RESUMO
Resumo Fundamento A formação de células espumosas ocorre devido ao aumento em lipoproteína plasmática de baixa densidade (LDL) e desregulação da inflamação, sendo importante para o desenvolvimento da aterosclerose. Objetivo Avaliar o perfil do fator de necrose tumoral alfa (TNF-α) e da interleucina-6 (IL-6) no método de formação da célula espumosa existente, otimizando esse protocolo. Métodos A LDL foi isolada, oxidada e marcada com sonda de isotiocianato de fluoresceína (FITC). As células espumosas foram geradas de célula derivada de monócitos humanos THP-1 e incubadas na ausência (controle) ou presença de FITC-ox-LDL (10, 50, 100, 150 ou 200 μg/mL), por 12, 24, 48 ou 72 horas. A FITC-ox-LDL na célula foi quantificada por microscopia. O ensaio de imunoabsorção enzimática foi avaliado para quantificar a IL-6 e o TNF-α, com um p <0,05 considerado significativo. Resultados Todas as concentrações de FITC-ox-LDL testadas apresentaram fluorescência mais alta em comparação com o controle, demonstrando maior acúmulo de lipoproteínas nas células. Quanto mais alta a concentração de FITC-ox-LDL, maior a produção de TNF-α e IL-6. A produção de IL-6 pelas células espumosas foi detectada até o valor de 150 µg/mL da LDL máxima de estímulo. Concentrações acima de 50 μg/mL de LDL estimularam maior liberação de TNF-α comparado ao controle. Conclusões Nosso modelo contribui para o entendimento da liberação de IL-6 e TNF-α em resposta a várias concentrações de ox-LDL usando o método otimizado para a formação de células espumosas.
Abstract Background The formation of foam cells occurs due to the increase in low-density plasma lipoprotein (LDL) and dysregulation of inflammation, which is important for the development of atherosclerosis. Objective To evaluate the profile of tumor necrosis factor-alpha (TNF-α) and Interleukin-6 (IL-6) in the existing foam cell formation method, optimizing this protocol. Methods The LDL was isolated, oxidized, and labeled with a Fluorescein isothiocyanate (FITC) probe. Foam cells were generated from THP-1 human monocyte-derived cells and incubated in the absence (control) or presence of FITC-ox-LDL (10, 50, 100, 150, or 200 μg/mL), for 12, 24, 48, or 72 hours. The accumulated FITC-ox-LDL in the cell was quantified by microscopy. The enzyme-linked immunosorbent assay was evaluated to quantify the IL-6 and TNF-α, with p < 0.05 considered significant. Results All the FITC-ox-LDL concentrations tested showed a higher fluorescence when compared to the control, showing a greater accumulation of lipoprotein in cells. The higher the concentration of FITC-ox-LDL, the greater the production of TNF-α and IL-6. The production of IL-6 by foam cells was detected up to the value of 150 µg/mL of the maximum stimulus for LDL. Concentrations above 50 μg/mL LDL stimulated greater release of TNF-α compared to control. Conclusions Our model contributes to the understanding of the release of IL-6 and TNF-α in response to different concentrations of ox-LDL, using an optimized method for the formation of foam cells.
RESUMO
BACKGROUND: The formation of foam cells occurs due to the increase in low-density plasma lipoprotein (LDL) and dysregulation of inflammation, which is important for the development of atherosclerosis. OBJECTIVE: To evaluate the profile of tumor necrosis factor-alpha (TNF-α) and Interleukin-6 (IL-6) in the existing foam cell formation method, optimizing this protocol. METHODS: The LDL was isolated, oxidized, and labeled with a Fluorescein isothiocyanate (FITC) probe. Foam cells were generated from THP-1 human monocyte-derived cells and incubated in the absence (control) or presence of FITC-ox-LDL (10, 50, 100, 150, or 200 µg/mL), for 12, 24, 48, or 72 hours. The accumulated FITC-ox-LDL in the cell was quantified by microscopy. The enzyme-linked immunosorbent assay was evaluated to quantify the IL-6 and TNF-α, with p < 0.05 considered significant. RESULTS: All the FITC-ox-LDL concentrations tested showed a higher fluorescence when compared to the control, showing a greater accumulation of lipoprotein in cells. The higher the concentration of FITC-ox-LDL, the greater the production of TNF-α and IL-6. The production of IL-6 by foam cells was detected up to the value of 150 µg/mL of the maximum stimulus for LDL. Concentrations above 50 µg/mL LDL stimulated greater release of TNF-α compared to control. CONCLUSIONS: Our model contributes to the understanding of the release of IL-6 and TNF-α in response to different concentrations of ox-LDL, using an optimized method for the formation of foam cells.
