No / P-cadherin serves as a major topobiological cue in mammalian epithelium. In human hair follicles (HFs), it is prominently expressed in the inner hair matrix that harbors the HF pigmentary unit. However, the role of P-cadherin in normal human pigmentation remains unknown. As patients with mutations in the gene that encodes P-cadherin show hypotrichosis and fair hair, we explored the hypothesis that P-cadherin may control HF pigmentation. When P-cadherin was silenced in melanogenically active organ-cultured human scalp HFs, this significantly reduced HF melanogenesis and tyrosinase activity as well as gene and/or protein expression of gp100, stem cell factor, c-Kit, and microphthalmia-associated transcription factor (MITF), both in situ and in isolated human HF melanocytes. Instead, epidermal pigmentation was unaffected by P-cadherin knockdown in organ-cultured human skin. In hair matrix keratinocytes, P-cadherin silencing reduced plasma membrane β-catenin, whereas glycogen synthase kinase 3 beta (GSK3β) and phospho-β-catenin expression were significantly upregulated. This suggests that P-cadherin-GSK3β/Wnt signaling is required for maintaining the expression of MITF to sustain intrafollicular melanogenesis. Thus, P-cadherin-mediated signaling is a melanocyte subtype-specific topobiological regulator of normal human pigmentation, possibly via GSK3β-mediated canonical Wnt signaling.

No / Although we have made significant progress in understanding the regulation of the UVR-exposed epidermal-melanin unit, we know relatively little about how human hair follicle pigmentation is regulated. Progress has been hampered by gaps in our knowledge of the hair growth cycle’s controls, to which hair pigmentation appears tightly coupled. However, pigment cell researchers may have overly focused on the follicular melanocytes of the nocturnal and UVR-shy mouse as a proxy for human epidermal melanocytes. Here, I emphasize the epidermis-follicular melanocyte pluralism of human skin, as research models for vitiligo, alopecia areata and melanoma, personal care/cosmetics innovation. Further motivation could be in finding answers to why hair follicle and epidermal pigmentary units remain broadly distinct? Why melanomas tend to originate from epidermal rather than follicular melanocytes? Why multiple follicular melanocyte sub-populations exist? Why follicular melanocytes are more sensitive to aging influences? In this perspective, I attempt to raise the status of the human hair follicle melanocyte and highlight some species-specific issues involved which the general reader of the pigmentation literature (with its substantial mouse-based data) may not fully appreciate.

Considerando o melanoma a forma mais agressivas de câncer de pele e mais resistente aos tratamentos convencionais, o presente estudo teve como objetivo avaliar a atividade de um novo complexo de platina (II) com derivado de hidantoína (CX42) em células de melanoma humano e murino. Foram utilizados também para comparação células da pele (fibroblastos, queratinócitos e melanócitos) e os compostos cisplatina (CIS) e complexo de hidantoína isolado (NN10). Ensaios de viabilidade, ciclo e morte celular foram realizados. Investigou-se também a atividade da enzima tirosinase, principal enzima que regula a síntese de melanina durante o processo conhecido como melanogênese. Como resultados, obteve-se a diminuição da viabilidade celular e parada de ciclo na fase G0/G1 nas células de melanoma, principalmente na linhagem murina B16F10, frente ao composto CX42. Em células B16F10, foi possível observar estímulo na melanogênese, com aumento da atividade da enzima tirosinase. O CX42 apresentou uma atividade com efeito citostático nas células de melanoma, não sendo observado efeito citotóxico nas células da pele. Ainda, com a prévia estimulação da melanogênese, o CX42 apresentou-se mais efetivo, aumentando a morte celular em 40% por apoptose. Portanto, conclui-se que o CX42 diminuiu a viabilidade celular e seu efeito foi mais intenso quando as células apresentavam-se pigmentadas, demonstrando indução da tirosinase e da morte celular, atributo importante para potenciais novos fármacos anti-melanoma. / Since melanoma is one of the most aggressive forms of skin cancer and more resistant to conventional treatments, the present study aimed to evaluate the activity of a new platinum complex (II) with hydantoin derivative (CX42) in melanoma cells human and murine. Skin cells (fibroblasts, keratinocytes and melanocytes) were used as control and the compounds cisplatin (CIS) and hydantoin compound alone (NN10) were also evaluated. Viability assays, cell cycle analysis and cell death characterization were performed. Likewise, the activity of tyrosinase, the key enzyme that regulates melanin synthesis during the process known as melanogenesis, was also investigated. The results demonstrated a reduction in cell viability and cell cycle arrest in G0/G1 phase of murine melanoma B16F10 treated with CX42. In B16F10 cells, it was also observed melanogenesis stimulation with increased activity of tyrosinase. The CX42 showed a selective cytostatic effect on melanoma cells, however no toxic effects were observed in skin cells. In addition, with prior stimulation of melanogenesis, the CX42 demonstrated to be more effective, increasing apoptosis cell death in 40%. Therefore, the results demonstrated that CX42 decreased cell viability and the intensity of its effect is associated with cell pigmentation, demonstrating an association between tyrosinase high activity and induction of cell death which is an important attribute for new potential anti - tumoral drugs.

