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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.
J Clin Med ; 9(12)2020 Nov 25.
Artigo em Inglês | MEDLINE | ID: mdl-33255545

RESUMO

Infections are among the most frequent and challenging events in diabetic foot ulcers (DFUs). Pathogenic bacteria growing in biofilms within host tissue are highly tolerant to environmental and chemical agents, including antibiotics. The present study was aimed at assessing the use of silver sulfadiazine (SSD) for wound healing and infection control in 16 patients with DFUs harboring biofilm-growing Staphylococcus aureus and Pseudomonas aeruginosa. All patients received a treatment based on a dressing protocol including disinfection, cleansing, application of SSD, and application of nonadherent gauze, followed by sterile gauze and tibio-breech bandage, in preparation for toilet surgery after 30 days of treatment. Clinical parameters were analyzed by the T.I.M.E. classification system. In addition, the activity of SSD against biofilm-growing S. aureus and P. aeruginosa isolates was assessed in vitro. A total of 16 patients with S. aureus and P. aeruginosa infected DFUs were included in the study. Clinical data showed a statistically significant (p < 0.002) improvement of patients' DFUs after 30 days of treatment with SSD with significant amelioration of all the parameters analyzed. Notably, after 30 days of treatment, resolution of infection was observed in all DFUs. In vitro analysis showed that both S. aureus and P. aeruginosa isolates developed complex and highly structured biofilms. Antibiotic susceptibility profiles indicated that biofilm cultures were significantly (p ≤ 0.002) more tolerant to all tested antimicrobials than their planktonic counterparts. However, SSD was found to be effective against fully developed biofilms of both S. aureus and P. aeruginosa at concentrations below those normally used in clinical preparations (10 mg/mL). These results strongly suggest that the topical administration of SSD may represent an effective alternative to conventional antibiotics for the successful treatment of DFUs infected by biofilm-growing S. aureus and P. aeruginosa.

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.
BMC Microbiol ; 19(1): 228, 2019 10 21.
Artigo em Inglês | MEDLINE | ID: mdl-31638894

RESUMO

BACKGROUND: Infective endocarditis (IE) is associated with high rates of mortality. Prolonged treatments with high-dose intravenous antibiotics often fail to eradicate the infection, frequently leading to high-risk surgical intervention. By providing a mechanism of antibiotic tolerance, which escapes conventional antibiotic susceptibility profiling, microbial biofilm represents a key diagnostic and therapeutic challenge for clinicians. This study aims at assessing a rapid biofilm identification assay and a targeted antimicrobial susceptibility profile of biofilm-growing bacteria in patients with IE, which were unresponsive to antibiotic therapy. RESULTS: Staphylococcus aureus was the most common isolate (50%), followed by Enterococcus faecalis (25%) and Streptococcus gallolyticus (25%). All microbial isolates were found to be capable of producing large, structured biofilms in vitro. As expected, antibiotic treatment either administered on the basis of antibiogram or chosen empirically among those considered first-line antibiotics for IE, including ceftriaxone, daptomycin, tigecycline and vancomycin, was not effective at eradicating biofilm-growing bacteria. Conversely, antimicrobial susceptibility profile of biofilm-growing bacteria indicated that teicoplanin, oxacillin and fusidic acid were most effective against S. aureus biofilm, while ampicillin was the most active against S. gallolyticus and E. faecalis biofilm, respectively. CONCLUSIONS: This study indicates that biofilm-producing bacteria, from surgically treated IE, display a high tolerance to antibiotics, which is undetected by conventional antibiograms. The rapid identification and antimicrobial tolerance profiling of biofilm-growing bacteria in IE can provide key information for both antimicrobial therapy and prevention strategies.


