Recent progress in the development of artificial skin concepts is a result of the increased demand for providing environment perception such as touch and flow sensing to robots, prosthetics and surgical tools. Tactile sensors are the essential components of artificial skins and attracted considerable attention that led to the development of different technologies for mimicking the complex sense of touch in humans. This dissertation work is devoted to the development of a bioinspired tactile sensing technology that imitates the extremely sensitive hair-like cilia receptors found in nature. The artificial cilia are fabricated from permanent magnetic, biocompatible and highly elastic nanocomposite material, and integrated on a giant magneto-impedance magnetic sensor to measure the stray field. A force that bends the cilia changes the stray field and is therefore detected with the magnetic sensor, providing high performance in terms of sensitivity, power consumption and versatility. The nanocomposite is made of Fe nanowires (NWs) incorporated into polydimethylsiloxane (PDMS). Fe NWs have a high remanent magnetization, due the shape anisotropy; thus, they are acting as permanent nano-magnets. This allows remote device operation and avoids the need for a magnetic field to magnetize the NWs, benefiting miniaturization and the possible range of applications. The magnetic properties of the nanocomposite can be easily tuned by modifying the NWs concentration or by aligning the NWs to define a magnetic anisotropy. Tactile sensors are realized on flexible and rigid substrates that can detect flow, vertical and shear forces statically and dynamically, with a high resolution and wide operating range. The advantage to operate the sensors in liquids and air has been utilized to measure flows in different fluids in a microfluidic channel. Various dynamic studies were conducted with the tactile sensor demonstrating the detection of moving objects or the texture of objects. Overall, the results confirm the possibility to easily control the sensors’ performance with the cilia arrangement and dimensions. The cost effective mold-based microfabrication process and magnetic operation enable a high degree of integration, which together with the extremely low power consumption make the artificial cilia sensor reported in this dissertation an attractive solution for many applications.

Os melanomas são o tipo mais mortal de câncer de pele, apesar da baixa incidência, 80% das mortes de câncer de pele são devem-se ao melanoma metastático. Novas abordagens farmacológicas e a busca por novos compostos para a terapêutica do melanoma, em aplicações isolados ou em combinação com outros fármacos é imprescindível. Esta busca ocorre principalmente no campo das terapias de alvos específicos, devido à aquisição de resistência tumoral e recidiva. O resveratrol (RES) é um polifenol com atividade anti-oxidante, e seu efeito anti-tumoral foi mostrado pela indução de morte celular, porém o seu estudo não foi aprofundado pela inviabilidade do uso de altas doses in vitro para observação de efeitos celulares. Outro composto, o 2-methoxiestradiol (2ME) é um metabólito do estrógeno cujo efeitos anti-câncer já foi demonstrado em melanoma, porém sem elucidação das vias de sinalização envolvidas. O efeito em células com resistência adquirida também nunca foram testados. Neste estudo ampliamos o painel de linhagens celulares de melanoma humano, e demonstramos que o 2ME induz morte celular, inibe a proliferação destas células sendo que esta inibição está associada a indução de senescência. Pela primeira vez foi observada a inibição de proliferação pelo 2ME em células com a mutação BRAF V600E resistentes ao vemurafenibe (inibidor de BRAF) e duplo resistentes ao vemurafenibe e trametinibe (inibidor de MEK). A inibição de proliferação foi acompanhada pela modulação de p21Cip1, Ciclina B1, pRb, proteínas envolvidas na regulação do ciclo celular. A exposição prolongada ao 2ME inibiu a formação de colônias em todas as linhagens de melanoma (não resistentes e resistentes), mas não teve o mesmo efeito em fibroblastos primários, mostrando efeito seletivo. Em modelo tridimensional de esferóides, foi observado que as linhagens resistentes (Sk-Mel-28R) e duplo resistentes (Sk- Mel-28RT) são mais invasivas que a parental (Sk-Mel-28). Neste modelo, o 2ME foi capaz de inibir a invasão e viabilidade destas células. No modelo de pele reconstituída, na ausência de tratamento, observa-se invasão das células de melanoma pela derme, porém este fenômeno é diminuído quando as peles são tratadas com 2ME. Estes resultados demonstram que o 2ME é um efetivo agente anti-melanoma, independente de sua resistência. / Melanomas are the deadliest type of skin cancer, and in spite of the low incidence, 80% of the skin cancer associated death cases are due to metastatic melanoma. New pharmacological approaches and the search of new compounds for melanoma therapeutics, for monotherapy or combination therapy, are essential. This search occurs mainly in the targeted therapy field because of the melanoma acquisition of resistance to the current treatments. Resveratrol (RES) is a polyphenol with anti-oxidant activity, and its anti-tumor effect has been shown through the induction of cell death. However, the study of this compound has been discontinued in this work due to the impossibility of using high doses in vitro for the observation of cellular effects. Another compound, 2-methoxyestradiol (2ME) is a metabolite from estrogen, and its anti-cancer effects has already been shown in melanoma, but with no elucidation of the signaling pathways involved. Furthermore, the effects in cells with acquired resistance have never been shown. In this study we used a broader panel of human melanoma cell lines and demonstrated that 2ME induces cell death, inhibits proliferation of these cells, and this inhibition is associated with the induction of cell senescence. The inhibition of proliferation caused by 2ME was observed for the first time in BRAF V600E cells that are resistant to Vemurafenib (BRAF inhibitor) and double resistant to Vemurafenib and Trametinib (MEK inhibitor). The proliferation inhibition was related to the modulation of p21Cip1,Cyclin B1 and pRb, which are proteins involved in cell cycle regulation. Long exposure to 2ME in colony formation assay showed the inhibition of colony in all melanoma cell lines (regardless of resistance and mutational status), but not in primary fibroblasts, showing selective effect. In three-dimensional spheroid model, it was observed that the resistant (Sk-Mel- 28R) and double-resistant (Sk-Mel-28RT) cell lines were more invasive than the parental cell line (Sk-Mel-28). In this model, 2ME was able to inhibit cell invasion and cell viability. In the skin reconstruct model, in the absence of treatment, melanoma cell invasion can be observed in the dermis layer. However, after the treatment with 2ME these cell invasion foci are inhibited. Altogether, these effects demonstrate that 2ME is an effective anti-melanoma agent, regardless of resistance.