Considerando o melanoma a forma mais agressivas de câncer de pele e mais resistente aos tratamentos convencionais, o presente estudo teve como objetivo avaliar a atividade de um novo complexo de platina (II) com derivado de hidantoína (CX42) em células de melanoma humano e murino. Foram utilizados também para comparação células da pele (fibroblastos, queratinócitos e melanócitos) e os compostos cisplatina (CIS) e complexo de hidantoína isolado (NN10). Ensaios de viabilidade, ciclo e morte celular foram realizados. Investigou-se também a atividade da enzima tirosinase, principal enzima que regula a síntese de melanina durante o processo conhecido como melanogênese. Como resultados, obteve-se a diminuição da viabilidade celular e parada de ciclo na fase G0/G1 nas células de melanoma, principalmente na linhagem murina B16F10, frente ao composto CX42. Em células B16F10, foi possível observar estímulo na melanogênese, com aumento da atividade da enzima tirosinase. O CX42 apresentou uma atividade com efeito citostático nas células de melanoma, não sendo observado efeito citotóxico nas células da pele. Ainda, com a prévia estimulação da melanogênese, o CX42 apresentou-se mais efetivo, aumentando a morte celular em 40% por apoptose. Portanto, conclui-se que o CX42 diminuiu a viabilidade celular e seu efeito foi mais intenso quando as células apresentavam-se pigmentadas, demonstrando indução da tirosinase e da morte celular, atributo importante para potenciais novos fármacos anti-melanoma. / Since melanoma is one of the most aggressive forms of skin cancer and more resistant to conventional treatments, the present study aimed to evaluate the activity of a new platinum complex (II) with hydantoin derivative (CX42) in melanoma cells human and murine. Skin cells (fibroblasts, keratinocytes and melanocytes) were used as control and the compounds cisplatin (CIS) and hydantoin compound alone (NN10) were also evaluated. Viability assays, cell cycle analysis and cell death characterization were performed. Likewise, the activity of tyrosinase, the key enzyme that regulates melanin synthesis during the process known as melanogenesis, was also investigated. The results demonstrated a reduction in cell viability and cell cycle arrest in G0/G1 phase of murine melanoma B16F10 treated with CX42. In B16F10 cells, it was also observed melanogenesis stimulation with increased activity of tyrosinase. The CX42 showed a selective cytostatic effect on melanoma cells, however no toxic effects were observed in skin cells. In addition, with prior stimulation of melanogenesis, the CX42 demonstrated to be more effective, increasing apoptosis cell death in 40%. Therefore, the results demonstrated that CX42 decreased cell viability and the intensity of its effect is associated with cell pigmentation, demonstrating an association between tyrosinase high activity and induction of cell death which is an important attribute for new potential anti - tumoral drugs.

