Submitted by Renata Lopes (renatasil82@gmail.com) on 2017-09-05T19:49:54Z No. of bitstreams: 1 michelecamposalves.pdf: 4465106 bytes, checksum: 619948227c4d2d419bdbeb66a9cad785 (MD5) / Approved for entry into archive by Adriana Oliveira (adriana.oliveira@ufjf.edu.br) on 2017-09-06T11:36:14Z (GMT) No. of bitstreams: 1 michelecamposalves.pdf: 4465106 bytes, checksum: 619948227c4d2d419bdbeb66a9cad785 (MD5) / Made available in DSpace on 2017-09-06T11:36:14Z (GMT). No. of bitstreams: 1 michelecamposalves.pdf: 4465106 bytes, checksum: 619948227c4d2d419bdbeb66a9cad785 (MD5) Previous issue date: 2013-08-12 / CAPES - Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / Atualmente, existe um interesse crescente nas potenciais atividades de proteção à saúde do chá verde (Camellia sinensis), que é caracterizado pela presença de grandes quantidades de polifenois, sendo a maioria deles representados por catequinas. Galato de epigalocatequina (EGCG) é o componente mais abundante e ativo, sendo geralmente utilizado como biomarcador junto com a cafeína (CAF) e o ácido gálico (AG). EGCG é pouco absorvido no tratogastrointestinal e está sujeito ao metabolismo de primeira passagem pelo fígado quando administrado por via oral, forma mais comumente utilizada para o chá verde. Um sistema transdérmico poderia então ser um mecanismo de liberação alternativo. Dessa forma, o presente trabalho teve como objetivo: desenvolver e validar métodos por cromatografia em fase líquida de alta eficiência (CLAE) para a quantificação simultânea de EGCG, CAF e AG em cápsulas e emulsões contendo extrato seco de chá verde e determinar na emulsão a liberação de EGCG in vitro e sua permeação cutânea em pele humana, utilizando modelo ex vivo. Os métodos utilizaram CLAE de fase inversa e detecção em arranjo de fotodiodo. A separação foi atingida utilizando as seguintes condições: coluna octadecilsilano; fase móvel composta por água, etanol, acetato de etila e ácido acético (84:12:3:1, v/v/v/v); temperatura do compartimento para a coluna de 35 °C; e um sistema de fluxo por gradiente, alternando o fluxo entre 0.7 e 1.4 mL.min-1. Os compostos foram separados em 35 min. Os métodos foram simples, seletivos, precisos, exatos e rápidos; e mostraram a confiabilidade necessária para que fossem utilizados no controle de qualidade das formulações testadas. O método para a emulsão provou ser adequado para os ensaios de liberação in vitro e de permeação ex vivo. A taxa de liberação in vitro de EGCG foi de 8896.01 mg.cm-2, seguindo o modelo de pseudo-primeira ordem, também conhecido como modelo de Higuchi. O teste de permeação ex vivo mostrou que EGCG não é capaz de exercer suas atividades biológicas sistemicamente quando utilizado a partir da emulsão testada, permanecendo apenas no estrato córneo. / Currently, there is an increased interest in potential health-protective activities of green tea (Camellia sinensis), which is characterized by the presence of polyphenols huge amounts, being the majority of them catechins. Epigallocatechin 3- gallate (EGCG) is the most abundant and active component, been generally used as biomarker together with caffeine (CAF) and gallic acid (GA). EGCG is poorly absorbed in the gastrointestinal tract and is subject to first-pass metabolism by the liver when administered orally, the most widely used forms of green tea. A transdermal system could then be an alternative delivery mechanism. In this light, the present work aimed: to develop and validate high performance liquid chromatography (HPLC) methods for simultaneous quantification of EGCG, CAF and GA in emulsions and capsules containing dry extract green tea and to determine in emulsion the in vitro EGCG release and its human skin cutaneous permeation using ex vivo model. The methods used reversed phase HPLC and photodiode array detection. Separation was achieved using the follow conditions: octadecylsilyl column; mobile phase composed by water, ethanol, ethyl acetate and acetic acid (84:12:3:1, v/v/v/v); column temperature of 35 °C; and flow gradient syst em, alternating flow between 0.7 and 1.4 mL.min-1. The compounds were separated within 35 min. The methods were simple, selective, precise, accurate and fast; and provide the reliability required for them to be used for quality control of tested formulations. The emulsion method proved to be suitable both for in vitro release and ex vivo permeation. EGCG in vitro release rate was found to be 8896.01 μg.cm–2, following pseudo-first-order model, also known as Higuchi’s model. Ex vivo permeation testing showed that EGCG is not able to exert its biological activities systemically when used from tested emulsion, remaining only in the stratum corneum.