O melanoma é a forma mais mortal de câncer de pele, origina-se de células produtoras de pigmentos, os melanócitos. Esses podem ser cutâneos ou não-cutâneos (encontrados no revestimento da membrana coróide do olho, nas meninges, e nos tratos gastrintestinal e geniturinário). O aumento da incidência de melanomas malignos nas últimas décadas, e sua alta taxa de mortalidade e grande resistência a maior parte das terapias, tem sido um enorme desafio para a comunidade científica. Particularmente, a falta de habilidade de indução à morte por apoptose em resposta à quimioterapia e outros estímulos externos permitem uma vantagem seletiva para progressão tumoral, formação de metástase e resistência à terapia em melanomas. O estresse oxidativo e espécies reativas de oxigênio (EROs) vêm sendo, há muito tempo, reconhecidos como importantes desencadeadores e moduladores da apoptose. Porém o exato papel do estresse oxidativo no processo apoptótico ainda é uma questão de debate. Antioxidantes tendem a possuir propriedades regulatórias de tradução de sinais que devem ou não estar ligadas as suas capacidades de inativar oxidantes. Porém em certas condições, um forte ambiente oxidante onde há falta de suporte para regenerar (reduzir) antioxidantes oxidados, permite que alguns antioxidantes assumam características de um pró-oxidante. Foi demonstrada a capacidade citotóxica de um potente antioxidante, 4-nerolidilcatecol (4-NC), extraído da planta Pothomorphe umbellata L. Miq, sobre linhagens tumorais de melanoma e sobre fibroblastos humanos normais. Esse composto foi capaz de induzir a parada do ciclo celular em G1, bem como diminuir a atividade de MMPs e em outras linhagens de melanoma foi capaz de induzir a morte celular por apoptose. Estudos subseqüentes mostraram que o mecanismo de ação deste composto inicia-se com a formação e acúmulo de EROs, além da inibição da enzima catalase. O 4-NC foi capaz de induzir a morte por apoptose via mitocondrial, aumentando os níveis das proteínas p53, Noxa, Mcl-1, clivando Bax e Bid e induzindo a clivagem das caspases 3 e 9. Além disso, em modelo de pele artificial contendo melanoma, o 4-NC foi capaz de conter a invasão do melanoma para a estrutura dérmica da pele reconstituída. Foram utilizadas como controle de diferenciação as proteínas Queratina 10 e 14, Involucrina e, como marcador do melanoma, a proteína S100. Parte desta invasão é contida devido à inibição da ativação das MMP-2 e -9 e ativação de TIMP-2 pelo 4-NC. Sendo assim, esse composto se mostra como um potencial quimioterápico no tratamento do melanoma humano. / Melanoma is the most agressive form of skin cancer, it arises from the pigment-producing cells, melanocytes. These may be cutaneous or non-cutaneous (found in the lining membrane of the eye choroid, the meninges, and gastrointestinal and genitourinary tracts). The increased incidence of malignant melanomas in recent decades, its high mortality rate and high resistance to most therapies has been a major challenge to the scientific community. It\'s particularly difficult to induce cell death by apoptosis in response to chemotherapy and other external stimuli, which may provide a selective advantage for tumor progression, metastasis formation and resistance to therapy in melanoma. Oxidative stress and reactive oxygen species (ROS) have been recognized for a long time as important triggers and modulators of apoptosis, but the exact role of oxidative stress in the apoptotic process is still a matter of discussion. Antioxidants tend to possess properties to regulate transduction signals that may not be related to their ability to inactivate oxidants. Under certain conditions, in a strong oxidizing environment where there is lack of support to regenerate (reduce) oxidized antioxidants, some antioxidants can assume characteristics of pro-oxidant. The 4-nerolidylcatechol (4-NC) is a potent antioxidant that is extracted from the plant Pothomorphe umbellata L. Miq. Its citotoxic potential has been demonstrated on melanoma tumor cell lines and on normal human fibroblasts. This compound was able to induce cell cycle arrest in G1, decrease the activity of MMPs and cell death by apoptosis. Subsequent studies showed that the mechanism of action of this compound starts with the formation and accumulation of ROS, and inhibition of the enzyme catalase. The 4-NC was able to induce apoptosis via mitochondria, increasing the levels of p53, Noxa, Mcl1, cleaving Bax and Bid and inducing cleavage of caspases 3 and 9. Furthermore, in a model of artificial skin containing melanoma 4-NC was able to contain the invasion of melanoma to the dermal part of the skin. Proteins keratin 10 and 14, involucrin and S100 were used as control of differentiation. Part of this invasion is restrained due to TIMP-2 activation and the inhibition of MMP-2 and -9 activation by 4-NC. Concluding, this compound can be used as a potential chemotherapeutic agent in the treatment of human melanoma.

