JANAINA ALINE GALVÃO BARROS

JANAINA ALINE GALVÃO BARROS

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Protein crosslinking onto gold nanoparticles by gamma radiation
2017 - VARCA, GUSTAVO H.C.; BARROS, JANAINA A.G.; BATISTA, JORGE G.S.; SASOUNIAN, RAFAELA; SILVA, GUSTAVO T.M.; ZAMARION, VITOR M.; KATTI, KATTESH V.; LUGAO, ADEMAR B.
The use of gold nanoparticles for diagnosis and treatment of cancer has received great attention over the last decade. Particularly, the possibility to use them for theranostics has increased the interest in the medical and scientific community. Weak technological aspects are related to the low biological affinity and non-specific toxicity. The use of albumin is of highlighted interest as albumin has been associated to inorganic particles to overcome biopharmaceutical challenges, including site-specific delivery and other biopharmaceutical advantages. The current work addresses the use of radiation and its effects over the crosslinking of bovine serum albumin onto gold nanoparticles. The idea of crosslinking the albumin onto gold surface aims to improve the stability of the protein layer onto gold nanoparticles in biological systems. Gold nanoparticles were synthesized by green technology using resveratrol and albumin capping was performed by physiosorption followed by irradiation at doses of 2.5, 5, 7.5, 10 and 15 kGy using 60Co as a radioactive source. Nanoparticle properties were assessed by dynamic light scattering, UV/Vis spectrophotometry and transmission electron microscopy. Protein crosslinking was monitored by fluorescence studies and stability of the nanoparticles was evaluated by zeta potential and titration with sodium chloride. The results evidenced the formation of a protein layer onto gold nanoparticles and revealed a protein crosslinking by means of bityrosine as a function of irradiation dose. Stability was considerably improved by the presence of the protein layer and the crosslinked protein layer.
Avaliação da toxicidade in vivo das Nanopartículas de Ouro reduzidas e estabilizadas com o fitoquímico Resveratrol
2018 - CAVALCANTE, A.K.; BATISTA, J.G.S.; BARROS, J.A.G.; ORMENIO, M.B.; DAMASCENO, K.C.; ROGERO, S.O.; ROGERO, J.R.; LUGAO, A.B.
As nanopartículas de ouro (AuNPs) com diferentes tamanhos e formas têm sido amplamente estudadas e utilizadas em diversas áreas, como por exemplo, em aplicações biomédicas. Dentre tais aplicações, encontramos a liberação de agentes antitumorais. A síntese de AuNPs geralmente envolve agentes de redução que apresentam problemas relacionados à toxicidade. A fim de resolver esta questão, metabólitos presentes em diversos extratos de plantas tem sido explorados para a preparação de diferentes nanopartículas. Os métodos que utilizam os fitoquímicos para redução de íons metálicos fornecem uma abordagem verde a nanotecnologia, conhecida como “green nanotechnology”. O fitoquímico resveratrol, um composto fenólico com potencial redutor, encontrado em 72 espécies de plantas, como uva, amora e amendoim, foi usado neste trabalho como agente redutor na preparação de AuNPs. O resveratrol além de ser um antioxidante, também é conhecido como fármaco antitumoral/anticâncer. Foi descrito na literatura, que a conjugação de Reveratrol com AuNPs aumenta em 65% a efetividade em testes realizados in vitro, utilizando células de câncer de pulmão humano, quando comparado ao resveratrol administrado isoladamente. Este trabalho teve como objetivo verificar o nível de toxicidade das nanopartículas de ouro, reduzidas e estabilizadas com resveratrol (RESV-AuNPs) em embriões de Zebrafish (Danio rerio), de acordo com o protocolo da OECD nº 236 (Fish Embryo Acute Toxicity Test- FET). Os embriões foram expostos as RESV-AuNPs por um período de 96 e 168 horas. O Zebrafish apresenta-se como um modelo in vivo alternativo, rápido, de alto rendimento, facilmente acessível e que possui uma boa correlação com modelos in vitro. As RESV-AuNPs demonstraram toxicidade nos dois períodos de exposição, sendo a letalidade dos organismos inferior a 10% em todas as concentrações utilizadas. O trabalho forneceu uma contribuição sobre a toxicidade de AuNPs sintetizadas e estabilizadas com o agente redutor resveratrol, utilizando como modelo animal embriões de Zebrafish.