L'objectif de cette thèse était de mettre en oeuvre le développement d'une thérapie des plaies cutanées profondes basée sur l'utilisation de nanoparticules (NP) biodégradables de poly(acide lactique) (NP-PLA) vectrices de médiateurs de la cicatrisation. Le but était d'accélérer la cicatrisation cutanée et de favoriser la reconstruction d'un derme fonctionnel. La méthode a été de (i) réduire la réaction inflammatoire pour en contenir les effets délétères et (ii) stimuler la réépithélialisation pour accélérer la cicatrisation et réduire le risque infectieux. Les moyens ont été l'utilisation d'un antioxydant, la vitamine E (VE) et d'un facteur de croissance des fibroblastes (le FGF2) vectorisés par des nanoparticules biocompatibles et biodégradables de poly(acide lactique) (PLA). Nos NP-PLA contiennent l'antioxydant (VE) dans leur coeur hydrophobe, et portent le facteur de croissance fibroblastique (FGF2) à leur surface. Ces formulations ont été (i) caractérisées par des méthodes physico-chimiques et (ii) testées par des méthodes in vitro pour évaluer leurs effets potentiels, en tant que système de délivrance de VE et de FGF2, sur la cicatrisation des plaies. Des modèles expérimentaux in vivo ont été développés et caractérisés pour mettre en évidence l'efficacité des NP-PLA fonctionnalisées pour la cicatrisation cutanée et la reconstruction dermique fonctionnelle. Nos résultats montrent que l'activité antioxydante de la VE n'est pas perturbée par l'encapsulation dans des NP-PLA et qu'elle est légèrement supérieure à celle de la VE libre dans un système in vitro. De même, l'activité biologique du FGF2 sur la prolifération et la migration des fibroblastes dans un système in vitro n'est pas altérée par son adsorption sur des NP-PLA. Aucune de ces deux NP-PLA fonctionnalisées n'a de cytotoxicité avérée in vitro. Deux modèles expérimentaux de plaies cutanées profondes ont été développés sur souris sans poils SKH1 saines : (i) Un modèle robuste de brûlure cutanée thermique de 3ème degré qui se caractérise par une inflammation massive de la plaie et par un stade de granulation tardif après 16 jours de cicatrisation. (ii) Un modèle de plaie d'excision cutanée a également été utilisé. Un modèle de cicatrisation retardée a été obtenu par induction chimique d'un diabète de type I stable avant réalisation des plaies d'excision ou de brûlure. Ces modèles de plaies cutanées ont été caractérisés tout au long du processus de cicatrisation par des études (i) macroscopiques de cinétique de fermeture des plaies, (ii) histologiques d'inflammation, de nécrose et de réépithélialisation, (iii) physiologiques de perfusion sanguine cutanée. L'expression de 84 gènes impliqués dans le processus de cicatrisation a été étudiée sur le tissu cicatriciel 14 jours après formation de la plaie. Pour conclure, nos résultats mettent en évidence le potentiel de vectorisation de molécules thérapeutiques des NP de PLA pour le développement de futures stratégies de délivrance ciblée de VE et de FGF2 dans les plaies cutanées profondes. Les modèles expérimentaux in vivo développés et caractérisés, ouvrent la voie aux études précliniques d'efficacité des NP-PLA fonctionnalisées dans le processus de cicatrisation des plaies profondes / The objective of this thesis was to develop a therapy of cutaneous deep wounds based on biodegradable poly (lactic-acid) nanoparticles (PLA-NP) releasing wound healing mediators. The goal was to accelerate wound healing and to promote the reconstruction of a functional dermis. Our method was (i) to reduce the inflammatory reaction in the aim of limiting its deleterious effects, (ii) to stimulate reepithelialization to accelerate wound healing and to reduce the risk of infections. The implementation of means was based on the use of an antioxidant (vitamin E, VE) and a fibroblast growth factor (FGF2) carried by biocompatible and biodegradable poly(lactic-acid) based nanoparticles. Our PLA-NP contained the antioxidant (VE), in their hydrophobic core and carried the fibroblastic growth factor (FGF2) on their surface. These formulations were (i) characterized by physico-chemical methods and (ii) tested by in vitro methods to evaluate their effects as a delivery system of VE and FGF2 on wound healing. Experimental in vivo models have been developed and characterized in the aim of studying the