Avaliação da atividade antitumoral, antioxidante e anti-edematogênica de nanocápsulas de núcleo lipídico contendo resveratrol e/ou curcumina

A busca por tratamentos mais efetivos é um dos principais estímulos para novas pesquisas na área farmacêutica, dando suporte ao emprego da nanotecnologia. Além disso, a utilização de substâncias presentes em plantas, frutas e especiarias para o tratamento de doenças como câncer, associadas à inflamação e ao estresse oxidativo, têm despertado grande interesse da comunidade científica. No presente estudo, nanocápsulas de núcleo lipídico contendo resveratrol e/ou curcumina, dois fitoquímicos com inúmeros benefícios à saúde, foram desenvolvidas com o intuito de contornar as suas limitações e aumentar a sua eficácia terapêutica. Primeiramente, uma avaliação dos principais benefícios da nanoencapsulação da curcumina, tanto em sistemas lipídicos como poliméricos, foi realizada. Uma criteriosa revisão da literatura acerca das aplicações in vivo da curcumina nanoencapsulada demonstrou que os sistemas carreadores nanoestruturados são uma excelente ferramenta para aumentar a sua biodisponibilidade, a sua vetorização ao sistema nervoso central, bem como, seus efeitos terapêuticos. Na primeira etapa experimental deste estudo, o efeito antitumoral da curcumina associada à nanocápsulas de núcleo lipídico foi avaliado em modelo experimental de gliomas. Após ensaios in vitro, a maior citotoxicidade da curcumina nanoencapsulada com relação à curcumina em solução foi dependente da linhagem celular utilizada. No modelo in vivo de gliomas foi observado uma redução no tamanho e na malignidade do tumor, e um aumento da sobrevida dos animais tratados com a curcumina associada às nanocápsulas. Em um segundo momento, nanocápsulas de núcleo lipídico contendo resveratrol e/ou curcumina com adequadas propriedades nanotecnológicas foram desenvolvidas. A associação dos polifenóis não influenciou nas características físico-químicas das suspensões. A nanoencapsulação foi capaz de diminuir a fotodegradação dos polifenóis e a co-encapsulação aumentou a fotoestabilidade do resveratrol. Ainda, a nanoencapsulação aumentou a atividade antioxidante dos polifenóis e um efeito mais pronunciado foi observado devido à co-encapsulação. Os resultados promissores obtidos in vitro estimularam a avaliação dos efeitos in vivo das nanocápsulas de núcleo lipídico contendo os polifenóis individualmente ou co-encapsulados. Para tal, utilizou-se o modelo animal de artrite induzida pelo Adjuvante Completo de Freund (CFA). Neste modelo, apenas os polifenóis nanoencapsulados foram capazes de reduzir o edema causado pela administração do CFA. Uma maior atividade anti-edematogênica foi observada após a administração das nanocápsulas contendo a associação dos polifenóis, estando esta atividade acompanhada de uma redução do dano à cartilagem e ao osso e reduzida infiltração linfocitária no soro dos animais. Os resultados nesta tese demonstraram a viabilidade tecnológica da preparação de nanocápsulas de núcleo lipídico contendo resveratrol e/ou curcumina, com potencial aplicação para o tratamento de tumores cerebrais e de doenças relacionadas ao estresse oxidativo e processos inflamatórios.The search for more effective treatments is one of the main stimuli for new researches in the pharmaceutical area, supporting the use of nanotechnology. Furthermore, the utilization of substances found in plants, fruits and spices to treat diseases like cancer, associated with inflammation and oxidative stress, have attracted great interest from the scientific community. In the present study, lipid-core nanocapsules containing resveratrol and/or curcumin, two phytochemicals with innumerable health benefits, were developed in order to circumvent its limitations and enhance their therapeutic efficacy. First, a review of the main benefits of curcumin- loaded to nanostructured systems, whether lipidic, polymeric or other type, were performed. A discerning review concerning in vivo applications of curcumin nanoencapsulated demonstrated that nanostructured carrier systems are an excellent tool to increase their bioavailability, the targeting to the central nervous system as well as its therapeutic effects. In the first experimental phase of this study, the antitumor effect of curcumin loaded in lipid-core nanocapsules was evaluated in experimental model of gliomas. After in vitro assays, the highest cytotoxicity of nanoencapsulated curcumin compared with solution of curcumin was dependent on the cell line used. In the in vivo model of gliomas, a reduction in the size and malignancy of the tumor and an increase in the survival rate were observed after treatments with curcumin-loaded nanocapsules. In a second step, lipid-core nanocapsules containing the association of resveratrol and curcumin with adequate nanotechnological properties were developed. The association of polyphenols did not influence the physicochemical characteristics of the suspensions. The nanoencapsulation was able to decrease the phodegradation of polyphenols and the co-encapsulation improved resveratrol photostability. Moreover, the nanoencapsulation increased the antioxidant activity of polyphenols and a more pronounced effect was observed due to the co-encapsulation. The promising in vitro results which were obtained stimulated the evaluation of in vivo effects of polyphenols encapsulated individually or associated in lipid-core nanocapsules. To that, a Complete Freund’s Adjuvant (CFA)-induced arthritis model was used. In this model, only nanoencapsulated polyphenols were able to reduce the swelling caused by the administration of CFA. A higher anti-edematogenic activity was observed after administration of nanocapsules containing the combination of polyphenols, which was accompanied by a reduction of the damage to the cartilage and bone and reduced lymphocyte infiltration in the serum of animals. The results of this thesis demonstrated the technical feasibility of preparation lipid-core nanocapsules containing resveratrol and/or curcumin, with a potential application for the treatment of brain tumors and diseases related with oxidative stress and inflammation process