This thesis reports the discovery of a wide negative di↵erential resistance (NDR) region in a graphite-silicone composite that was utilized to create a strain-tuned flexible oscillator. Encoding the strain into frequency mimics the behavior of mechanoreceptor neurons in the skin and demonstrates a flexible and electronically active material suitable for state of the art bio-electronic applications. The NDR was investigated over a range of composite filling fractions and temperatures; alongside theoretical modelling to calculate the tunneling current through a graphite-silicone barrier. This led to the understanding that the NDR is the result of a semi-metal to insulator transition of embedded graphene bilayers within the graphite nanoparticles. The transition, brought about by a transverse bias across specifically orientated particles, opens a partial band-gap at the Fermi level of the bilayer. NDR in a flexible material has not been observed before and has potential for creating a flexible active device. The electromechanical properties of the composite were considered through a bend induced bilayer strain. The piezoresistance was found to be dominated by transient resistance spiking from the breaking of conduction lines, which then reform according to the viscoelasticity of the polymer matrix. The resistance spiking was embraced as a novel method for sensitive di↵erential pressure detection, used in the development of two applications. Firstly, it was employed for the detection of ultrasound waves and found to have an acoustic pressure detection threshold as low as 48 Pa. A commensurability was observed between the composite width and ultrasound wavelength which was shown to be consistent with the formation of standing waves, described by Bragg’s law. Secondly, a differential pressure array of 64 composite pixels was fabricated and demonstrated to image pressures under 3.8 kPa at a resolution of 10 dpi. The NDR active region was incorporated into an LC circuit where it was demonstrated to sustain oscillations of up to 12.5 kHz. The composite was then strained and an intrinsic frequency was observed which had a linear dependence on the strain with a frequency shift of 84 Hz / % strain. Lastly the composite was used in a strain-tuned amplifier circuit and shown to provide a gain of up to 4.5. This thesis provided the groundwork for a completely flexible electronically active device for futuristic bio-electronic skins with resolutions and sensitivities rivalling those of human tactile sensing.

O desenvolvimento de novos modelos de pele e novas metodologias in vitro segue uma tendência mundial na busca pela redução ou substituição de testes em animais. Nesse contexto, kits de epiderme humana reconstruída (RHE) apresentam-se como uma plataforma promissoras para essa proposta e, alguns modelos encontram-se validados para ensaios de irritação e corrosão cutânea in vitro. Entretanto, em países como o Brasil, enfrentam-se questões alfandegárias e perda do material por perecibilidade, dificultando e até impedindo, a importação desses kits para utilização por parte das indústrias e laboratórios nacionais. Em contrapartida, o desenvolvimento de um modelo de RHE apresenta-se como um avanço tecnológico e ganho de autonomia para esses países. Assim, no capítulo 1 explorou-se o desenvolvimento de um modelo nacional de RHE (USP-RHE) que atendesse às exigências internacionais descritas no guia OECD 439. O modelo desenvolvido apresentou uma epiderme bem diferenciada e atendeu aos parâmetros de qualidade (histologia, viabilidade e função barreira) bem como da funcionalidade, a qual é expressa na capacidade de distinção entre irritantes e não irritantes, apresentando 85,7% de especificidade, 100% sensibilidade e 92,3% de acurácia quando comparada com a classificação in vivo obtida pelo ensaio do linfonodo local (LLNA). No capítulo 2, células monocíticas THP-1 em monocamada foram capazes de distinguir entre agentes sensibilizantes e não sensibilizantes por meio da expressão de CD86, CD54 e liberação de IL-8. Após a obtenção de RHE e THP-1 funcionais, um cross-talking foi estabelecido gerando uma RHE imunocompetente. A RHEI distinguiu satisfatoriamente entre agentes sensibilizantes e não sensibilizantes por meio da expressão de CD86 e CD54 na membrana das células THP-1. A liberação de IL-8 também foi avaliada na RHEI, mas, não demonstrou ser um bom indicador para a avaliação de sensibilização, ao contrário de IL-1α, que distinguiu satisfatoriamente agentes sensibilizantes de não-sensibilizantes, mas não foi capaz de hierarquizá-los. No capítulo 3, avaliou-se o papel de interleucinas do tipo Th2 e da depleção de colesterol na membrana plasmática no desenvolvimento de características morfológicas e moleculares da dermatite atópica (DA) in vitro em um modelo de RHE. Os resultados demonstram que o uso de IL-4, IL-13 e IL-25 em combinação com a depleção de colesterol na membrana plasmática mimetiza in vitro, as principais características da DA. No capítulo 4, buscou-se avaliar os efeitos imunossupressores da radiação ultravioleta na RHEI. Os ensaios foram realizados em diferentes períodos de exposição, entretanto, não foi possível observar tais efeitos. Os resultados justificam-se pela ausência da liberação de IL-10 pelo RHE imunocompetente, por exemplo, e demonstram uma limitação do RHE imunocompetente para avaliações de inativação