Navegando por Autor "Marcolino, Luciana Maria Cortez"
Agora exibindo 1 - 6 de 6
Resultados por página
Opções de Ordenação
Item Efeito da terapia fotodinâmica com curcumina em promastigotas e curcumina nanoencapsulada em amastigotas de Leishmania braziliensis e Leishmania major(2021-09-24) Ferreira-Strixino, Juliana; Soares, Cristina Pacheco; Sant'Anna, Luciana Barros; Ribeiro, Martha Simões; Tada, Dayane Batista; Marcolino, Luciana Maria Cortez; São José dos CamposA leishmaniose tegumentar é uma doença zoonótica, transmitida por fêmeas de flebotomíneos e causada por protozoários do gênero Leishmania. É considerada pela OMS um problema de saúde pública que nos últimos anos apresentou aumento significativo, tanto no número de casos quanto na distribuição geográfica. O tratamento convencional é tóxico e contraindicado para pacientes que apresentam algumas comorbidades e pode causar diversos efeitos colaterais graves. Tendo em vista uma alternativa de tratamento não agressivo vem se estudando a Terapia Fotodinâmica, que consiste em um tratamento não invasivo, não apresenta efeitos colaterais, de fácil acesso que utiliza substâncias de baixo custo, como a curcumina. Este estudo teve como objetivo avaliar os efeitos da curcumina livre em promastigotas e curcumina nanoencapsuladas em amastigotas de espécies de Leishmanias. Para isso os promastigotas foram incubados com curcumina livre divididos em grupo escuro e irradiado, além do grupo controle escuro e apenas irradiado. A viabilidade celular foi avaliada pelo teste de exclusão com azul de tripan, a atividade mitocondrial por MTT, as alterações morfológicas por MEV. Para analisar o tipo de morte celular utulizou-se os marcadores Anexina V e iodeto de propídio e para avaliar o potencial de membrana mitocondrial o JC-1, para isso foram obtidas as imagens por microscopia confocal e a fluorescência emitida pelos marcadores foram quantificadas utilizando o software ImageJ. Para analisar a produção de espécies reativas de oxigênio utilizou-se o reagente H2DCF-DA e a fragmentação de DNA foi verificada pelo teste do cometa. Verificou-se a diminuição da viabilidade dos parasitas bem como alterações na atividade mitocondrial. A TFD provocou alterações no potencial de membrana mitocondrial das duas espécies, independentemente do tempo de análise (imediatamente ou 18 horas após tratamento). A produção de ERO foi concentraçãodependente e foi observada apenas nos grupos submetidos à TFD e a espécie L. braziliensis apresentou maior produção de ERO que L. major, no entanto L. braziliensis foi mais resistente que L. major quando analisada a viabilidade após TFD. Os danos no DNA também foram observados nas duas espécies, é sugestivo que a apoptose prevaleceu na espécie L. braziliensis. A morfologia das células foi afetada, principalmente na concentração mais elevada de curcumina, deixando as células arredondadas e flagelo encurtado. Os resultados apoiam o uso da curcumina como fotossensibilizador na TFD contra células de Leishmania. Foram testadas também nanopartículas de BSA associada a curcumina em amastigotas de L. braziliensis e L. major. Para isso macrófagos e macrófagos infectados com Leishmanias foram incubados com a curcumina encapsulada em BSA catiônica e aniônica. Foram realizados testes de viabilidade pelo método de exclusão com azul de tripan, onde foi verificado que apenas a concentração 0,7 µmol. L1 da BSA-Cur catiônica demonstrou redução na viabilidade das células e as concentrações de 0,1 e 0,08 µmol. L-1 da BSA-Cur aniônica reduziu a viabilidade das células, já a concentração de 0,7 µmol. L-1 foi citotóxica. A análise através da microscopia confocal foi realizada para verificar se as nanopartículas eram internalizadas pelos macrófagos e pode-se verificar que as nanopartículas catiônicas e aniônicas se encontravam dentro dos macrófagos e protozoários. Por fim foi realizado teste de viabilidade com os fibroblastos e verificou-se que as nanopartículas não afetaram a viabilidade dessa linhagem celular, no entanto a curcumina livre promoveu diminuição da viabilidade dos fibroblastos após tratamento. Os resultados obtidos foram satisfatórios, demonstrando a capacidade das nanopartículas reduzirem a viabilidade das células em menores concentrações do que a curcumina livre.Item Effect of serial photodynamic therapy with curcumin on Leishmania