No / This study shows that prostaglandins in human FM55 melanoma cells and epidermal melanocytes are produced by COX-1. Prostaglandin production in FM55 melanoma cells was unrelated to that of melanin suggesting that the two processes can occur independently. ¿-Melanocyte stimulating hormone (¿-MSH), which had no effect on melanin production in FM55 cells, stimulated PGD2 production in these cells without affecting PGE2. While cAMP pathways may be involved in regulating PGD2 production, our results suggest that ¿-MSH acts independently of cAMP, possibly by regulating the activity of lipocalin-type PGD synthase. This ¿-MSH-mediated effect may be associated with its role as an immune modulator. / The Wellcome Trust

Yes / Skin pigmentation is directed by epidermal-melanin units, characterized by long-lived and dendritic epidermal melanocytes (MC) that interact with viable keratinocytes (KC) to contribute melanin to the epidermis. Previously we reported that MC:KC contact is required for melanosome transfer, that this can be enhanced by filopodial and by UVR/UVA irradiation, which can up-regulate melanosome transfer via Myosin X-mediated control of MC filopodia. Both MC and KC express Ca2+-dependent E-cadherins. These homophilic adhesion contacts induce transient increases in intra-KC Ca2+, while ultraviolet radiation (UVR) raises intra-MC Ca2+ via calcium selective ORAI1 ion channels; both are associated with regulating melanogenesis. However, how Ca2+ triggers melanin transfer remains unclear, and here we evaluated the role of E-Cadherin in UVR-mediated melanin transfer in human skin cells. MC and KC in human epidermis variably express filopodia-associated E-Cadherin, Cdc42, VASP and β-catenin, all of which were upregulated by UVR/UVA in human MC in vitro. Knockdown of E-cadherin revealed that this cadherin is essential for UVR-induced MC filopodia formation and melanin transfer. Moreover, Ca2+ induced a dose-dependent increase in filopodia formation and melanin transfer, as well as increased β-catenin, Cdc42, Myosin X, and E-Cadherin expression in these skin cells. Together these data suggest that filopodial proteins and E-Cadherin, which are upregulated by intracellular (UVR-stimulated) and extracellular Ca2+ availability, are required for filopodia formation and melanin transfer. This may open new avenues to explore how Ca2+ signalling influences human pigmentation.

Notre travail s’est intéressé à l’albinisme oculocutané en étudiant ses aspects clinico- moléculaires. Malgré l’analyse approfondie des gènes connus d’albinisme oculocutané, 15 % des patients restent sans mutation identifiée indiquant que les mutations sont situées dans des régions géniques non analysées par les techniques classiques de diagnostic moléculaire, ou qu’il existe d’autres gènes d’albinisme oculocutané. Nous avons établi une base de données clinico- biologiques décrivant les caractéristiques de plus de 400 patients analysés. Des outils de diagnostic moléculaire ont été développés à la recherche de mutations situées dans les introns et les régions régulatrices et de réarrangements géniques. Différentes stratégies ont également été utilisées pour rechercher des gènes candidats. La puce à façon a permis l’identification de grands réarrangements dans les gènes TYR, OCA2 et SLC45A2 et un réarrangement complexe du gène OCA2 chez 2 patients non apparentés. L'analyse de gènes candidats nous a permis d'identifier, chez 5 patients non apparentés présentant un albinisme oculocutané non syndromique, des mutations dans le gène SLC24A5, très récemment associé à l’AOC6. Le séquençage d’exome de 6 patients a mis en évidence des gènes candidats pour lesquels des analyses complémentaires sont poursuivies afin de confirmer leur implication dans la pathogenèse de l’AOC.Les résultats de ce travail permettent de redéfinir les aspects cliniques et moléculaires de l’AOC, d’identifier de nouveaux mécanismes moléculaires à l’origine de l’AOC ainsi que des gènes candidats dont la fonction dans le développement pigmentaire reste à élucider. L’identification de nouveaux gènes impliqués dans l’AOC pourrait permettre de mieux comprendre et de mieux prendre en charge les patients avec un AOC. / Our work focused on oculocutaneous albinism (OCA) by studying its clinical and molecular aspects. Despite a thorough analysis of the known genes involved in oculocutaneous albinism, 15% of patients remain without diagnostic at the molecular level indicating that mutations are located in unexplored regions and are undetected by standard techniques or that other genes are involved in albinism. We established a clinicomolecular database describing more than 400 patients and developped molecular tools in order to improve molecular diagnostic including a custom high resolution array-CGH dedicated to the four OCA genes (TYR, OCA2, TYRP1 and SLC45A2). We also used different strategies to identify new genes. Array-CGH allows us to detect large deletion in TYR, OCA2 and SLC45A2 and a complexe rearrangement in OCA2 in 2 unrelated patients. We identified, in 5 patients presenting with a non syndromic OCA, mutations in SLC24A5, recently associated with OCA6. Exome sequencing of 6 different patients allows us to identify candidate genes, for which further studies are required to confirm their involvement in OCA pathogenesis. The results of this work allowed us to delineate clinical and genetics aspects of more than 400 OCA patients and to identify new molecular mechanisms leading to OCA and candidates genes for which exact nature of their functions has to be understood. Giving the complexity of pigmentary system development and its regulation, identification of new genes leading to OCA could help to better understand OCA and take care of patients