Radiation-induced "one pot" synthesis for cell therapies
2017 - LUGAO, ADEMAR B.; FAZOLIN, GABRIELA N.; VARCA, GUSTAVO H.C.; SANTOS, JORGE G.; BARROS, JANAINA; FUCASE, TAMARA; SANTOS, JONNATAN J.; LEAL, JESSICA; GRASSELLI, MARIANO; KATTI, KATTESH V.
The dream of Marie Curie lab’s expressed by Regaud and Lacassagne in 1927 was to administer radiations with penetration range of molecular dimensions to the organism and selectively fixed in the protoplasm of cells one seeks to destroy. Gold nanoparticles can be employed as a radiation sensitizer by utilizing mainly Auger effect and photoelectrons. Auger electrons are released in large numbers with low kinetic energy therefore these electrons damage cells over a very short range: less than the size of a single cell, on the order of nanometers. Gold-198 is a beta and gamma emitter can be employed for therapy as well as diagnostic. The radioactive properties of gold include: Au (βmax=0.96 MeV; t1/2 = 2.7 days) and Au (βmax =0.46 MeV; t1/2 = 3.14 days), making it a strong candidate for theranostics. However, Gold or Gold-198 need to internalize selectively in tumor cells. Conjugation with proteins and peptides can make them very selective. While radioactive nanoparticles can offer a much higher dose payload than ions for therapy and diagnostic, in addition to the huge surface to bind targeting species presented by the nanoparticles, functionalization with proteins may potentially increases the particle uptake by tumors or tissues. Albumin and Papain features a set of characteristics that assure applications as natural drug carriers with particular attractive properties in oncology. Albumin may be easily crosslinked and engineered towards loading of large number of hydrophobic molecules as well as hydrophilic ones. They can be bound in a reversible way and the delivery controlled by endogenous mechanism. Alternatively to conventional systems, albumin can be crosslinked by radiation in such way that dialdehydes or toxic chemicals are totally avoided . Conjugation of such materials with sugars, peptides, antibodies, proteins among others is routinely used nowadays for targeting. The main purpose of this work was the development of one pot in situ synthesis of radioactive gold 198 nanoparticle encapsulated by albumin for application in cancer Theranostics. While crosslinked albumin may provide a nontoxic coating on AuNPs with a controllable hydrodynamic diameter, conventional AuNP can be activated by nuclear reactor to produce AuNP. The gamma or beta radiation originated from the gold nanoparticle was used to crosslink the Albumin layer. The use of a radioactive particle able to emit radiation for crosslinking of the Albumin layer and simultaneous theranostic application was tried for the first time. The elegant procedure and simplicity of the production process combined with the properties of Au and the safety of AuNP/BSA make this new particle an exciting advancement in cancer therapy and diagnosis. Gold conjugated protein nanoparticles and protein nanoparticles itself were also produced in an radiation induced one pot process. Crosslinking and protein damage werea cessed by different techniques.
Evaluation of the in vitro and in vivo toxicity of gold nanoparticles synthesized by green nanotechnology
2017 - BATISTA, JORGE G.S.; BARROS, JANAINA A.G.; VARCA, GUSTAVO H.C.; ROGERO, SIZUE O.; CAVALCANTE, ADRIANA K.; MAZIERO, JOANA S.; ROGERO, JOSE R.; LUGAO, ADEMAR B.
Researchers and laboratories around the world have studied gold nanoparticles. In medicine, several studies demonstrate the applicability of gold nanoparticles (AuNPs) in the treatment and diagnosis of cancer. Green nanotechnology uses phytochemical agents to synthesize and stabilize nanoparticles. The phytochemical epigallocatechin-gallate (EGCG) reduces and stabilize gold nanoparticles by functionalizing the surface of the molecule. Such chemical groups allow binding to overexpressed receptors on some types of tumors as demonstrated in studies performed with PC3 prostate cancer cells. With the advancement of nanotechnology, a large number of nanoparticles are produced on a daily basis. However apart from their possible applications it is necessary to evaluate the environmental impact of these molecules as well as find ways for proper disposal. The embryonic zebrafish (Danio rerio) trial has recently emerged as an interesting method for evaluating in vivo nanotoxicity providing a more complex system analysis than in typical cell cultures and less expensive if compared to large-scale biocompatibility studies performed in rats and mice. The objective of this study was to evaluate the in vitro and in vivo toxicity of EGCG-AuNPs by means of the cytotoxicity by neutral red uptake methodology according to the International Standard Organization [ISO 10993-5, 2009] and in vivo test based on the OECD guideline on Fish Embryo Toxicity Test (FET) (OECD, 2013). The spectrophotometric band at 535 nm observed is characteristic of the formation of AuNPs. Nanoparticles synthesized with EGCG presented a size of 32 ± 4 nm as determined by transmission electron microscopy and the hydrodynamic diameter of these particles was about 60 ± 18 nm obtained by dynamic light scattering. The EGCG-AuNPs showed no cytotoxicity up to 4.2 μg.L-1. In the FET test regarding the acute ecotoxicity assay the LC50/96 hours revealed no toxicity at concentrations up to 1.8 mM.