potential beneficial effect of functionalized PLA-NP on wound healing and functional reconstruction of dermis. Our results show that the antioxidant activity of VE was not inhibited by encapsulation into PLA-NP and was lightly increased compared with free VE in an in vitro system. The biological activity of FGF2 on proliferation and migration of fibroblasts in an in vitro system was not altered by adsorption onto PLA-NP as well. No cytotoxicity of these functionalized PLA-NP was detected in vitro. Two experimental models of deep cutaneous wounds were developed on the healthy SKH1 hairless mouse: (i) A robust third degree thermal burn model that was characterized by massive inflammation of the wound and a late granulation stage after 16 days of healing. (ii) A model of excisional skin wound was also used. A model of delayed wound healing was established by chemical induction of stable type I diabetes prior to excision and burn injuries. The healing process of these models of cutaneous wounds was characterized by (i) macroscopic studies of wound closure, (ii) histological studies of inflammation, necrosis and reepithelialization, and (iii) by physiological studies of cutaneous blood perfusion. A study of the expression of 84 genes involved in the healing process was carried out on the scar tissue 14 days post-wound. In conclusion, our results highlight the potential efficacy of PLA-NP as a vector of therapeutic molecules for the development of future strategies for targeted delivery of VE and FGF2 in deep skin wounds. The developed and characterized in vivo experimental models open the way to preclinical studies of efficacy of functionalized PLA-NP on the healing process of deep wounds

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / This work had as main objective the development of nanostructured formulations containing rutin, using nanoparticles prepared with alternative vegetables oils. In the first chapter it was demonstrated the feasibility of preparing nanocapsules and nanoemulsions using grape seed or almond kernel oil. Nanocapsule suspensions and nanoemulsions were prepared by the interfacial deposition of preformed polymer and spontaneous emulsification, respectively. All formulations presented nanometric mean size, polydispersity index below 0.30, negative zeta potential, pH values between 6.5 and 7.5 remaining stable after 6 storage months. These formulations promoted the protection of the active against UV degradation, regardless of the type of the oily phase or vesicle. In the second chapter, formulations prepared with grape seed oil were selected for the development of rutin-loaded nanoparticles, as well as to evaluate its in vitro antioxidant activity and photostability. All formulations presented nanometric size, low polydispersity index, acid pH values, negative zeta potential and encapsulation efficiency close to 100 %. Nanoparticles were able to protect rutin against UV photodegradation, if compared to rutin ethanolic solution. In the study of the in vitro antioxidant activity, rutin-loaded nanocapsules and nanoemulsions showed a lower rutin decay rate compared to the rutin ethanolic solution when exposed to UV radiation in the presence of OH radical. However, its presence in nanocapsules led to a prolonged in vitro antioxidant activity compared to the rutin-loaded nanoemulsions. Finally, in the third chapter we studied the development of hydrogels containing rutin (free or associated to polymeric nanocapsules). Their activity on the cutaneous wound healing in rats was evaluated. The developed formulations showed adequate properties regarding their cutaneous administration. In vivo response concerning the healing effect of hydrogels was evaluated by the regression of skin lesions after six days of treatment. Markers of oxidative stress in the lesions of rats were also evaluated, as levels of lipid peroxidation analyzed by the method of thiobarbituric acid reactive substances (TBARS), determination of protein carbonyls levels, total proteins levels, glutathione (GSH) levels, vitamin C and evaluation of the antioxidant enzyme catalase (CAT). This last chapter showed for the first time the feasibility of the dermatological use of such formulations containing rutin to promote the in vivo wound healing. / Este trabalho teve como principal objetivo o desenvolvimento