da reposta imune. Neste trabalho, concluímos que foi possível obter uma RHE competitiva, similar aos modelos internacionais validados e que pode ser utilizada como plataforma para ensaios de irritação e sensibilização cutânea, além de ser uma plataforma para estudos da dermatite atópica. No modelo é possível estudar a ativação do sistema imune, o que o torna promissor como uma plataforma para avaliação de resposta imunológica in vitro. Conclui-se, portanto, que os objetivos foram amplamente atendidos além de oferecermos um protocolo de livre acesso para reprodução por outros laboratórios e um modelo para validação futura. / The development of new in vitro skin models and new methodologies follows a global trend in search for reductions or replacement of animal testing. In this context, Reconstructed Human Epidermis kits (RHE) are presented as a promising platform in the search for alternative methods to animal use, and some models are validated for skin irritation and corrosion in vitro tests. However, in countries such as Brazil, who face customs issues and loss of material due to perishability, making it challenging and even compromising the importation of these kits for use by industries and laboratories. In contrast, the development of an RHE model is presented as a technological breakthrough and gain of autonomy for these countries. Thus, in Chapter 1 we explored the development of a national model of RHE (USP-RHE) that meet international requirements described in OECD TG 439. The developed model presented a well-differentiated epidermis and met the quality parameters, for instance, histology, viability, and barrier function as well as the functionality expressed in the capacity of screening between irritants and nonirritants, with 85.7 % of specificity, 100 % of sensitivity and 91.7% of accuracy in comparision to in vivo UN GHS classification from Local limph node assay (LLNA). In chapter 2, monocytic THP-1 cell line, as monolayers, were able to distinguish between sensitizers and non-sensitizers by expression of CD86, CD54, and IL-8 release. In this model, functional RHE and THP-1 were used in a cross-talking, and thus an immunocompetent RHE (RHEI) was generated. The RHEI has distinguished satisfactorily between sensitizers and non-sensitizers through CD86 and CD54 expression that was larger and more sensitive in this model. The release of IL-8 was also evaluated in RHEI, however, did not demonstrate to be a good parameter for this evaluation, unlike IL-1α, which satisfactorily distinguished sensitizers from non-sensitizers, but was not able to hierarchize them. In chapter 3, we evaluated the role of Th2-related cytokines and plasma membrane cholesterol depletion (CD) in the development of atopic dermatitis (AD) morphological and molecular characteristics in an in vitro model of RHE. The results showed that combination of IL-4, IL-13 and IL-25 in combination with CD can reproduce the major features of AD in vitro. In Chapter 4, we sought to evaluate the ultraviolet radiation-induced immunosuppressive effects in RHE. The tests were performed at different times. However, it was not possible to observe such effects. The results are justified by the absence of IL-10 release by RHEI, for example, and show a limitation of RHEI for rating inactivation of the immune response. In this work, we conclude that it was possible to obtain a competitive RHE similar to the validated international models that can be used as a platform for irritation and skin sensitization tests, besides being a platform for the study of atopic dermatitis. Using this model is possible to explore the activation of immune system, which makes it promising as a platform for the evaluation of immune response in vitro. We conclude, therefore, that the objectives have been met as well as it is offering an open source protocol for breeding by other laboratories, thus offering the RHE model developed here for future validation tests.

O desenvolvimento de novos modelos de pele e novas metodologias in vitro segue uma tendência mundial na busca pela redução ou substituição de testes em animais. Nesse contexto, kits de epiderme humana reconstruída (RHE) apresentam-se como uma plataforma promissoras para essa proposta e, alguns modelos encontram-se validados para ensaios de irritação e corrosão cutânea in vitro. Entretanto, em países como o Brasil, enfrentam-se questões alfandegárias e perda do material por perecibilidade, dificultando e até impedindo, a importação desses kits para utilização por parte das indústrias e laboratórios nacionais. Em contrapartida, o desenvolvimento de um modelo de RHE apresenta-se como um avanço tecnológico e ganho de autonomia para esses países. Assim, no capítulo 1 explorou-se o desenvolvimento de um modelo nacional de RHE (USP-RHE) que atendesse às exigências internacionais descritas no guia OECD 439. O modelo desenvolvido apresentou uma epiderme bem diferenciada e atendeu aos parâmetros de qualidade (histologia, viabilidade e função barreira) bem como da funcionalidade, a qual é expressa na capacidade de distinção entre irritantes e não irritantes, apresentando 85,7% de especificidade, 100% sensibilidade e 92,3% de acurácia quando comparada com a classificação in vivo obtida pelo ensaio do linfonodo local (LLNA). No capítulo 2, células monocíticas THP-1 em monocamada foram capazes de distinguir entre agentes