Assuntos
Antibacterianos/farmacologia , Bactérias/classificação , Bactérias/efeitos dos fármacos , Biofilmes/efeitos dos fármacos , Endocardite Bacteriana/diagnóstico , Endocardite/microbiologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Antibacterianos/uso terapêutico , Bactérias/isolamento & purificação , Farmacorresistência Bacteriana Múltipla , Endocardite/tratamento farmacológico , Endocardite/cirurgia , Endocardite Bacteriana/tratamento farmacológico , Endocardite Bacteriana/cirurgia , Feminino , Humanos , Masculino , Testes de Sensibilidade Microbiana , Pessoa de Meia-Idade , Filogenia , Resultado do Tratamento
6.
J Nat Prod ; 81(1): 49-56, 2018 01 26.
Artigo em Inglês | MEDLINE | ID: mdl-29300477

RESUMO

Bioassay-guided fractionation of the methanol extract of the shoots of Myrsine africana led to the isolation of the new compound myricetin 3-O-(2″,4″-di-O-acetyl)-α-l-rhamnopyranoside (9) and 11 known compounds. The known compounds quercetin 3-O-(3″,4″-di-O-acetyl)-α-l-rhamnopyranoside (8), rutin (10), quercetin 3-O-α-l-rhamnopyranoside (11), and myricetin 3-O-α-l-rhamnopyranoside (12) are reported for the first time from the methanol extract of the shoots of M. africana. Compounds 10 and 12 showed significant inhibition of tyrosinase with 50% inhibition (IC50 values) of the enzyme at 0.13 ± 0.003 and 0.12 ± 0.002 mM, respectively, which was supported by the docking fitness scores obtained through molecular docking analysis. In addition, compounds 1-12 displayed significant antioxidant activity with IC50 values ranging 1.90 to 3.90 µM.


Assuntos
Agaricales/efeitos dos fármacos , Flavonoides/química , Flavonoides/farmacologia , Glicosídeos/química , Glicosídeos/farmacologia , Myrsine/química , Antioxidantes/química , Antioxidantes/farmacologia , Simulação de Acoplamento Molecular/métodos , Extratos Vegetais/química , Extratos Vegetais/farmacologia , Quercetina/análogos & derivados , Quercetina/química , Quercetina/farmacologia
7.
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
8.
Sci Rep ; 7(1): 13663, 2017 10 20.
Artigo em Inglês | MEDLINE | ID: mdl-29057950

RESUMO

Vitiligo is characterized by death or functional defects of epidermal melanocytes through still controversial pathogenic process. Previously, we showed that mitochondria-driven pre-senescent phenotype diminishes the capability of vitiligo melanocytes to cope with stressful stimuli. In the current study, we investigated markers of mitochondrial energy metabolism including the PGC1a axis, and then we determined the index of mitochondrial impairment using a cytomic approach. We found in cultured epidermal vitiligo melanocytes, compared to healthy ones, low ATP, increased proton leakage, and altered expression of several glycolytic enzymes (hexokinase II, pyruvic dehydrogenase kinase 1 and pyruvic kinase M2), We suggest that the low ATP production may be sufficient in steady-state conditions but it is unable to cover further needs. We also found in vitiligo melanocyrtes hyper-activation of the PGC1α axis, finalized to counteract the energy defect. Cytomic analysis, supported by MitoTracker Red pattern and ex-vivo immunohistochemistry, suggested an increased mitochondrial mass, possibly useful to ensure the essential ATP level. Finally, pharmacological cardiolipin stabilization reverted the energetic impairment, confirming the initial mitochondrial role. In conclusion, we report new insight in the pathogenetic mechanism of viitligo and indicate that the mitochondrial failure rescue by cardiolipin manipulation may be a new intriguing target in treatment development.


Assuntos
Cardiolipinas/metabolismo , Mitocôndrias/metabolismo , Vitiligo/metabolismo , Trifosfato de Adenosina/metabolismo , Epiderme/metabolismo , Epiderme/patologia , Glucose/metabolismo , Humanos , Melanócitos/metabolismo , Melanócitos/patologia , Mitocôndrias/patologia , Cultura Primária de Células , RNA Mensageiro/metabolismo , Vitiligo/patologia
9.
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
10.
Proc Natl Acad Sci U S A ; 113(30): E4397-406, 2016 07 26.
Artigo em Inglês | MEDLINE | ID: mdl-27412859