de formulações nanoestruturadas contendo rutina, a partir de nanopartículas contendo óleos vegetais até então não estudados no preparo destas formulações. No primeiro capítulo foi demonstrada a viabilidade de preparar nanocápsulas e nanoemulsões contendo o óleo de semente de uva e de amêndoas doce como fase oleosa. As suspensões de nanocápsulas e nanoemulsões foram preparadas pelo método da deposição interfacial do polímero pré-formado e emulsificação espontânea, respectivamente. Todas as formulações apresentaram tamanho médio nanométrico, índice de polidispersão inferior a 0,30, potencial zeta negativo e valores de pH entre 6,5 e 7,5, permanecendo estáveis após 6 meses de armazenamento. As formulações promoveram a fotoproteção do ativo frente à degradação UV, independente do tipo de fase oleosa e do tipo de vesícula. No segundo capítulo, as formulações contendo o óleo de semente de uva foram selecionadas para o estudo do desenvolvimento de nanopartículas contendo rutina, bem como avaliação de sua atividade antioxidante in vitro e fotoestabilidade. Após o preparo, todas as formulações apresentaram tamanho nanométrico de partículas, baixo índice de polidispersão, valores de pH ácido, potencial zeta negativo e eficiência de encapsulação próxima à 100%. As nanopartículas protegeram a rutina frente à fotodegradação UV, quando comparadas à solução etanólica. No estudo da atividade antioxidante in vitro, as nanocápsulas e nanoemulsões apresentaram uma menor taxa de decaimento da rutina comparada com a solução etanólica, quando expostas à radiação UV em presença do radical ·OH. No entanto, a presença das nanocápsulas levou a uma atividade antioxidante in vitro mais prolongada comparada com as nanoemulsões contendo rutina. Finalmente, no terceiro capítulo estudamos o desenvolvimento de hidrogéis contendo rutina livre ou associada a nanocápsulas poliméricas, avaliando a sua atividade sobre a cicatrização de lesões cutâneas em ratos. As formulações desenvolvidas apresentaram propriedades adequadas para aplicação tópica. A resposta in vivo do efeito cicatrizante foi avaliada através da regressão de lesões na pele após 6 dias de tratamento. Marcadores do estresse oxidativo nas lesões dos ratos foram também avaliados, como os níveis da peroxidação lipídica através do método de TBARS, níveis de proteína carbonilada, níveis de proteínas totais, níveis de glutationa (GSH), vitamina C e avaliação da enzima antioxidante catalase (CAT). Este terceiro capítulo demonstrou pela primeira vez a potencialidade do emprego dermatológico de hidrogéis contendo rutina para a promoção de cicatrização de feridas in vivo.

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / Nanostructured lipid carriers (NLCs) have been developed in order to improve cutaneous hydration. Due to its ultrafine particle size it is possible a film formation on the skin which decreases transepidermal water loss, improving occlusive effect and hydration. In this context, the aim of this work was to develop a formulation containing NLCs based in Ouratea sp. butter (Ochnaceae) as a strategy to increase cutaneous hydration. Firstly, it was performed a characterization of the components such as, stearic acid (SA), Ouratea sp. butter (OB), Phospholipon 90G®, and its physical mixtures by Differential Scanning Calorimetry (DSC), Thermogravimetry (TG) and X ray Diffraction (XRD). The NLCs were prepared using the diffusion solvent method and evaluated for particle size, polidispersity index, zeta potential, pH and conductivity. Moreover, NLCs were characterized by DSC, XRD and Transmission electron microscopy (TEM). Finally, the NLCs were incorporated into a cream base and were evaluated for rheological behavior, occlusive effect and potential cutaneous hydration in vitro. From the raw materials characterization was possible to observe in which the PM1 (SA + OB) 1:1 showed a slightly shift of the melting point corresponding to SA (57 to 50°C) and presented an amorphous portion at ~ 20° in the XRD analysis. These observations could be attributed to a decrease of crystallinity when compared with solid lipid pure, highly ordered. With the addition of Phospholipon® 90G (PM2), the melting point was highly shifted to lower temperature (57 to 34°C), and presented a pronounced amorphous at ~ 20° in XRD. Since the NLCs need to remain solid