sensibilizantes e não sensibilizantes por meio da expressão de CD86, CD54 e liberação de IL-8. Após a obtenção de RHE e THP-1 funcionais, um cross-talking foi estabelecido gerando uma RHE imunocompetente. A RHEI distinguiu satisfatoriamente entre agentes sensibilizantes e não sensibilizantes por meio da expressão de CD86 e CD54 na membrana das células THP-1. A liberação de IL-8 também foi avaliada na RHEI, mas, não demonstrou ser um bom indicador para a avaliação de sensibilização, ao contrário de IL-1α, que distinguiu satisfatoriamente agentes sensibilizantes de não-sensibilizantes, mas não foi capaz de hierarquizá-los. No capítulo 3, avaliou-se o papel de interleucinas do tipo Th2 e da depleção de colesterol na membrana plasmática no desenvolvimento de características morfológicas e moleculares da dermatite atópica (DA) in vitro em um modelo de RHE. Os resultados demonstram que o uso de IL-4, IL-13 e IL-25 em combinação com a depleção de colesterol na membrana plasmática mimetiza in vitro, as principais características da DA. No capítulo 4, buscou-se avaliar os efeitos imunossupressores da radiação ultravioleta na RHEI. Os ensaios foram realizados em diferentes períodos de exposição, entretanto, não foi possível observar tais efeitos. Os resultados justificam-se pela ausência da liberação de IL-10 pelo RHE imunocompetente, por exemplo, e demonstram uma limitação do RHE imunocompetente para avaliações de inativação da reposta imune. Neste trabalho, concluímos que foi possível obter uma RHE competitiva, similar aos modelos internacionais validados e que pode ser utilizada como plataforma para ensaios de irritação e sensibilização cutânea, além de ser uma plataforma para estudos da dermatite atópica. No modelo é possível estudar a ativação do sistema imune, o que o torna promissor como uma plataforma para avaliação de resposta imunológica in vitro. Conclui-se, portanto, que os objetivos foram amplamente atendidos além de oferecermos um protocolo de livre acesso para reprodução por outros laboratórios e um modelo para validação futura. / The development of new in vitro skin models and new methodologies follows a global trend in search for reductions or replacement of animal testing. In this context, Reconstructed Human Epidermis kits (RHE) are presented as a promising platform in the search for alternative methods to animal use, and some models are validated for skin irritation and corrosion in vitro tests. However, in countries such as Brazil, who face customs issues and loss of material due to perishability, making it challenging and even compromising the importation of these kits for use by industries and laboratories. In contrast, the development of an RHE model is presented as a technological breakthrough and gain of autonomy for these countries. Thus, in Chapter 1 we explored the development of a national model of RHE (USP-RHE) that meet international requirements described in OECD TG 439. The developed model presented a well-differentiated epidermis and met the quality parameters, for instance, histology, viability, and barrier function as well as the functionality expressed in the capacity of screening between irritants and nonirritants, with 85.7 % of specificity, 100 % of sensitivity and 91.7% of accuracy in comparision to in vivo UN GHS classification from Local limph node assay (LLNA). In chapter 2, monocytic THP-1 cell line, as monolayers, were able to distinguish between sensitizers and non-sensitizers by expression of CD86, CD54, and IL-8 release. In this model, functional RHE and THP-1 were used in a cross-talking, and thus an immunocompetent RHE (RHEI) was generated. The RHEI has distinguished satisfactorily between sensitizers and non-sensitizers through CD86 and CD54 expression that was larger and more sensitive in this model. The release of IL-8 was also evaluated in RHEI, however, did not demonstrate to be a good parameter for this evaluation, unlike IL-1α, which satisfactorily distinguished sensitizers from non-sensitizers, but was not able to hierarchize them. In chapter 3, we evaluated the role of Th2-related cytokines and plasma membrane cholesterol depletion (CD) in the development of atopic dermatitis (AD) morphological and molecular characteristics in an in vitro model of RHE. The results showed that combination of IL-4, IL-13 and IL-25 in combination with CD can reproduce the major features of AD in vitro. In Chapter 4, we sought to evaluate the ultraviolet radiation-induced immunosuppressive effects in RHE. The tests were performed at different times. However, it was not possible to observe such effects. The results are justified by the absence of IL-10 release by RHEI, for example, and show a limitation of RHEI for rating inactivation of the immune response. In this work, we conclude that it was possible to obtain a competitive RHE similar to the validated international models that can be used as a platform for irritation and skin sensitization tests, besides being a platform for the study of atopic dermatitis. Using this model is possible to explore the activation of immune system, which makes it promising as a platform for the evaluation of immune response in vitro. We conclude, therefore, that the objectives have been met as well as it is offering an open source protocol for breeding by other laboratories, thus offering the RHE model developed here for future validation tests.