Submitted by Irene Nascimento (irene.kessia@ufpe.br) on 2016-09-08T19:08:46Z No. of bitstreams: 2 license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) Tese_Karina_Final_CD.pdf: 2797138 bytes, checksum: 5e86db88edfdb58a92aceb27cf908735 (MD5) / Made available in DSpace on 2016-09-08T19:08:46Z (GMT). No. of bitstreams: 2 license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) Tese_Karina_Final_CD.pdf: 2797138 bytes, checksum: 5e86db88edfdb58a92aceb27cf908735 (MD5) Previous issue date: 2015-10-16 / A partir dos parâmetros eletrônicos obtidos com a Teoria do Funcional Densidade, realizamos um estudo Quantitativo da Relação Estrutura Atividade (QSAR) de derivados de cetonatiossemicarbazonas e de ácidos kójico e benzoico para analisar o potencial de inibição da melanogênese destes compostos. Utilizando técnicas computacionais em conjunção com uma Regressão linear múltipla, obtivemos uma expressão capaz de prever a concentração inibitória (IC50) destes compostos e dos demais aqui propostos. Para a previsão da IC50 foram utilizados os seguintes parâmetros eletrônicos e físico-químicos: afinidade eletrônica (EA), gap de energia (HHL), momento de dipolo (μ) e o logaritmo do coeficiente de partição [octanol/água](LogP). Para as cetonatiossemicarbazonas os descritores eletrônicos que proporcionaram uma boa correlação linear com a IC50 experimental foram: a carga atômica do nitrogênio N2 e a EA. Para as demais moléculas avaliadas nesta pesquisa, além desses parâmetros, foram incluídos o potencial de ionização (IP), a energia do atracamento molecular (G), eletronegatividade absoluta (), dureza (), maciez (S), logaritmo do coeficiente de solubilidade (LogS), volume molar (VM) e o coeficiente de Hansch (). Como resultado da QSAR, os descritores que proporcionaram a melhor correlação linear com a IC50 experimental foram: a HHL e o VM. Na análise QSAR dos derivados dos ácidos kójico e benzoico foi utilizado um conjunto de treinamento formado por dez moléculas e um grupo de teste constituído por duas moléculas para uma validação cruzada tipo boostrap. Os cálculos de G se restringiram a encontrar o valor da energia livre de interação dos derivados dos ácidos kójico e benzoico e da enzima tirosinase por meio da formação do complexo ligante-tirosinase. Os valores das energias de interação obtidos para as moléculas propostas se revelaram promissores, visto que apresentaram valores mais baixos do que o obtido para o complexo ácido kójico-tirosinase. Em todas as análises QSAR, os valores dos parâmetros estatísticos de validação, como coeficiente de correlação, desvio-padrão, teste de Fischer e do nível geral de confiabilidade do modelo, estão dentro do esperado para um bom modelo estatístico. Os modelos obtidos fornecem uma boa previsão das atividades biológicas investigadas neste trabalho apontam para novos compostos candidatos com potencial para inibição da melanogênese. / We have carried ant studies of QSAR using electronic structure derived parameters be the means of Density Functional Theory (DFT) calculations for derivatives of ketonethiosemicarbazones, kojic acid and benzoic acid. The aim was to evaluate the melanogenesis inhibiting potential of these compounds, by using this procedure and performing a Multiple Linear Regression we obtained an expression able to predict the inhibitory concentration (IC50) of the compounds studied here. In predicting the IC50 we used the electronic and physical-chemical parameters of electron affinity (EA), energy gap (HHL), dipole of moment (μ) and the [octanol/water] logarithm of the partition coefficient (LogP). For ketonethiosemicarbazones the electronic parameters that provide a good linear correlation with experimental IC50 are the atomic charge of nitrogen N2 and the EA. For the other molecules analyzed in this study, in addition to these parameters it is included ionization potential (IP), molecular docking of energy (G), absolute electronegativity (), hardness(), softness (S), partition coefficient of molar solubility (LogS), molar volume (VM) and Hansch coefficient (). We found that the parameters leading to the best linear correlation with experimental IC50 are the interaction energy and the molar volume. In the QSAR analysis of derivatives of the benzoic and kojic acids it is used a training group formed by ten molecules and a test group formed of two molecules to realize a bootstrap-type cross validation. The calculations of the molecular docking are restricted to values of free energy of derivatives of the benzoic and kojic acids and to the enzyme tyrosinase forming the tyrosinase-ligand complex. In all QSAR analysis, the statistical validation such as correlation coefficient, standard deviation, Fisher test and the model reliability are in that range expected for a good statistical model. As a conclusion, we show that our model gives a good prediction of the biological activities, which allow us to indicate new compounds with potential in inhibiting melanogenesis.