braziliensis and Leishmania amazonensis promastigotes(CDRR Editors) Maciel, Lucas Tobias Rodrigues; Marcolino, Luciana Maria Cortez; Maciel, Fernanda Bueno Sant’Anna Pereira; Pinto, Juliana Guerra; Ferreira-Strixino, JulianaPhotodynamic Therapy (PDT) consists of using a light source and a photosensitive drug at an appropriate wavelength and molecular oxygen to trigger cell death through the production of reactive oxygen species. Because it is a localised therapy, PDT is shown to be ideal for skin diseases. American cutaneous Leishmaniasis (ACL) is a highly prevalent protozoan disease worldwide that presents different clinical evolutions and may result in ulcerations and disfiguring lesions on the skin and cartilage. This study was aimed at evaluating the effect in vitro of PDT applied serially using curcumin as a photosensitiser. For this, a concentration of 125 µg.mL-1 of curcumin was used on Leishmania braziliensis and Leishmania amazonensis strains, with a light fluence of 10 J.cm-2 and irradiance of 110 mW.cm-2. The tests done were viability analysis by trypan blue exclusion test, analysis of photosensitizer (PS) internalization by confocal microscopy and morphological alterations by May-Grunwald/Giemsa staining. We observed that there was internalisation of the PS before the first and second application of PDT, with L. braziliensis and L. amazonensis strains mortality of 92% and 82% respectively, after the second application, and induction of alterations in the structural conformation, such as cell size and non-evidence of nucleus and flagellum, demonstrating that PDT was effective. We conclude that serial PDT was effective in inducing the mortality of promastigotes forms of L. braziliensis and L. amazonensis in vitro, thus highlighting its potential for the treatment of leishmaniasis.Item Evaluation of the Photodynamic Therapy with Curcumin on L. braziliensis and L. major Amastigotes(MDPI) Pereira, André Henrique Correia; Marcolino, Luciana Maria Cortez; Pinto, Juliana Guerra; Strixino, Juliana FerreiraCutaneous leishmaniasis (CL) is a neglected disease prevalent in tropical countries with the ability to cause skin lesions. Photodynamic therapy (PDT) represents a specific and topical option for the treatment of these lesions. This study evaluated the response of macrophages infected with L. braziliensis and L. major to PDT with curcumin. Curcumin concentrations were evaluated in serial dilutions from 500.0 to 7.8 μg/mL using LED (λ = 450 ± 5 nm), with a light dose of 10 J/cm2. The Trypan blue viability test, ultrastructural analysis by scanning electron microscopy (SEM), mitochondrial polarity by Rhodamine 123 (Rho 123), curcumin internalization by confocal microscopy, and counting of the recovered parasites after the PDT treatment were performed. The lowest concentrations of curcumin (15.6 and 7.8 μg/mL) presented photodynamic inactivation. Cell destruction and internalization of curcumin in both macrophages and intracellular parasites were observed in microscopy techniques. In addition, an increase in mitochondrial membrane polarity and a decrease in the number of parasites recovered was observed in the PDT groups. This study indicates that PDT with curcumin has the potential to inactivate infected macrophages and might act as a basis for future in vivo studies using the parameters herein discussed.Item Molecular effects of photodynamic therapy with curcumin on Leishmania major promastigotes(Parasitology Research, Springer-Verlag GmbH Germany) Marcolino, Luciana Maria Cortez; Pinto, Juliana Guerra; Ferreira, Isabelle; Godoi, Bruno Henrique; Canevari, Renata de Azevedo; Ferreira-Strixino, JulianaLeishmaniasis is a neglected disease mainly affecting low-income populations. Conventional treatment involves several side effects, is expensive, and, in addition, protozoa can develop resistance. Photodynamic therapy (PDT) is a promising alternative in treating the disease. PDT involves applying light at a specific wavelength to activate a photosensitive compound (photosensitizer, PS), to produce reactive oxygen species (ROS). Curcumin and its photochemical characteristics make it a good candidate for photodynamic therapy. Studies evaluating gene expression can help to understand the molecular events involved in the cell death caused by PDT. In the present study, RNA was