at room and body temperature, it is suggested that using a lower proportion of the Phospholipon® 90G in NLCs formulations. From the CMC determination of SDS surfactant (8,02 mmol L-1) was possible to suggest a range concentration of the SDS in the NLCs preparations. The formulation with the highest SDS concentration (24 mmol L-1) demonstrated the lowest size particle (~ 240 nm), however there were no difference in the zeta potential over formulation that using 10 mmol L-1 of SDS (~ -50 mV). The NLCs presented a decrease in the enthalpy (ΔHNLC8 = 22.2, ΔHNLC10 = 23.2, ΔHNLC24 = 7.2 J/g) and also an increase in the width peak when compared with solid lipid SA pure (ΔHSA = 164.1). In addition, the XRD analysis demonstrated that NLCs presented main peaks both formulations are similar to those found in the SA (23.93°, 25.09°, 27.98°), but with a lower intensity. These observations suggest a lipid matrix less ordered which result in NLC formation. TEM results confirmed previous data related to NLCs particle size (200 to 300 nm). After the incorporation of the NLC10 into a cream base, it was possible to observe an increase in the occlusive effect and potential cutaneous hydration. Therefore, NLCs containing Ouratea sp. butter can be an attractive nanotechnology way to increase cutaneous hydration. / Carreadores Lipídicos nanoestruturados (CLNs) têm sido desenvolvidos visando uma melhor hidratação cutânea. Devido ao seu reduzido tamanho de partícula, há formação de um filme sobre a pele o qual diminui a perda de água transepidermal, aumentando a hidratação. Neste contexto o objetivo deste trabalho foi desenvolver uma formulação contendo CLNs à base de manteiga de Ouratea sp. (Ochnaceae) como uma estratégia para o aumento do efeito oclusivo e da hidratação cutânea. Primeiramente foi realizada a caracterização das matérias-primas como, ácido esteárico (AE), Manteiga Ouratea sp.(MO), Phospholipon 90G® e suas misturas físicas utilizando Calorimetria Exploratória Diferencial (DSC), Termogravimetria (TG) e Difração de raios-X (DRX). Os CLNs foram preparados pelo método de difusão do solvente e avaliados por diâmetro médio de partícula, índice de polidispersão, potencial zeta, pH e condutividade. Os CLNs ainda foram caracterizados por DSC e DRX, Microscopia eletrônica de Transmissão (MET). Por fim, os CLNs foram incorporados no creme base e foram avaliados quanto ao comportamento reológico, efeito oclusivo e potencial hidratação cutânea in vitro. Através da caracterização das matérias-primas foi possível observar que a MF1 (AE + MO) 1:1 mostrou um pequeno deslocamento do ponto de fusão correspondente ao AE (57 a 50°C) e apresentou uma porção amorfa em ~ 20° nas análises de DRX. Essas observações podem ser atribuídas à diminuição da cristalinidade quando comparado com o lipídeo sólido puro, altamente ordenado. Com a adição do Phospholipon 90G® (MF2), o ponto de fusão foi deslocado para uma temperatura menor (57 a 34°C), e apresentou uma pronunciada porção amorfa em ~ 20° no DRX. Partindo do pressuposto que os CLNs devem se manter sólidos a temperatura ambiente e corpórea, é sugerida a utilização de uma menor proporção de Phospholipon® 90G nas formulações de CLNs. A partir da determinação da CMC do tensoativo SDS (8,02 mmol L-1) foi possível sugerir uma faixa de concentração de SDS nas preparações de CLNs. A formulação com a maior concentração de SDS (24 mmol L- 1) demonstrou o menor diâmetro de partícula (~ 240 nm), entretanto, não houve diferenças no potencial zeta em relação a formulação em que foi utilizado 10 mmol L-1 of SDS (~ -50 mV). CLNs apresentaram uma diminuição nos valores de entalpia (ΔHCLN8 = 22,2; ΔHCLN10 = 23,2; ΔHCLN24 = 7,2 J/g) e também um aumento da largura do pico quando comparados com o AE puro (ΔHSA = 164,1). Além disso, as análises de DRX demonstraram que os CLNs apresentaram picos principais similares àqueles encontrados no AE (23,93°; 25,09°; 27,98°), porém com uma menor intensidade. Os resultados de MET confirmaram os dados de diâmetro médio de partícula dos CLNs (200 a 300 nm). Após a incorporação do CLN10 no creme base, foi possível observar um aumento no efeito oclusivo e na hidratação cutânea. Dessa forma os CLNs contendo manteiga de Ouratea sp. podem ser uma alternativa nanotecnológica atrativa para o aumento da hidratação cutânea.

No description available.