RESUMO

The intracellular serine amidase, fatty acid amide hydrolase (FAAH), degrades a heterogeneous family of lipid-derived bioactive molecules that include amides of long-chain fatty acids with taurine [N-acyl-taurines (NATs)]. The physiological functions of the NATs are unknown. Here we show that genetic or pharmacological disruption of FAAH activity accelerates skin wound healing in mice and stimulates motogenesis of human keratinocytes and differentiation of human fibroblasts in primary cultures. Using untargeted and targeted lipidomics strategies, we identify two long-chain saturated NATs-N-tetracosanoyl-taurine [NAT(24:0)] and N-eicosanoyl-taurine [NAT(20:0)]-as primary substrates for FAAH in mouse skin, and show that the levels of these substances sharply decrease at the margins of a freshly inflicted wound to increase again as healing begins. Additionally, we demonstrate that local administration of synthetic NATs accelerates wound closure in mice and stimulates repair-associated responses in primary cultures of human keratinocytes and fibroblasts, through a mechanism that involves tyrosine phosphorylation of the epidermal growth factor receptor and an increase in intracellular calcium levels, under the permissive control of transient receptor potential vanilloid-1 receptors. The results point to FAAH-regulated NAT signaling as an unprecedented lipid-based mechanism of wound-healing control in mammalian skin, which might be targeted for chronic wound therapy.


Assuntos
Pele/metabolismo , Taurina/metabolismo , Cicatrização , Amidoidrolases/genética , Amidoidrolases/metabolismo , Animais , Células Cultivadas , Fibroblastos/efeitos dos fármacos , Fibroblastos/metabolismo , Humanos , Recém-Nascido , Queratinócitos/efeitos dos fármacos , Queratinócitos/metabolismo , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout , Pele/efeitos dos fármacos , Pele/patologia , Especificidade por Substrato , Taurina/química , Taurina/farmacologia
11.
Cell Signal ; 26(4): 716-23, 2014 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-24412755

RESUMO

Transfer of melanin-containing melanosomes from melanocytes to neighboring keratinocytes results in skin pigmentation. Pharmacological modulation of melanosomal transfer has recently gained much attention as a strategy for modifying normal or abnormal pigmentation. In this study, while investigating the impact of pyridinyl imidazole (PI) compounds, a class of p38 MAPK inhibitors, on melanocyte differentiation we observed that some, but not all PIs interfere with the physiological melanosome sorting producing a strong retention of melanin in the intracellular compartment associated with a general reduction of melanin synthesis. Electron microscopy studies illustrated an accumulation of melanosomes inside melanocytes with enrichment in immature melanosome at stages II and III at the end of dendrites. We identified cyclin G-associated kinase GAK, a protein expressed ubiquitously in various tissues, as the off-target responsible of intracellular melanin accumulation and we report evidence that reduced GAK-dependent cathepsin maturation is implicated in melanosome sorting deficiency. The co-regulation of GAK and cathepsin B and L expression with the melanogenic biosynthetic pathway in normal human melanocytes as well as in B16-F0 melanoma cells strengthen the idea that these proteins represent new possible targets for prevention and treatment of irregular pigmentation.


Assuntos
Imidazóis/farmacologia , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Melanócitos/efeitos dos fármacos , Proteínas Serina-Treonina Quinases/metabolismo , Animais , Catepsinas/metabolismo , Células Cultivadas , Humanos , Imidazóis/química , Peptídeos e Proteínas de Sinalização Intracelular/antagonistas & inibidores , Peptídeos e Proteínas de Sinalização Intracelular/genética , Melaninas/biossíntese , Melanócitos/citologia , Melanócitos/metabolismo , Melanossomas/metabolismo , Melanossomas/ultraestrutura , Camundongos , Monofenol Mono-Oxigenase/metabolismo , Proteínas Serina-Treonina Quinases/antagonistas & inibidores , Proteínas Serina-Treonina Quinases/genética , Piridinas/química , Interferência de RNA , RNA Interferente Pequeno/metabolismo , Pigmentação da Pele , Proteínas Quinases p38 Ativadas por Mitógeno/antagonistas & inibidores , Proteínas Quinases p38 Ativadas por Mitógeno/genética , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo
12.
J Invest Dermatol ; 134(4): 1001-1011, 2014 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-24166135