extracted from promastigotes from the control and treated groups after applying PDT. RT-qPCR was performed to verify the expression of the putative ATPase beta subunit (ATPS), ATP synthase subunit A (F0F1), argininosuccinate synthase 1 (ASS), ATP-binding cassette subfamily G member 2 (ABCG2), glycoprotein 63 (GP63), superoxide dismutase (FeSODA), and glucose-6-phosphate dehydrogenase (G6PDH) genes (QR). The results suggest that PDT altered the expression of genes that participate in oxidative stress and cell death pathways, such as ATPS, FeSODA, and G6PD. The ATP-F0F1, ASS, and GP63 genes did not have their expression altered. However, it is essential to highlight that other genes may be involved in the molecular mechanisms of oxidative stress and, consequently, in the death of parasites.Item Photodynamic therapy of cationic and anionic BSA-curcumin nanoparticles on amastigotes of Leishmania braziliensis and Leishmania major and Leishmania amazonensis(Photodiagnosis and Photodynamic Therapy, Elsevier) Marcolino, Luciana Maria Cortez; Ambrósio, Jéssica Aparecida Ribeiro; Pinto, Juliana Guerra; Ferreira, Isabelle; Simioni, Andreza Ribeiro; Ferreira-Strixino, JulianaCutaneous leishmaniasis is a neglected disease prevalent in tropical countries, and conventional treatment can cause several serious side effects. Photodynamic therapy (PDT) can be considered a promising treatment alternative, as it is non-invasive therapy that has no side effects and uses accessible and low-cost substances, such as curcumin. This study evaluated the PDT response with cationic and anionic BSA nanoparticles encapsulated with curcumin in macrophages infected with L. braziliensis, L. major, and L. amazonensis. The nanoparticle system was characterized using a steady-state technique, scanning electron microscopy (SEM) study, and its biological activity was evaluated using macrophage cell lines infected with different Leishmania species. All spectroscopy measurements demonstrated that BSA curcumin (BSACur) has good photophysical properties, and confocal microscopy shows that macrophages and protozoa internalized the nanoparticles. The viability test demonstrated that at low concentrations, such as 0.1, 0.7, and 1.0 μmol. L 1, there was a decrease in cell viability after PDT application. Furthermore, a decrease in the number of parasites recovered was observed in the PDT groups. The results allowed us to conclude that curcumin loaded into BSA nanoparticles may have potential application in drug delivery systems for PDT protocols, demonstrating reduced cell viability at lower concentrations than free curcumin.Item Zinc pthalocyanine loaded poly (lactic acid) nanoparticles by double emulsion methodology for photodynamic therapy against 9 L/LacZ gliosarcoma cells(Taylor & Francis Group) Oliveira Junior, Benedito Marcio de; Teodoro, Jéssica Beatriz Miranda; Ambrósio, Jéssica Aparecida Ribeiro; Gonçalves, Érika Peterson; Beltrame Junior, Milton; Marcolino, Luciana Maria Cortez; Pinto, Juliana Guerra; Ferreira-Strixino, Juliana; Simioni, Andreza RibeiroDevelopment delivery systems, such as nanoparticles, represent a growing area in biomedical research. Nanoparticles (NP) were prepared using a double-emulsion method to load zinc(II) phthalocyanine (ZnPc). NP were obtained using poly (lactic acid) (PLA). ZnPc is a second generation of photosensitizer used in photodynamic therapy (PDT). ZnPc loaded PLA nanoparticles (NPLAZnPc) were prepared by double-emulsion method, characterized and available in cellular culture. The mean nanoparticle size presented particle size was 384.7 ± 84.2 nm with polydispersity index (PDI) of 0.150 ± 0.015, and the encapsulation efficiency was of 83%. The nanoparticle formulations presented negative zeta potential values (27.5 ± 1.0 mV), explaining their colloidal stability. ZnPc loaded nanoparticles maintain its photophysical behavior after encapsulation. Photosensitizer release from nanoparticles was sustained over 168 h with a biphasic ZnPc release profile. An in vitro phototoxic effect in range of 80% was observed in 9 L/ LacZ gliosarcoma cells at laser light doses (10 J cm2) with 3.0 mg mL1 of NPLA-ZnPc. All the physical–chemical, photophysical and photobiological measurements performed allow us to conclude that ZnPc loaded PLGA nanoparticles is a promising drug deliverysystem for PDT.