Submitted by Kamila Costa (kamilavasconceloscosta@gmail.com) on 2015-06-29T18:32:01Z No. of bitstreams: 2 Dissertação-Priscilla T Ribeiro.pdf: 1928471 bytes, checksum: 2e387f8cd9c9d72ef4f2f30ecf57cebc (MD5) ficha catalográfica.pdf: 92086 bytes, checksum: 2bcb4a421e2ed5a5806b0418cacbd08d (MD5) / Approved for entry into archive by Divisão de Documentação/BC Biblioteca Central (ddbc@ufam.edu.br) on 2015-07-06T18:25:53Z (GMT) No. of bitstreams: 2 Dissertação-Priscilla T Ribeiro.pdf: 1928471 bytes, checksum: 2e387f8cd9c9d72ef4f2f30ecf57cebc (MD5) ficha catalográfica.pdf: 92086 bytes, checksum: 2bcb4a421e2ed5a5806b0418cacbd08d (MD5) / Approved for entry into archive by Divisão de Documentação/BC Biblioteca Central (ddbc@ufam.edu.br) on 2015-07-06T18:31:15Z (GMT) No. of bitstreams: 2 Dissertação-Priscilla T Ribeiro.pdf: 1928471 bytes, checksum: 2e387f8cd9c9d72ef4f2f30ecf57cebc (MD5) ficha catalográfica.pdf: 92086 bytes, checksum: 2bcb4a421e2ed5a5806b0418cacbd08d (MD5) / Made available in DSpace on 2015-07-06T18:31:15Z (GMT). No. of bitstreams: 2 Dissertação-Priscilla T Ribeiro.pdf: 1928471 bytes, checksum: 2e387f8cd9c9d72ef4f2f30ecf57cebc (MD5) ficha catalográfica.pdf: 92086 bytes, checksum: 2bcb4a421e2ed5a5806b0418cacbd08d (MD5) Previous issue date: 2015-02-27 / FAPEAM - Fundação de Amparo à Pesquisa do Estado do Amazonas / The Passiflora nitida is a species of Amazonian flora still little studied that added to its antioxidant potential, reported in the literature, mainly due to the presence of phenols and flavonoids, arouses the interest for use in a product. The objective of this study was to develop a semi-solid formulation with rejuvenating activity and whitening of the skin. The plant drug was collected at Embrapa-AM, the dry extract obtained by sprinkler in spray dryer was prepared in three different concentrations (plant-drug solvent). In the characterization of the raw material plant tests as loss on drying, particle size by sieving, extractive content, ash content, phytochemical screening were performed with the plant drug. With the dry extract was made using laser particle size tests, analysis of specific surface area, fluorescence x-rays, thermal analysis, dry residue, phenol and total flavonoids, DPPH, ABTS, tyrosinase inhibition activity, chelating activity and reducing potential. Further, semisolid formulations were developed conducted the pre-test stability, primary stability, accelerated stability, rheological studies, and spreadability. Subsequently, tests were performed in cell culture to evaluate the cytotoxicity and the ability to reduce melanogenesis and inhibiting tyrosinase. It was also determined the total flavonoid content by UV-Visible esctrofotometria both the dry extract as in the formulation. In characterizing the plant drug loss on drying was 9.08%, the powder was classified as coarse powder. The extractive yield was 17.16% and the ash content was 5.43%. In the phytochemical screening results were strongly positive for flavônicos glycosides. The dry extract was classified as fine powder with average particle diameter of 25.06 μM and pores of the mesopore type. The chemicals found by the fluorescence X-ray technique are Ca, K, Cl, S, P, Si, Mg, Na and Br. In the thermal analysis, there are four thermal events observed in the DSC curve and four loss event mass in the TG / DTG curve. For the dry extract 7.5% the result of total phenols was 18.9%, total flavonoid 4.1%. IC50 of DPPH was 31.5 μg/mL, IC50 of ABTS was 21.8 μg/mL, IC50 inhibition of tyrosinase was 437 μg/mL. The extract showed no chelating activity and reducing potential of 29.7% at a concentration of 50 μg/mL. In the development of formulations, eighteen bases were manipulated and from below, various formulations were tested in pre-stability. Four formulations followed for primary stability and for accelerated stability. In accelerated stability LB12 with 3% Passiflora nitida was the formulation selected, the rheological profile showed pseudoplastic behavior with thixotropy and good spreadability. The dry extract and the formulation showed no cytotoxicity, ability to reduce both showed melanogenesis, but did not inhibit tyrosinase in cell culture. The total flavonoid content was 1.2% for dry extract and 0.2% for the formulation. The dried extract of Passiflora nitida has