Background: Topical negative pressure wound therapy (TNPWT) is a regularly used method in modern wound treatment with a growing and diverse potential for clinical use. So far positive effects on microcirculation have been observed and examined, although precise statements on the underlying mechanism appear unsatisfying. Objective: The aim of our study was to extend the understanding of the effect of TNPWT on tissue perfusion and determine the time frame and the extent to which the tissue perfusion changes due to TNPWT. Material and methods: TNPWT was applied to the anterior thighs of 40 healthy individuals for 30 min, respectively. Before and up to 90 min after the application, measurements of the amount of regional haemoglobin (rHb), capillary venous oxygen saturation (sO2), blood flow (flow) and velocity were conducted with spectrophotometry (combining white light spectrometry and laser Doppler spectroscopy) within two different depths/skin layers. A superficial measuring probe for depths up to 3 mm and a deep measuring probe for up to 7 mm were used. Results: All parameters show significant changes after the intervention compared to baseline measurements. The greater effect was seen superficially. The superficially measured rHb, sO2 and flow showed a significant increase and stayed above the baseline at the end of the protocol. Whereas deeply measured, the rHb initially showed a decrease. The flow and sO2 showed a significant increase up to 60 min after the intervention. Conclusion: The application of TNPWT on healthy tissue shows an increase in capillary-venous oxygen saturation and haemoglobin concentration of at least 90 min after intervention. A possible use in clinical practice for preconditioning to enhance wound healing for high-risk patients to develop wound healing disorder, requires further studies to investigate the actual duration of the effect.

Le but de notre étude visait à déterminer si les matrices ECM (extracellular matrix) préparés à partir d'un modèle vertébré (Axolotl) capables de régénérer ses tissus suite à une blessure sont plus efficaces pour stimuler les réponses régénératives chez les animaux non régénérant (par exemple les mammifères). Nous avons testé la capacité de matrice ECM axolotl à améliorer la guérison des plaies cutanées dans des souris et nous les avons comparés à une matrice disponible commercialement (échafaudage Symbios PerioDerm) pour leur efficacité à favoriser la guérison des plaies. Des lésions d'excision ont été créées sur le dos de souris et les animaux ont été regroupés dans différents groupes; a-) ECM de peau axolotl décellularisée (groupe Axolotl), b-) matrice de derme acellulaire Symbios Perioderm (groupe PerioDerm), c-) grillage en titane (groupe témoin); respectivement. Les tissus des plaies ont été récoltés à des moments précis : 7 jours et 30 jours après la blessure pour évaluer la guérison des plaies. La guérison des blessures ayant reçu les différentes matrices a été comparées entre elles en utilisant le test de transillumination et des analyses histologiques. Les résultats indiquent que la ECM de peau d’axolotl décellularisée est bien tolérée par les souris, car aucun rejet n'a été observé. Le groupe qui a reçu l'ECM de la peau axolotl décellularisé a démontré une réépithélialisation, une densité cellulaire, une teneur en collagène (avec une organisation similaire à un tissu intact) et une vascularisation (angiogenèse) élevées par rapport aux groupes PerioDerm et témoins. La présence de follicules pileux était également observé dans le groupe axolotl (qui n'est pas présent dans PerioDerm et groupes de contrôle). Sur la base de nos résultats, l'hypothèse de base semble être correcte en ce qu'une matrice ECM provenant d'un régénérateur puissant semble favoriser la guérison plus efficacement chez les animaux normalement non régénérants. Cependant, des recherches supplémentaires devront être menées pour confirmer ces résultats. / The aim of our study sought to determine whether ECM scaffolds prepared from a vertebrate model (Axolotl) capable of regenerating tissues following injury are more effective at stimulating regenerative responses in non-regenerating animals (e.g., mammals). We tested the ability of axolotl decellularized ECM scaffolds to improve skin wound healing in mammalian models and compare the axolotl skin ECM scaffold to a commercially available one (Symbios PerioDerm scaffold) for efficiency in promoting wound healing. Excisional lesions were created on the back of mice, and animals in different groups were treated by; a-) decellularized axolotl skin ECM (Axolotl group), b-) Symbios Perioderm acellular dermis scaffold (PerioDerm group), d-) Titanized mesh only (Control group); respectively. Wound tissues were harvested at time points: 7- and 30-days post-wounding to assess the scaffolds impact on wound healing. Wound healing was compared between the Axolotl, PerioDerm and Control groups using transillumination test and histological analyses, Results indicate that the decellularized axolotl skin ECM is well tolerated by mammalian models, as no immune rejection was observed. The axolotl group that received the decellularized Axolotl Skin ECM demonstrated high reepithelialization, cellular density, collagen content (in a porous pattern similar to intact skin), vascularization (angiogenesis) compared to PerioDerm and control groups. The presence of hair follicles was also observed in the axolotl group (which is not present in PerioDerm and control groups). Based on our results, the basic hypothesis appears to be correct in that an ECM scaffold from a strong regenerator seems to promote healing more efficiently in non-regenerating animals. However, further research should be conducted to confirm these findings.

