Avaliação citotóxica de nanopartículas magnéticas utilizando fotohipertermia no tratamento de melanoma
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2019-03-23
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Universidade Federal de Goiás
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Justification: Nanophototherapy has been shown a promising technique in non-invasive treatment of high selectivity with active uptake to the tumor target, showing good intracellular hyperthermic heat delivery efficiency, which promotes therapeutic effects such as tumor regression and increased expression of Heat Shock Proteins (HSPs). These proteins have also been shown to trigger systemic processes such as thermal immunoactivation and abscopal effect. It has been invested in the search for new nanoformulations that present the characteristics of biocompatibility required for nanophotohyperthermic applications. In this context, a study was conducted on the cytotoxic and antitumor profile in order to contribute to the development of new drugs, which are more efficient and safer for cancer treatment. Objectives: To evaluate cell viability, cytotoxic effect, mechanism of death after intracellular thermal treatment of nanophotohyperthermia with MALB magnetic nanoparticle (BSA+MnFe2O4) on murine melanoma tumor cells (B16-F10) and normal murine fibroblast cell line (L-929). Methodology: Cells were cultured in 6-well plates with DMEM containing 10% FBS, 1% of penicillin, in a humidified incubator at 37°C with 5% CO2. It were plated 5x105 cells per well after 3h initiated incubation with MALB for 12h or 24h at a final concentration of 914μg.mL-1 (proportion of 547 cells per ug of MALB). After the endocytosis time, the cells were washed twice in PBS and centrifuged obtaining pellets to receive nanophotohyperthermia treatment for 30min (ʎ=808nm, potency=4-6W/cm2). The heating was captured by infrared thermal camera images. After intracellular heat treatment at 42.5 or 46°C were verified the cytotoxicity, cell viability, the mechanism of death and ability to form new colonies. Statistical analysis of ANOVA was done using GraphPad Prism 5. Results: MTT assay results non-heating promoted by nanophotohyperthermia make it clear that MALB and MnFe2O4 coated with sodium citrate do not present cytotoxicity for both lineages. The MALB is selective to the melanoma tumor line. Evidence was found that MALB has active uptake to the tumor target, due to albumin mounted on the nanocarrier. It was observed that MALB [914μg.mL-1] previously incubated for 12 or 24h more laser application has led tumor cells B16-F10 to high intracellular hyperthermia temperatures range (42.5 and 46°C) resulting in 79% of death by late apoptosis (annexin V/+ and PI/+), due to thermal necrosis and only 17% of living cells. Through iron mass revealed by Ferene-S assay has been found that MALB 24h incubation was endocytosed 10.67 ± 0,5 pg Fe+/cell (B16-F10), the equivalent 3.5x higher than control (-) and still twice greater than for L-929. Conclusion: MALB showed good intracellular heat delivery efficiency in a dose-dependent manner. The nanophotohyperthermia heating tests presented good results of destruction of micro-phantom tumors for melanoma model in vitro (B16-F10) and still showed high selectivity to the tumoral lineage, due to the presence of albumin (BSA) in its nanostructure, revealing that this nanocarrier has active tumor targeted characteristic
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OLIVEIRA, A. L. S. Avaliação citotóxica de nanopartículas magnéticas utilizando fotohipertermia no tratamento de melanoma. 2019. 93 f. Dissertação (Mestrado em Ciências Biológicas) - Universidade Federal de Goiás, Goiânia, 2019.