O melanoma é a forma mais mortal de câncer de pele, origina-se de células produtoras de pigmentos, os melanócitos. Esses podem ser cutâneos ou não-cutâneos (encontrados no revestimento da membrana coróide do olho, nas meninges, e nos tratos gastrintestinal e geniturinário). O aumento da incidência de melanomas malignos nas últimas décadas, e sua alta taxa de mortalidade e grande resistência a maior parte das terapias, tem sido um enorme desafio para a comunidade científica. Particularmente, a falta de habilidade de indução à morte por apoptose em resposta à quimioterapia e outros estímulos externos permitem uma vantagem seletiva para progressão tumoral, formação de metástase e resistência à terapia em melanomas. O estresse oxidativo e espécies reativas de oxigênio (EROs) vêm sendo, há muito tempo, reconhecidos como importantes desencadeadores e moduladores da apoptose. Porém o exato papel do estresse oxidativo no processo apoptótico ainda é uma questão de debate. Antioxidantes tendem a possuir propriedades regulatórias de tradução de sinais que devem ou não estar ligadas as suas capacidades de inativar oxidantes. Porém em certas condições, um forte ambiente oxidante onde há falta de suporte para regenerar (reduzir) antioxidantes oxidados, permite que alguns antioxidantes assumam características de um pró-oxidante. Foi demonstrada a capacidade citotóxica de um potente antioxidante, 4-nerolidilcatecol (4-NC), extraído da planta Pothomorphe umbellata L. Miq, sobre linhagens tumorais de melanoma e sobre fibroblastos humanos normais. Esse composto foi capaz de induzir a parada do ciclo celular em G1, bem como diminuir a atividade de MMPs e em outras linhagens de melanoma foi capaz de induzir a morte celular por apoptose. Estudos subseqüentes mostraram que o mecanismo de ação deste composto inicia-se com a formação e acúmulo de EROs, além da inibição da enzima catalase. O 4-NC foi capaz de induzir a morte por apoptose via mitocondrial, aumentando os níveis das proteínas p53, Noxa, Mcl-1, clivando Bax e Bid e induzindo a clivagem das caspases 3 e 9. Além disso, em modelo de pele artificial contendo melanoma, o 4-NC foi capaz de conter a invasão do melanoma para a estrutura dérmica da pele reconstituída. Foram utilizadas como controle de diferenciação as proteínas Queratina 10 e 14, Involucrina e, como marcador do melanoma, a proteína S100. Parte desta invasão é contida devido à inibição da ativação das MMP-2 e -9 e ativação de TIMP-2 pelo 4-NC. Sendo assim, esse composto se mostra como um potencial quimioterápico no tratamento do melanoma humano. / Melanoma is the most agressive form of skin cancer, it arises from the pigment-producing cells, melanocytes. These may be cutaneous or non-cutaneous (found in the lining membrane of the eye choroid, the meninges, and gastrointestinal and genitourinary tracts). The increased incidence of malignant melanomas in recent decades, its high mortality rate and high resistance to most therapies has been a major challenge to the scientific community. It\'s particularly difficult to induce cell death by apoptosis in response to chemotherapy and other external stimuli, which may provide a selective advantage for tumor progression, metastasis formation and resistance to therapy in melanoma. Oxidative stress and reactive oxygen species (ROS) have been recognized for a long time as important triggers and modulators of apoptosis, but the exact role of oxidative stress in the apoptotic process is still a matter of discussion. Antioxidants tend to possess properties to regulate transduction signals that may not be related to their ability to inactivate oxidants. Under certain conditions, in a strong oxidizing environment where there is lack of support to regenerate (reduce) oxidized antioxidants, some antioxidants can assume characteristics of pro-oxidant. The 4-nerolidylcatechol (4-NC) is a potent antioxidant that is extracted from the plant Pothomorphe umbellata L. Miq. Its citotoxic potential has been demonstrated on melanoma tumor cell lines and on normal human fibroblasts. This compound was able to induce cell cycle arrest in G1, decrease the activity of MMPs and cell death by apoptosis. Subsequent studies showed that the mechanism of action of this compound starts with the formation and accumulation of ROS, and inhibition of the enzyme catalase. The 4-NC was able to induce apoptosis via mitochondria, increasing the levels of p53, Noxa, Mcl1, cleaving Bax and Bid and inducing cleavage of caspases 3 and 9. Furthermore, in a model of artificial skin containing melanoma 4-NC was able to contain the invasion of melanoma to the dermal part of the skin. Proteins keratin 10 and 14, involucrin and S100 were used as control of differentiation. Part of this invasion is restrained due to TIMP-2 activation and the inhibition of MMP-2 and -9 activation by 4-NC. Concluding, this compound can be used as a potential chemotherapeutic agent in the treatment of human melanoma.