No description available.

A rutina é empregada como antioxidante e na prevenção da fragilidade capilar. Pode ser veiculada em emulsões tópicas adequadas para atingir o local de ação. Estudos de penetração in vitro através da pele humana seria a situação ideal, entretanto, há dificuldades de sua obtenção e manutenção de sua viabilidade. Entre os demais modelos de membrana, a muda de pele de cobra apresenta-se como estrato córneo puro, fornecendo barreira similar ao humano e é obtida sem a morte do animal. Os objetivos desta pesquisa foram: (1) desenvolver e avaliar a estabilidade de emulsões cosméticas, contendo rutina e promotores de penetração cutânea, tais como, uréia (U), isopropanol (IP) e propilenoglicol (PG); (2) avaliar a liberação da referida substância ativa das emulsões e; (3) avaliar a penetração e a retenção cutânea in vitro da rutina da formulação de melhor desempenho. Emulsões foram desenvolvidas com rutina a 5,0% p/p e U, IP e PG, associados ou não e em proporções distintas, segundo planejamento fatorial com dois níveis com ponto central. Quantificou-se a rutina das emulsões por espectrofotometria a 361,0 nm, método previamente validado. A liberação da rutina nas formulações foi realizada em células de difusão vertical com membrana de acetato de celulose e água destilada e álcool etílico absoluto 99,5% (1:1), como fluido receptor. O experimento foi conduzido em um período de seis horas, a 37,0 ±. 0,5 °C e agitação constante de 300 rpm.>f. emulsão de melhor desempenho quanto à liberação foi estudada quanto à estabilidade (Testes de Estabilidade Acelerada). Para o estudo de penetração e retenção cutânea da rutina dessa formulação foi utilizada muda de pele de cobra de Crotalus durissus. Empregou-se o método espectrofotométrico validado a 410,0 nm para a quantificação da rutina após liberação, penetração e retenção cutânea. Todas as emulsões foram consideradas adequadas após desenvolvimento das formulações. A uréia (isolada e em associação com IP e PG) e o isopropanol (isolado e em associação com PG) influenciaram negativamente a liberação da rutina das emulsões em diversos parâmetros. A rutina liberada e acumulada da formulação contendo PG a 5,0% p/p possuiu valor de 648,80 ±. 53,01 &#181g/cm2. Fora do esperado, a preparação contendo o número maior de promotores (U 5,0% p/p, IP 5,0% p/p e PG 5,0% p/p) resultou em liberação de menor magnitude igual a 419,76 ±. 17,98 &#181g/cm2. A presença do PG apresentou-se mais eficiente na liberação da rutina, mas não na sua penetração através da muda de pele de C. durissus, retendo 0,931 ± 0,0391 µg de rutina/mg de muda de pele de cobra. Nas condições de armazenamento a 25,0 ±2,0 °C; 5,0 ±0,5 °C e 45,0 ±. 0,5 °C, a emulsão com PG e rutina apresentou-se quimicamente estável durante 30 dias. De acordo com os resultados, a emulsão contendo PG apresentou liberação mais expressiva da rutina, no entanto, não ocorreu a penetração cutânea, mas apenas sua retenção no estrato córneo de C. durissus. A preparação manteve-se estável em todas as condições de armazenamento. / Rutin is employed as antioxidant and to prevent the capillary fragility and, when incorporated in cosmetic emulsions, it must target the action site. In vitro cutaneous penetration studies through human skin is the ideal situation, however, there are difficulties to obtain and to maintain this tissue viability. Among the membrane models, the shed snake skin presents itself as pure stratum corneum, providing barrier function similar to human and it is obtained without the animal sacrifica. The objectives of this research were: (1) development and stability evaluation of cosmetic emulsions containing rutin and penetration enhancers, as urea (U), isopropanol (IP) and propylene glycol (PG); (2) release evaluation of the mentioned active substance from the emulsions and; (3) evaluation of rutin in vitro cutaneous penetration and retention from the emulsion of the best performance. Emulsions were developed with rutin 5.0% w/wand U, IP and PG, associated or not according to factorial design with two levels and central point. Active substance on the formulations was quantified by a validated spectrophotometric method at 361.0 nm. Rutin release from emulsions was performed in vertical diffusion cells with cellulose acetate membrane and distilled water and ethanol 99.5% (1:1), as receptor