FUNDAMENTO: A formação de células espumosas ocorre devido ao aumento em lipoproteína plasmática de baixa densidade (LDL) e desregulação da inflamação, sendo importante para o desenvolvimento da aterosclerose. OBJETIVO: Avaliar o perfil do fator de necrose tumoral alfa (TNF-α) e da interleucina-6 (IL-6) no método de formação da célula espumosa existente, otimizando esse protocolo. MÉTODOS: A LDL foi isolada, oxidada e marcada com sonda de isotiocianato de fluoresceína (FITC). As células espumosas foram geradas de célula derivada de monócitos humanos THP-1 e incubadas na ausência (controle) ou presença de FITC-ox-LDL (10, 50, 100, 150 ou 200 µg/mL), por 12, 24, 48 ou 72 horas. A FITC-ox-LDL na célula foi quantificada por microscopia. O ensaio de imunoabsorção enzimática foi avaliado para quantificar a IL-6 e o TNF-α, com um p <0,05 considerado significativo. RESULTADOS: Todas as concentrações de FITC-ox-LDL testadas apresentaram fluorescência mais alta em comparação com o controle, demonstrando maior acúmulo de lipoproteínas nas células. Quanto mais alta a concentração de FITC-ox-LDL, maior a produção de TNF-α e IL-6. A produção de IL-6 pelas células espumosas foi detectada até o valor de 150 µg/mL da LDL máxima de estímulo. Concentrações acima de 50 µg/mL de LDL estimularam maior liberação de TNF-α comparado ao controle. CONCLUSÕES: Nosso modelo contribui para o entendimento da liberação de IL-6 e TNF-α em resposta a várias concentrações de ox-LDL usando o método otimizado para a formação de células espumosas.
Assuntos
Aterosclerose , Células Espumosas , Humanos , Interleucina-6 , Fluoresceína , Fator de Necrose Tumoral alfa , Fluoresceína-5-Isotiocianato , Lipoproteínas LDL , IsotiocianatosRESUMO
Exercise can generate alterations in body composition and modulate the immune system. The objective of this study was to verify whether a circuit resistance training (CRT) protocol can increase lean body mass (LM), and reduce fat body mass (FM) and the percent of FM (%FM) of sedentary women, without inducing inflammatory responses, indicated by serum cytokine levels. The initial hypothesis was that CRT would improve body composition, without changing serum cytokine levels. The study consisted of 14 healthy, sedentary women, aged 33-45 years (mean +/- SD, 40.23 +/- 3.98 years), with a normal body mass index. They participated in 3 sessions per week of CRT, which included 2 rounds in 9 stations with 1 set of 8-12 repetition maximum at each station, for 10 weeks. During the 10-week CRT period, participants maintained their pretraining nutritional standard. Body composition was analysed with dual-energy X-ray absorptiometry both pre- and post-training. Blood samples were collected after 96 h of rest pre- and post-training, and 5 min, 24 h, and 48 h after the second and last training sessions to measure serum cytokine levels by flow cytometry. The nutritional standard was accompanied throughout the study period with 24-h dietary recall. Increases in LM (35.937 +/- 4.926 to 39.130 +/- 4.950 kg) and decreases in FM (21.911 +/- 8.150 to 17.824 +/- 4.235 kg) and %FM (37.10 +/- 10.84 to 31.19 +/- 6.06), without concurrent changes in serum cytokine levels, and in the nutritional standard (alpha = 0.05). The proposed CRT improved body composition and did not induce any changes in serum cytokine levels characteristic of the inflammatory response in women.