RESUMO

Peroxisome proliferator-activated receptor γ (PPARγ) antagonizes inflammatory signals by interfering with NF-κB nuclear translocation. Consistently, PPARγ agonists have been proposed in various inflammatory skin disorders, but their wide use has been limited by severe side effects. Classes of compounds with specific PPARγ agonism have been designed to selectively target inflammatory pathways. Among these compounds, GED-0507-34L has been developed and recently used in phase II clinical trials for inflammatory bowel diseases. This study was aimed at assessing the role of GED-0507-34L in preclinical models of inflammatory skin diseases. The compound modulated PPARγ function and suppressed the inflammatory process inhibiting NF-κB nuclear translocation with the consequent reduction of inflammatory cytokines expression, such as IL-6, IL-8, IL-12, IL-21, IL-23, tumor necrosis factor-α (TNF-α), and cyclooxygenase-2 (COX-2) in normal human keratinocytes and lymphocytes treated with lipopolysaccharide (LPS) or TNF-α. Moreover, an altered proliferation and expression of differentiation markers induced by TNF-α were also counteracted. In psoriasis-like skin lesions elicited in mice by IL-21, topical application of GED-0507-34L reduced cellular infiltrate and epidermal hyperplasia, normalizing the differentiation process. The results indicate that GED-0507-34L possesses anti-inflammatory properties useful for the management of patients with inflammatory skin diseases including psoriasis. Phase I trial on patients is ongoing.


Assuntos
Transporte Ativo do Núcleo Celular/efeitos dos fármacos , Compostos de Anilina/uso terapêutico , Inflamação/tratamento farmacológico , PPAR gama/metabolismo , Fenilpropionatos/uso terapêutico , Propionatos/uso terapêutico , Pele/patologia , Compostos de Anilina/química , Animais , Biópsia , Adesão Celular , Proliferação de Células , Células Cultivadas , Ciclo-Oxigenase 2/metabolismo , Citocinas/metabolismo , Epiderme/metabolismo , Humanos , Queratinócitos/citologia , Lipopolissacarídeos/química , Camundongos , Camundongos Endogâmicos C57BL , NF-kappa B/metabolismo , Fenilpropionatos/química , Propionatos/química , Ligação Proteica , Psoríase/metabolismo , Interferência de RNA , Pele/efeitos dos fármacos
13.
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
14.
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
15.
PLoS One ; 7(10): e48097, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-23094106

RESUMO

In filamentous fungi, peroxisomes are crucial for the primary metabolism and play a pivotal role in the formation of some secondary metabolites. Further, peroxisomes are important site for fatty acids ß-oxidation, the formation of reactive oxygen species and for their scavenging through a complex of antioxidant activities. Oxidative stress is involved in different metabolic events in all organisms and it occurs during oxidative processes within the cell, including peroxisomal ß-oxidation of fatty acids. In Aspergillus flavus, an unbalance towards an hyper-oxidant status into the cell is a prerequisite for the onset of aflatoxin biosynthesis. In our preliminary results, the use of bezafibrate, inducer of both peroxisomal ß-oxidation and peroxisome proliferation in mammals, significantly enhanced the expression of pex11 and foxA and stimulated aflatoxin synthesis in A. flavus. This suggests the existence of a correlation among peroxisome proliferation, fatty acids ß-oxidation and aflatoxin biosynthesis. To investigate this correlation, A. flavus was transformed with a vector containing P33, a gene from Cymbidium ringspot virus able to induce peroxisome proliferation, under the control of the promoter of the Cu,Zn-sod gene of A. flavus. This transcriptional control closely relates the onset of the antioxidant response to ROS increase, with the proliferation of peroxisomes in A. flavus. The AfP33 transformant strain show an up-regulation of lipid metabolism and an higher content of both intracellular ROS and some oxylipins. The combined presence of a higher amount of substrates (fatty acids-derived), an hyper-oxidant cell environment and of hormone-like signals (oxylipins) enhances the synthesis of aflatoxins in the AfP33 strain. The results obtained demonstrated a close link between peroxisome metabolism and aflatoxin synthesis.


Assuntos
Aflatoxinas/biossíntese , Aspergillus flavus/genética , Ácidos Graxos/metabolismo , Proteínas Fúngicas/genética , Regulação Fúngica da Expressão Gênica/efeitos dos fármacos , Peroxissomos/metabolismo , Antioxidantes/metabolismo , Aspergillus flavus/efeitos dos fármacos , Aspergillus flavus/metabolismo , Bezafibrato/farmacologia , Proteínas Fúngicas/metabolismo , Vetores Genéticos , Hipolipemiantes/farmacologia , Metabolismo dos Lipídeos/efeitos dos fármacos , Oxirredução , Estresse Oxidativo , Regiões Promotoras Genéticas , Espécies Reativas de Oxigênio/metabolismo , Superóxido Dismutase/genética , Superóxido Dismutase/metabolismo , Tombusvirus/genética
16.
J Invest Dermatol ; 131(6): 1182-5, 2011 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-21566575