in vitro antioxidant activity and capacity reduction of melanogenesis. The results provided the establishment of dry extract of the quality control parameters, and provide the data as: formulation, physical and chemical specifications and stability, for the regularization of the cosmetic product at the National Health Surveillance Agency Sanitária- ANVISA. In this way, managed to get a semi-solid cosmetic product with Amazon bioactive, - Passiflora nitida dry extract, safe, stable and with the antioxidant activities and whitening of the skin, proven in vitro. / A Passiflora nitida é uma espécie da flora amazônica ainda pouco estudada que somada ao seu potencial antioxidante, relatado na literatura, devido principalmente à presença de fenóis e flavonoides, desperta o interesse para aplicação em um produto. O objetivo do presente trabalho foi desenvolver uma formulação semissólida com atividade rejuvenescedora e clareadora da pele. A droga vegetal foi coletada na Embrapa-AM, o extrato seco obtido por aspersão em spray dryer foi preparado em três diferentes concentrações (droga vegetal-solvente). Na caracterização da matéria-prima vegetal testes como perda por dessecação, granulometria por tamisação, teor extrativo, teor de cinzas, triagem fitoquímica foram realizados com a droga vegetal. Com o extrato seco foram realizados os testes de granulometria a laser, análise da área superficial específica, fluorescência de raios-x, análises térmicas, resíduo seco, teor de fenóis e flavonoides totais, DPPH, ABTS, atividade de inibição da tirosinase, atividade quelante e potencial redutor. Na sequência, foram desenvolvidas formulações semissólidas e realizado os testes de pré-estabilidade, estabilidade preliminar, estabilidade acelerada, estudo reológico, espalhabilidade. Posteriormente, foram realizados testes em cultura de células para avaliar a citotoxicidade e a capacidade de reduzir a melanogênese e inibir a tirosinase. Também foi determinado o teor de flavonoides totais por esctrofotometria UV-Visível tanto no extrato seco como na formulação. Na caracterização da droga vegetal a perda por dessecação foi de 9,08 %, o pó foi classificado como grosso. O teor extrativo foi de 17,16 % e o teor de cinzas foi de 5,43 %. Na triagem fitoquímica o resultado foi fortemente positivo para heterosídeos flavônicos. O extrato seco foi classificado como pó finíssimo, com diâmetro médio de partículas de 25,06 μM e poros do tipo mesoporos. Os elementos químicos encontrados pela técnica de fluorescência de raios-x foram Ca, K, Cl, S, P, Si, Mg, Na e Br. Na análise térmica, temos quatro eventos térmicos observados na curva de DSC e quatro eventos de perda de massa na curva de TG/DTG. Para o extrato seco a 7,5% o resultado de fenóis totais foi de 18,9 %, flavonoides totais de 4,1 %. A CI50 do DPPH foi de 31,5 μg/mL, CI50 do ABTS foi de 21,8 μg/mL, CI50 da inibição de tirosinase foi de 437 μg/mL. O extrato não apresentou atividade quelante e o potencial redutor foi de 29,7 % na concentração de 50 μg/mL. No desenvolvimento das formulações, dezoito bases foram manipuladas, a partir das quais várias formulações foram testadas na pré-estabilidade. Quatro formulações seguiram para estabilidade preliminar e estabilidade acelerada. A formulação selecionada na estabilidade acelerada foi a LB12 com 3% de Passiflora nitida. A LB12 3% apresentou comportamento pseudoplástico com tixotropia e boa espalhabilidade. O extrato seco e a formulação não apresentaram citotoxicidade e ambos apresentaram capacidade de reduzir a melanogênese. Entretanto, não houve inibição da tirosinase em cultura de células. O teor de flavonoides totais foi de 1,2 % para o extrato seco e de 0,2% para a formulação. O extrato seco de Passiflora nitida possui, in vitro, atividade antioxidante e capacidade de redução da melanogênese. Os resultados possibilitam o estabelecimento de parâmetros de controle de qualidade do extrato seco, e fornecem dados como: formulação, especificações físico-químicas e estabilidade, para a regularização do produto cosmético junto a Agência Nacional de Vigilância Sanitária- ANVISA. Desta maneira, conseguimos obter um produto cosmético semissólido com o bioativo amazônico - extrato seco de Passiflora nitida, seguro, estável e com as atividades antioxidante e clareadora da pele, comprovadas in vitro.