Protocolo clínico para acidente vascular cerebral: desenvolvimento de um instrumento informativo

The objective was to know what health professionals understand about the Clinical Protocol and Therapeutic Guidelines - Thrombolysis in Acute Ischemic Stroke and to develop an informative instrument about it, to be used in the Emergency Care Unit, the study scenario. This is an exploratory, descriptive research, with a qualitative approach, carried out with five nurses and five doctors in an Emergency Care Unit, in the state of Rio Grande do Sul, Brazil. Data were collected during the months of October to November 2019, through semi-structured interviews, and were subjected to content analysis. The results enabled five categories: Clinical protocol as a criterion for thrombolysis; how to use medication and imaging exams; as standardization / organization of care; standardization, cost reduction and clinical improvement of the patient; need for qualification of human, physical and structural resources. In conclusion, the professionals understand that the realization of the protocol would assist in the organization of the service and standardization, as well as in the reduction of expenses and clinical improvement of the patient.Objetivou-se conhecer o que os profissionais de saúde compreendem sobre o Protocolo Clínico e Diretrizes Terapêutica - Trombólise no Acidente Vascular Cerebral Isquêmico Agudo e, desenvolver um instrumento informativo acerca deste, para ser utilizado na Unidade de Pronto Atendimento, cenário do estudo. Trata-se de uma pesquisa exploratória, descritiva, de abordagem qualitativa, realizada com cinco enfermeiros e cinco médicos em uma Unidade de Pronto Atendimento, no estado do Rio Grande do Sul, Brasil. Os dados foram coletados, durante os meses de outubro a novembro de 2019, por meio de entrevista semiestruturada, sendo submetidos a análise de conteúdo. Os resultados possibilitaram cinco categorias: Protocolo clínico como critério para trombólise; como utilização de medicação e exames de imagem; como padronização/organização do atendimento; padronização, redução de gastos e melhora clínica do paciente; necessidade de qualificação de recursos humanos, físicos e estruturais. Como conclusão, os profissionais entendem que a realização do protocolo auxiliaria na organização do serviço e padronização, bem como na redução de gastos e melhora clínica do paciente.El objetivo era saber qué entienden los profesionales de la salud sobre el Protocolo Clínico y las Pautas Terapéuticas - Trombolisis en el accidente cerebrovascular isquémico agudo y desarrollar una herramienta de información al respecto, que se utilizará en la Unidad de Atención de Emergencia, el escenario del estudio. Esta es una investigación exploratoria, descriptiva, con un enfoque cualitativo, realizada con cinco enfermeras y cinco médicos en una Unidad de Atención de Emergencia, en el estado de Rio Grande do Sul, Brasil. Los datos se recopilaron durante los meses de octubre a noviembre de 2019, a través de entrevistas semiestructuradas, y se sometieron a análisis de contenido. Los resultados permitieron cinco categorías: protocolo clínico como criterio para la trombolisis; cómo usar medicamentos y exámenes de imágenes; como estandarización/organización de la atención; estandarización, reducción de costos y mejora clínica del paciente; necesidad de calificación de recursos humanos, físicos y estructurales. En conclusión, los profesionales entienden que la realización del protocolo ayudaría en la organización del servicio y la estandarización, así como en la reducción de gastos y la mejora clínica del paciente