RESUMO

Cutaneous pigmentation is regulated by a complex melanogenic network in which skin cells synthesize growth factors and cytokines. Mutations in genes encoding these regulators modify their expression and/or functionality, leading to altered signaling pathways and contributing to altered skin phenotypes. In this issue, Amyere et al. report a genome-wide analysis of seven families with familial progressive hyperpigmentation and hypopigmentation, identifying three new mutations in KITLG. The study underlines the relevance of investigating candidate genes implicated in the onset of pigmentary disorders. Furthermore, Amyere et al. suggest that different pigmentary diseases can result from the same mutation or different mutations in the same gene, and they offer hope for the development of new and efficacious treatment strategies.


Assuntos
Hiperpigmentação/genética , Hipopigmentação/genética , Proteínas Proto-Oncogênicas c-kit/fisiologia , Pigmentação da Pele/genética , Fator de Células-Tronco/fisiologia , Estudo de Associação Genômica Ampla , Humanos , Hiperpigmentação/etiologia , Hipopigmentação/etiologia , Mutação , Neurofibromatose 1/genética , Transdução de Sinais , Fator de Células-Tronco/genética
17.
Exp Dermatol ; 19(9): 813-20, 2010 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-20545756

RESUMO

Azelaic acid (AzA), a nine-carbon dicarboxylic acid, is an agent for the topical treatment of acne. It has also been shown to be effective in rosacea; however, the mechanism of action has not been clarified. Because inflammation is a common feature of both conditions, we investigated the effects of azelaic acid on the inflammatory response of normal human keratinocytes to ultraviolet B light, which is a photosensitizer agent in rosacea. AzA, at 20 mM, a concentration achievable following topical application of a 15% gel, suppresses ultraviolet B light-induced interleukins-1beta, -6 and tumor necrosis factor-alpha mRNA expression and protein secretion. Mechanistically, azelaic acid significantly reduced the ultraviolet B light-induced nuclear translocation of nuclear factor kB p65 subunit and the phosphorylation of the p38 mitogen and stress-activated protein kinase. Moreover, as peroxisome proliferators-activated receptor gamma, (PPARgamma) which has a crucial role in the control of inflammation, is activated by fatty acids and products of lipid peroxidation, we further investigated the effect of azelaic acid on the expression of this nuclear receptor. AzA induced peroxisome proliferators-activated receptor-gamma mRNA and its transcriptional activity. The PPARgamma antagonist GW9662 abrogated the inhibitory effects of AzA on the UVB-induced pro-inflammatory cytokines release and on the cell proliferation. Our study provides new insights into the molecular mechanisms of the activity of azelaic acid and lands additional evidences for its therapeutic effects on inflammatory skin diseases, such as rosacea.


Assuntos
Acne Vulgar/tratamento farmacológico , Fármacos Dermatológicos/farmacologia , Ácidos Dicarboxílicos/farmacologia , Queratinócitos/efeitos dos fármacos , NF-kappa B/metabolismo , Células Cultivadas , Citocinas/metabolismo , Fármacos Dermatológicos/uso terapêutico , Ácidos Dicarboxílicos/uso terapêutico , Avaliação Pré-Clínica de Medicamentos , Genes Reporter , Humanos , Queratinócitos/metabolismo , Queratinócitos/efeitos da radiação , PPAR gama/agonistas , PPAR gama/genética , PPAR gama/metabolismo , Fosforilação/efeitos dos fármacos , RNA Mensageiro/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Rosácea/tratamento farmacológico , Transcrição Genética/efeitos dos fármacos , Raios Ultravioleta , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo
18.
Transl Oncol ; 3(2): 80-90, 2010 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-20360932

