofenol Mono-Oxigenase/genética , Oxidantes/farmacologia , RNA Mensageiro/biossíntese , Espécies Reativas de Oxigênio/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Pigmentação da Pele/fisiologia
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