Intracellular Delivery of Poorly Soluble Polyphenols: Elucidating the Interplay of Self-Assembling Nanocarriers and Human Chondrocytes

Increased molecular understanding of multifactorial diseases paves the way for novel therapeutic approaches requiring sophisticated carriers for intracellular delivery of actives. We designed and characterized self-assembling lipid-core nanocapsules for coencapsulation of two poorly soluble natural polyphenols curcumin and resveratrol. The polyphenols were identified as high-potential therapeutic candidates intervening in the intracellular inflammation cascade of chondrocytes during the progress of osteoarthritis. To elucidate the interplay between chondrocytes and nanocapsules and their therapeutic effect, we pursued a complementary analytical approach combining label-free visualization with biological assays. Primary human chondrocytes did not show any adverse effects upon nanocapsule application and coherent anti-Stokes Raman scattering images visualized their intracellular uptake. Further, by systematically blocking different uptake mechanisms, an energy independent uptake into the cells could be identified. Additionally, we tested the therapeutic effect of the polyphenol-loaded carriers on inflamed chondrocytes. Treatment with nanocapsules resulted in a major reduction of nitric oxide levels, a well-known apoptosis trigger during the course of osteoarthritis. For a more profound examination of this protective effect on joint cells, we pursued studies with atomic force microscopy investigations. Significant changes in the cell cytoskeleton as well as prominent dents in the cell membrane upon induced apoptosis were revealed. Interestingly, these effects could not be detected for chondrocytes which were pretreated with the nanocapsules. Overall, besides presenting a sophisticated carrier system for joint application, these results highlight the necessity of establishing combinatorial analytical approaches to elucidate cellular uptake, the interplay of codelivered drugs and their therapeutic effect on the subcellular level

Intracellular Delivery of Poorly Soluble Polyphenols: Elucidating the Interplay of Self-Assembling Nanocarriers and Human Chondrocytes

Increased molecular understanding of multifactorial diseases paves the way for novel therapeutic approaches requiring sophisticated carriers for intracellular delivery of actives. We designed and characterized self-assembling lipid-core nanocapsules for coencapsulation of two poorly soluble natural polyphenols curcumin and resveratrol. The polyphenols were identified as high-potential therapeutic candidates intervening in the intracellular inflammation cascade of chondrocytes during the progress of osteoarthritis. To elucidate the interplay between chondrocytes and nanocapsules and their therapeutic effect, we pursued a complementary analytical approach combining label-free visualization with biological assays. Primary human chondrocytes did not show any adverse effects upon nanocapsule application and coherent anti-Stokes Raman scattering images visualized their intracellular uptake. Further, by systematically blocking different uptake mechanisms, an energy independent uptake into the cells could be identified. Additionally, we tested the therapeutic effect of the polyphenol-loaded carriers on inflamed chondrocytes. Treatment with nanocapsules resulted in a major reduction of nitric oxide levels, a well-known apoptosis trigger during the course of osteoarthritis. For a more profound examination of this protective effect on joint cells, we pursued studies with atomic force microscopy investigations. Significant changes in the cell cytoskeleton as well as prominent dents in the cell membrane upon induced apoptosis were revealed. Interestingly, these effects could not be detected for chondrocytes which were pretreated with the nanocapsules. Overall, besides presenting a sophisticated carrier system for joint application, these results highlight the necessity of establishing combinatorial analytical approaches to elucidate cellular uptake, the interplay of codelivered drugs and their therapeutic effect on the subcellular level