Os melanomas são o tipo mais mortal de câncer de pele, apesar da baixa incidência, 80% das mortes de câncer de pele são devem-se ao melanoma metastático. Novas abordagens farmacológicas e a busca por novos compostos para a terapêutica do melanoma, em aplicações isolados ou em combinação com outros fármacos é imprescindível. Esta busca ocorre principalmente no campo das terapias de alvos específicos, devido à aquisição de resistência tumoral e recidiva. O resveratrol (RES) é um polifenol com atividade anti-oxidante, e seu efeito anti-tumoral foi mostrado pela indução de morte celular, porém o seu estudo não foi aprofundado pela inviabilidade do uso de altas doses in vitro para observação de efeitos celulares. Outro composto, o 2-methoxiestradiol (2ME) é um metabólito do estrógeno cujo efeitos anti-câncer já foi demonstrado em melanoma, porém sem elucidação das vias de sinalização envolvidas. O efeito em células com resistência adquirida também nunca foram testados. Neste estudo ampliamos o painel de linhagens celulares de melanoma humano, e demonstramos que o 2ME induz morte celular, inibe a proliferação destas células sendo que esta inibição está associada a indução de senescência. Pela primeira vez foi observada a inibição de proliferação pelo 2ME em células com a mutação BRAF V600E resistentes ao vemurafenibe (inibidor de BRAF) e duplo resistentes ao vemurafenibe e trametinibe (inibidor de MEK). A inibição de proliferação foi acompanhada pela modulação de p21Cip1, Ciclina B1, pRb, proteínas envolvidas na regulação do ciclo celular. A exposição prolongada ao 2ME inibiu a formação de colônias em todas as linhagens de melanoma (não resistentes e resistentes), mas não teve o mesmo efeito em fibroblastos primários, mostrando efeito seletivo. Em modelo tridimensional de esferóides, foi observado que as linhagens resistentes (Sk-Mel-28R) e duplo resistentes (Sk- Mel-28RT) são mais invasivas que a parental (Sk-Mel-28). Neste modelo, o 2ME foi capaz de inibir a invasão e viabilidade destas células. No modelo de pele reconstituída, na ausência de tratamento, observa-se invasão das células de melanoma pela derme, porém este fenômeno é diminuído quando as peles são tratadas com 2ME. Estes resultados demonstram que o 2ME é um efetivo agente anti-melanoma, independente de sua resistência. / Melanomas are the deadliest type of skin cancer, and in spite of the low incidence, 80% of the skin cancer associated death cases are due to metastatic melanoma. New pharmacological approaches and the search of new compounds for melanoma therapeutics, for monotherapy or combination therapy, are essential. This search occurs mainly in the targeted therapy field because of the melanoma acquisition of resistance to the current treatments. Resveratrol (RES) is a polyphenol with anti-oxidant activity, and its anti-tumor effect has been shown through the induction of cell death. However, the study of this compound has been discontinued in this work due to the impossibility of using high doses in vitro for the observation of cellular effects. Another compound, 2-methoxyestradiol (2ME) is a metabolite from estrogen, and its anti-cancer effects has already been shown in melanoma, but with no elucidation of the signaling pathways involved. Furthermore, the effects in cells with acquired resistance have never been shown. In this study we used a broader panel of human melanoma cell lines and demonstrated that 2ME induces cell death, inhibits proliferation of these cells, and this inhibition is associated with the induction of cell senescence. The inhibition of proliferation caused by 2ME was observed for the first time in BRAF V600E cells that are resistant to Vemurafenib (BRAF inhibitor) and double resistant to Vemurafenib and Trametinib (MEK inhibitor). The proliferation inhibition was related to the modulation of p21Cip1,Cyclin B1 and pRb, which are proteins involved in cell cycle regulation. Long exposure to 2ME in colony formation assay showed the inhibition of colony in all melanoma cell lines (regardless of resistance and mutational status), but not in primary fibroblasts, showing selective effect. In three-dimensional spheroid model, it was observed that the resistant (Sk-Mel- 28R) and double-resistant (Sk-Mel-28RT) cell lines were more invasive than the parental cell line (Sk-Mel-28). In this model, 2ME was able to inhibit cell invasion and cell viability. In the skin reconstruct model, in the absence of treatment, melanoma cell invasion can be observed in the dermis layer. However, after the treatment with 2ME these cell invasion foci are inhibited. Altogether, these effects demonstrate that 2ME is an effective anti-melanoma agent, regardless of resistance.

Le toucher est considéré comme l’un des sens primordiaux à modéliser chez un robot afin de lui permettre de générer des comportements plus souples et plus agiles comme attraper un objet, toucher (ou être touché par) une personne. Même si les capteurs tactiles actuels sont encore très limités en comparaison à la peau humaine, combinés à la vision et à la proprioception, le développement de nouveaux capteurs proches de la peau humaine pourrait démultiplier les capacités d’interactions d’un robot afin d’interagir directement avec une personne en toute sécurité et de partager avec lui son environnement physique et social. A la différence de la peau humaine, les principaux capteurs tactiles utilisés en robotique actuellement ne sont capables de détecter des variations de pression et de poids que sur de petites surfaces uniquement. De plus, ceux-ci sont souvent très rigides et n’ont pas les propriétés élastiques de déformation de la peau humaine. Les travaux de cette thèse se basent sur le développement d’une interface tactile proche d’une "peau artificielle" en terme de surface de recouvrement (qui peuvent atteindre plusieurs dizaines de centimètres carrés) et de localisation des points de contact de quelques dizaines de millinewtons. Deux aspects principaux sont développés : (i) aspect d’ingénierie comprenant le développement d’un prototype de peau artificielle conçue pour un robot humanoïde afin de lui conférer une perception tactile, et (ii) aspect cognitifs qui s’appuient sur l’intégration de multiples rétroactions sensorielles (tactile, visuelle, proprioceptive) dans le but d’avoir un robot qui puisse interagir physiquement avec des personnes.Le prototype tactile développé est basé sur la reconstruction du champ électrique à la surface d’un matériau conducteur, suivant le principe de la Tomographie par Impédance Électrique (TIE). Notre innovation principale a été d’implémenter des techniques d’apprentissage par réseau de neurones artificiels afin de reconstruire l’information sans utiliser les techniques analytiques d’inversion de matrice coûteuse en temps de calcul. De plus, nous montrons que l’utilisation de réseaux de neurones artificiels permet d’avoir un système beaucoup plus biomimétique, indispensable pour comprendre la perception