RESUMO

The paracrine networks of the human melanoma microenvironment are able to influence tumor growth and progression. Among the paracrine growth factors involved in skin homeostasis, the KGF/FGF7 secreted by dermal fibroblasts promotes the epidermal proliferation and differentiation as well as the release from keratinocytes of other paracrine mediators. To evaluate the possible role played by KGF in affecting the behavior of different subtypes of melanoma carrying activating mutations or overexpression of the SCF receptor c-KIT, we used human melanoma cell lines, characterized by different expression levels of c-KIT and opposing responsivity to SCF, and HaCaT keratinocytes. Quantitative real-time reverse transcription-polymerase chain reaction assay and ELISA test on KGF-treated keratinocytes showed enhanced expression and secretion of SCF in response to KGF and dependent on functional KGF receptor. Immunofluorescence microscopy and biochemical analysis showed, in one of the selected melanoma cell models, SCF-dependent c-KIT activation induced by stimulation with the culture supernatants collected from KGF-treated keratinocytes. In keratinocyte-melanoma cocultures stained for the Ki67 proliferation marker, incubation with KGF induced enhanced growth not only of the keratinocytes but also of the melanoma cells, which could be blocked by the c-KIT inhibitor imatinib, demonstrating the establishment of a KGF-induced paracrine signaling network owing to the coexpression of biologically active SCF released from keratinocytes and functional c-KIT on melanoma cells.

19.
Growth Factors ; 27(6): 448-55, 2009 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-19919532

RESUMO

Bovine colostrum represents a rich source of growth factors, which are known to play a central role in wound healing. The aim of our study was to investigate the possible mitogenic and motogenic effects induced by colostrum on human keratinocytes. Cell proliferation evaluated by 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide test and 5-Bromo-2'-deoxyuridine incorporation revealed that colostrum exerts a growth promoting activity. Scratch assay and immunofluorescence of actin cytoskeleton showed its effectiveness also in inducing cell migration. Furthermore, colostrum treatment increases the levels of tyrosine phosphorylated proteins and the activated forms of the extracellular signal-regulated kinases 1 and 2 and such effects appear to be repressed by the tyrosine kinase inhibitor genistein. Our results indicate that the biological activities of colostrum are specifically mediated by the growth factor-induced activation of tyrosine kinase receptors and underline the relevance of the synergistic action exerted by the growth factors in stimulating keratinocyte proliferation and migration essential for tissue repair.


Assuntos
Movimento Celular/fisiologia , Proliferação de Células/efeitos dos fármacos , Colostro/fisiologia , Queratinócitos/efeitos dos fármacos , Animais , Bovinos , Linhagem Celular , Feminino , Regulação da Expressão Gênica , Humanos , Queratinócitos/citologia , Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Proteína Quinase 3 Ativada por Mitógeno/metabolismo , Gravidez , Receptores Proteína Tirosina Quinases/metabolismo , Cicatrização/fisiologia
20.
Eur J Dermatol ; 19(5): 469-73, 2009.
Artigo em Inglês | MEDLINE | ID: mdl-19505863

RESUMO

Two patients with a generalized, progressive dyschromatosis disorder are described and investigated as a model to study the role of fibroblast-derived mediators on skin pigmentation. The patients (father and daughter) had had a widespread hyperpigmentation since early life which then progressively worsened with the appearance of hyperpigmented macules, café-au-lait macules and freckles, also involving the lips, palms and soles, intermixed with small hypopigmented spots. These features resembled those of familial progressive hyperpigmentation (FPH). Histology revealed a normal epidermis with pronounced keratinocyte hyperpigmentation and the presence of dermal melanophages. Ultrastructural analysis showed basal and suprabasal keratinocytes enriched in melanosome complexes. Immunohistochemical staining displayed an increased expression of hepatocyte growth factor (HGF), stem cell factor (SCF) and keratinocyte growth factor (KGF) in fibroblast-like cells of the upper dermis in hyperpigmented lesions of both patients, compared to control healthy skin. Our data suggest that a persistent activation of fibroblasts abnormally stimulating melanocyte functions is involved in hyperpigmentation disorders.


Assuntos
Fator 7 de Crescimento de Fibroblastos/fisiologia , Fibroblastos/química , Fator de Crescimento de Hepatócito/fisiologia , Hiperpigmentação/genética , Fator de Células-Tronco/fisiologia , Adulto , Feminino , Fator 7 de Crescimento de Fibroblastos/análise , Fator de Crescimento de Hepatócito/análise , Humanos , Hiperpigmentação/etiologia , Imuno-Histoquímica , Masculino , Pessoa de Meia-Idade , Fator de Células-Tronco/análise
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