Bien qu’au 11ème rang des cancers les plus fréquents et au 12ème rang des cancers les plus mortels, le mélanome reste un problème médical majeur préoccupant. En effet, cette pathologie, au stade métastatique, reste réfractaire à la chimiothérapie et aux thérapies ciblées. Un certain nombre d’arguments définissent la résistance à l’apoptose comme un point crucial dans l’échec aux traitements anti-cancéreux. Ces mécanismes de résistance et chimiorésistance spécifiques aux mélanomes, déclenchés en réponse aux traitements traditionnels, sont la conséquence d’une dérégulation des voies apoptotiques suite à l’activation des protéines anti-apoptotiques, l’inactivation des protéines pro-apoptotiques avec renforcement des signaux de survie (voies de survie PI3K/Akt, NF-κB et MAPK/ERK). Une étude effectuée sur la lignée murine de mélanome B16-F0 a introduit la mélanogenèse comme une forme de résistance à l’apoptose induite par l’acide ursolique (AU), un triterpène pentacyclique d’origine naturelle ; ainsi les cellules entrant en apoptose sont capables de déclencher une résistance qui se manifeste par une surproduction de la mélanine tout en retardant la mort cellulaire. D’autres études ont montré l’implication de la COX-2 dans un mécanisme de résitance à l’apoptose dans plusieurs types de cancers dans le but de retarder leur mort cellulaire. Dans cette optique, nous nous sommes intéressés à étudier l’implication de la mélanogenèse et de la voie COX-2/PGE2 dans la résistance à l’apoptose dans le mélanome et plus précisément dans un modèle d’apoptose induite par l’AU sur la lignée humaine de mélanome M4Beu. Par la suite, nous avons décrit une interaction probable entre ces deux voies distinctes, la mélanogenèse et la voie COX-2/PGE2. Dans un autre contexte, nous avons montré que l’AU inhibe les voies de survie PI3K/Akt et ERK1/2, ce qui favorise ses effets pro-apoptotique et anti-prolifératif. Notre étude permet de mieux explorer les mécanismes de résistance spécifiques aux mélanomes tout en suggérant l’effet bénéfique de l’AU comme adjuvant naturel aux traitements chimio-thérapeutiques traditionnels. / Despite the deployment of targeted therapies, the incidence and mortality rates of cutaneous melanoma is increasing very fast making it a pre-eminent public health threat. Previously, we had showed that B16-F0 murine melanoma cells undergoing apoptosis are able to delay their own death induced by ursolic acid (UA), a natural pentacyclic triterpenoid compound. We had demonstrated that tyrosinase and TRP-1 up-regulation in apoptotic cells and the subsequent production of melanin were implicated in an apoptosis resistance mechanism. Several resistance mechanisms to apoptosis have been characterized in melanoma such as hyperactivation of DNA repair mechanisms, drug efflux systems, and reinforcement of survival signals (PI3K/Akt, NF-κB and MAPK/ERK pathways). Otherwise, other mechanisms of apoptosis resistance involving different proteins, such as cyclooxygenase-2 (COX-2), have been described in many cancer types. In this study, we demonstrated the involvement of melanogenesis and COX-2/PGE2 pathway in resistance to UA-induced apoptosis in human M4Beu melanoma cells. Then, we established the evidence that an interaction exists between these two pathways by investigating on the one hand the effect of inhibiting melanogenesis by N-phenylthiourea (PTU) on COX-2 expression and its product PGE2, and on the other hand the effect of inhibiting COX-2 activity using NS-398 on tyrosinase expression and melanin production. Furthermore, we showed that anti-proliferative and proapoptotic effects of UA were mediated through modulation of multiple signaling pathways including Akt and ERK-1/2 proteins. Our study not only uncovers underlying molecular mechanisms of UA action in human melanoma cancer cells but also suggest its great potential as an adjuvant in treatment and cancer prevention.