du toucher chez l’être humain.Nous avons ensuite abordé le problème de l’intégration des informations tactiles et motrices. Après avoir recouvert un bras manipulateur avec la peau artificielle, nous avons fait apprendre un réseau de neurones son schéma corporel et adapter sa compliance par retour tactile. Le fonctionnement du moteur est basé sur le contrôle par admittance du bras robotique. Des expériences montrent que les réseaux de neurones peuvent contrôler l’interaction adaptative entre le bras du robot avec une personne grâce à l’estimation du couple appris selon la position où la force tactile avait été appliquée lors de la phase d’apprentissage.Enfin, nous nous sommes intéressées à la problématique de la représentation du corps au niveau neuronal, comment les êtres humains perçoivent leur propre corps à travers tous les sens (visuel, tactile et proprioceptif). Nous avons proposé un modèle biologique au niveau du cortex pariétal qui s’appuie sur l’intégration de multiples rétroactions sensorielles du corps du robot (son bras) et sur la synchronisation des rétroactions visuelles et proprioceptives. Nos résultats montrent l’apprentissage d’une image corporelle et l’espace péri-personnel avec l’émergence de neurones qui codent une information spatiale visuo-tactile relative au déplacement du bras et centrée soit sur le bras robotique soit centrée sur l’objet. / The touch perception is considered as one of the crucial senses to be recreated in a robot so that it could generate a more flexible and agile behavior. For instance, grasping an object, as well as touch or be touched by a person. Although modern touch sensors are still very limited compared to the human skin, combined with vision and proprioception, the development of new sensors similar to human skin could multiply the robot’s capacity to interact directly and safely with a person, as well as to share his or her physical and social environment.Unlike human skin, the main touch sensors used in modern robotics are only capable of detecting the pressure and weight variations on small batches of surface. Moreover, they are often quite stiff and do not have the elastic deformation capacity intrinsic to the human skin. The purpose of this thesis is to develop a touch interface close to "artificial skin" in terms of the covered area (which can reach several square decimeters) and localization of the contact points (several dozen millinewtons). Two main aspects have been developed: (i) the engineering aspect including the development of an artificial skin prototype for a humanoid robot designed to impart a tactile perception, and (ii) the cognitive aspect that is based on the integration of multiple sensory feedbacks (tactile, visual, proprioceptive) in order to conceive a robot that can physically interact with people.The developed tactile prototype is based on the reconstruction of the electric field on the surface of a conductive material, following the principle of Electrical Impedance Tomography (EIT). Our main innovation was to implement the neural network learning techniques to reconstruct the information without using the inverse matrix analytical techniques which imply time consuming computation. Moreover, we show that the application of artificial neural networks allows to obtain a much more biomimetic system, essential to understand the perception of the human touch.Then, we addressed the issue of integrating tactile and motor information. After having covered a manipulator arm with artificial skin, we have learn a neural network its body schema and enables it to adjust its compliance with tactile feedback. The functioning of the motor is based on the admittance control of the robot arm. Experiments show that neural networks can control the adaptive interaction between the robot arm and a human being by estimating the torque perceived according to the position where the touch force had been applied during the learning phase.Finally, we turned our attention to the issue of the body representation at the neuronal level, namely, how human beings perceive their own body through all their senses (visual, tactile, and proprioceptive). We have proposed a biological model in the parietal cortex, which is based on the integration of multiple sensory feedbacks from the robot’s body (its arm) and on the synchronization of visual and proprioceptive feedback. Our results show the capacity to perceive the body image with the emergence of neurons that encode a spatial visual-tactile information of the arm movement and is centered on either the robotic arm or on the object.

The next technological revolution, Industry 4.0, is envisioned as a digitally connected ecosystem where machines and gadgets are driven by artificial intelligence. By 2025, more than 75 billion devices are projected to serve this revolution. Many of which are to be integrated into the fabrics of everyday life in the form of smart wireless sensors. Still, two major challenges should be addressed to realize truly wireless and wearable sensors. First, the sensors should be flexible and stretchable, allowing for comfortable wearing. Second, the electronics should scavenge the energy it requires entirely from the environment, thus, eliminating the need for batteries, which are bulky, create ecological problems, etc. By addressing these two challenges, this dissertation paves the way for truly wearable sensors. The first part of the dissertation introduces a biocompatible magnetic skin with exceptional physical properties. It is highly-flexible, breathable, durable, and realizable in any desired shape and color. Attached to the skin of a user, the magnetic skin itself does not require any wiring, allowing to place the electronics and delicate components of the wireless sensor in a convenient nearby location to track the magnetic field produced by the magnetic skin. To demonstrate the performance of the magnetic skin, wearable systems are implemented as an assistive technology for severe quadriplegics, a touchless control solution for eliminating cross contaminations, and for monitoring blinking and eye movement for sleep laboratories. The second part of the dissertation is about wirelessly powering wireless sensors. In doing so, radio frequency (RF) rectifiers are a bottleneck, especially for ambient RF energy harvesting. Therefore, two RF rectifiers are introduced in standard CMOS technologies. The first architecture utilizes double-sided diodes to reduce the reverse leakage current, thus achieving a high dynamic range of 6.7 dB, -19.2 dBm sensitivity, and 86% efficiency. The second rectifier implements a dual-mode technique to lower the effective threshold voltage by 37%. Consequently, it achieves a 38% efficiency at −35 dBm input power and a 10.1 dB dynamic range while maintaining the same efficiency and sensitivity. Ultimately, combining these wireless powering techniques with the magnetic skin allows for truly wireless and wearable solutions.