fluid. The experiment was conducted for six hours, at 37.0 ± 0.5 °c with constant stirring of 300 rpm. Formulation with best profile of rutin release had its stability studied by the Accelerated Stability Assays. Rutin cutaneous penetration and retention from the mentioned emulsion was performed with shed snake skin from Crotalus durissus. Spectrophotometry at 410.0 nm, previously validated, determined the active substance after release and cutaneous penetration/retention. Ali emulsions were considered apparently stables after development. Urea (isolated and associated with IP and PG) and isopropanol (isolated and associated with PG) have influenced negatively the rutin release in several parameters. Emulsion with PG 5.0% w/w presented rutin released and accumulated equal to 648.80 ± 53.01 µg/cm2. Unexpectedly, the formulation containing all enhancers (U 5.0% w/w, IP 5.0% w/w and PG 5.0% w/w) has decreased the amount released of the active substance (419.76 ± 17.98 µg/cm2). Emulsion with PG presented more adequate for rutin release, but PG did not provide rutin cutaneous penetration through C. durissus skin, retaining 0.931 ± 0.0391 &$181;g rutin/mg shed snake skin. The referred formulation was chemically stable for 30 days after they have been stored at 25.0 ± 2.0 °c, 5.0 ± 0.5 °c and 45.0 ± 0.5 °C. In conclusion, emulsion with PG provided rutin release more expressively, although, it has not been verified the active cutaneous penetration, but only its retention on the Crotalus durissus stratum corneum. Formulation was stable in all storage conditions.

Na busca por espécies vegetais com atividade antileishmania, selecionaram-se, para o estudo, duas espécies bem conhecidas da família Asteraceae: Artemisia annua L. e Bidens pilosa L. Ambas são reconhecidamente utilizadas na medicina popular, como antiprotozoárias. Apesar de terem sido amplamente estudadas em diversos aspectos, alguns permaneceram inexplorados, até o momento, e foram abordados, neste trabalho. As duas espécies foram analisadas quanto aos aspectos químico e biológico de extratos (hidroetanólico e infuso) e frações orgânicas selecionados, em função da atividade antileishmania in vitro, frente às formas promastigotas de Leishmania amazonensis. Os extratos foram obtidos a partir de órgãos vegetais, em estados de conservação diferentes (in natura, droga) e coletados em fenofases distintas. Extratos e frações orgânicas das espécies estudadas mostraram promissora atividade antileishmania in vitro e baixo nível de citotoxicidade in vitro em células epiteliais humanas (HEP-2). No estudo químico dos extratos e frações bioativos, realizaram-se análises qualitativas e/ou quantitativas de terpenos, flavonóides e de marcadores específicos (artemisinina, quercetina e rutina), avaliando-se a variação da composição dos mesmos, nas diferentes fenofases consideradas. Discutiram-se as possíveis relações existentes entre a composição química e a atividade biológica verificada. Aspectos inéditos do estudo morfoanatômico de partes aéreas de A. annua foram descritos. / Plants are potential sources of new antileishmanial drugs. Two well-known antiprotozoal species were selected, from the Asteraceae family, for this study: Artemisia annua L and Bidens pilosa L. Despite the traditional and scientific accumulated knowledge, some aspects were not investigated before and were the subject of this work. Several extracts (infusions and ethanol 96 °GL) and selected fractions from both species were evaluated according to the different parameters, such as: plant organs and/or parts, growth stages and drying state of starting materials (fresh, drug). Fractions were selected among those more active against promastigotes of Leishmania amazonensis. Ethanol extracts and their fractions showed a high level of in vitro antileishmanial activity and a low cytotoxicity on epithelial human cells (HEP-2). Qualitative and/or quantitative analysis of extracts and fractions were performed for terpenes, flavonoids and selected markers (artemisinin, quercetin and rutin) in order to characterize them and evaluate variations during the different growth stages. Correlations of the chemical composition and the biological activity were discussed. The main anatomical characters of the aerial parts of A. annua were described for the first time and illustrated by photomicrographs.