Synthesis of radioactive gold 198 nanoparticle encapsulated by albumin as cancer theranostics agent
2016 - LUGAO, A.B.; GERALDES, A.N.; LEAL, J.; VARCA, G.H.C.; BATISTA, J.G.S.; GRASSELLI, M.; KATTI, K.; BARROS, J.
Albumin is a natural drug nanocarrier as it has equivalent diameter of about 5 nm and has natural affinity for hydrophobic and hydrophilic drugs. They can be bound in a reversible way and the delivery controlled by endogenous mechanism. Albumin can be crosslinked by radiation alternatively to conventional systems, in such way that dialdehydes or toxic chemicals are avoided. On the theranostic side, radioactive ions are commonly employed for diagnostic and therapeutic applications. As an example, radioactive gold nanoparticles are currently employed in radiotherapy whether to increase local dose deposition in tissue during radiotherapy or as a local emitter of gamma and beta rays. The radioactive properties of gold include: 198Au (?max=0.96 MeV; t1/2 = 2.7 days) and 199Au (?max =0.46 MeV; t1/2 = 3.14 days), making it a strong candidate for theranostics . Conjugation of such materials with sugars, peptides, antibodies, proteins among others is routinely used nowadays for targeting. While radioactive nanoparticles can offer a much higher dose payload than ions for therapy and diagnostic, in addition to the the huge surface to bind targeting species presented by the nanoparticles, functionalization with proteins may potentially increases the particle uptake by tumors or tissues. The main purpose of this work was the development of one pot in situ synthesis of radioactive gold 198 nanoparticle encapsulated by albumin for application in cancer Theranostics. While crosslinked albumin may provide a nontoxic coating on AuNPs with a controllable hydrodynamic diameter, conventional AuNP can be activated by nuclear reactor to produce 198AuNP. The gamma or beta radiation originated from the gold nanoparticle was used to crosslink the Albumin layer. The use of a radioactive particle able to emit radiation for crosslinking of the Albumin layer and simultaneous theranostic application was tried for the first time. The elegant procedure and ease of produc¬tion combined with the properties of 198Au and the safety of HSA-198AuNP make this new particle an exciting advancement in cancer therapy and diagnosis. For such purpose, radioactive tetrachloroauric acid H198AuCl4 was produced from gold foils of high purity by neutron irradiation in IPEN research nuclear reactor. 198AuNP/BSA were synthesized by stirring aqueous solutions of BSA and radioactive tetrachloroauric acid H198AuCl4. The reaction mixtures were stirred continuously at 25 °C. The color of the mixture become purple-red from pale yellow within 15 minutes indicating the formation of gold nanoparticles. The reaction mixture was stirred for an additional 20 minutes. The 198AuNP/BSA formed were characterized by UV-Vis spectrophotometry, DLS and TEM analysis. The spectra were recorded at first day of preparation and after 1 month. Radioactive gold nanoparticle encapsulated by crosslinked Albumin was prepared in reproducible way. The gold nanoparticle core size measured by TEM was about 20 nm and about 60 to 70 nm with the albumin layer as measured by DLS. Bityrosine formation was measured by fluorescence and it was an evidence of intramolecular and intermolecular crosslinking. In conclusion the technique was suitable for the in situ production of the crosslinked albumin functionalized radioactive gold nanoparticles. Acknowledgements The authors would like to thank Conselho Nacional de Desenvolvimento Cientifico e Tecnológico (CNPq) project number 402887/2013-1 and 401438/2014-7 and International Atomic Energy Agency (IAEA) (CRP code F22064 ) for financial support.
Radiation crosslinked albumin capped gold nanoparticles for theranostics
2016 - VARCA, GUSTAVO H.C.; BARROS, JANAINA A.G.; BATISTA, JORGE G. dos S.; KATTI, KATTESH V.; LUGAO, ADEMAR B.