Efeito do tratamento com eugenol e deidrodieugenol em modelo experimental de enfisema pulmonar induzido por elastase
Data
2022-10-24
Autores
Medeiros, Laura Taguchi Perez [UNIFESP]
Taguchi, Laura [UNIFESP]
Orientadores
Prado, Carla Máximo [UNIFESP]
Tipo
Tese de doutorado
Título da Revista
ISSN da Revista
Título de Volume
Resumo
A Doença Pulmonar Obstrutiva Crônica é uma doença de alta prevalência, morbidade e mortalidade, caracterizada por uma inflamação pulmonar crônica difusa. Nos períodos de exacerbação há agravamento da obstrução do fluxo aéreo e remodelamento pulmonar favorecendo redução progressiva da função pulmonar e piora na qualidade de vida. Nenhum tratamento convencional não reduz a progressão da doença e desencadeia efeitos colaterais. Os compostos foram extraídos de folhas secas de Nectandra leucanta: eugenol (4-alil-2-metoxifenol) e deidrodieugenol ou biseugenol (3,3’-dimetoxi-5,5’-di-2-propenil-1,1’-bifenil-2,2’-diol). Ambas as substâncias apresentam propriedades terapêuticas anti-inflamatórias e antioxidantes com efeitos benéficos já comprovados. Objetivo. Avaliar os efeitos do tratamento com eugenol e biseugenol nas alterações pulmonares em modelo experimental de enfisema pulmonar induzida pela elastase. Materiais e Métodos. Inicialmente realizou-se curva para avaliação da capacidade antioxidante dos compostos in vitro. Em paralelo, camundongos C57BL/6 receberam elastase intranasal no dia 0. Os tratamentos com eugenol e o seu dímero biseugenol foram realizados por injeção intraperitoneal nos dias consecutivos 21 a 28 do protocolo. No 28º dia, duas horas após a última dose, foi avaliada a função pulmonar, coletado o fluido do lavado broncoalveolar, sangue periférico e medula óssea para avaliação de células inflamatórias. Os pulmões foram retirados, fixados em pressão constante e submetidos às técnicas histológicas habituais. Por análise morfométrica, foi quantificado no tecido pulmonar o diâmetro alveolar médio (coloração de H&E) e proporção de fibras colágenas (coloração de picrosírius). Também foi realizado imuno-histoquímica para análise de citocinas inflamatórias e estresse oxidativo. A análise estatística foi realizada pelo Programa GrapPadPrisma e um p<0,05 foi considerado significativo. Resultados. O biseugenol apresentou maior capacidade em potencial antioxidante do que o eugenol. A elastase aumentou o diâmetro alveolar médio, as células inflamatórias, e aquelas que expressam citoinas IL1β, IL-6 e IL-17, iNOS e fatores de transcrição (Nrf2 e NF-κB) no tecido pulmonar quando comparados aos animais controle. Os tratamentos com eugenol e biseugenol reduziram o diâmetro alveolar médio, o número de células positivas para Nrf2, NF-kB, citocinas e iNOS no pulmão. Não foram observadas diferenças entre os grupos experimentais na contagem de células no sangue periférico e na função pulmonar (Gtis, Htis, Ers, Rsr e Raw) nem na expressão de células positivas para SOD 1 e 2 no tecido pulmonar entre os grupos avaliados. Para análise dos dados utilizou-se análise de variância um fator e Student-Newman-Keuls, por meio do programa GraphPad Prisma, considerando p<0,05 significativo. Conclusão: Estes dados indicam que o eugenol e o biseugenol foram efetivos no controle do enfisema, da inflamação e iNOS no pulmão de camundongos, podendo assim serem ferramentas importantes e eficazes, aliados a terapias farmacológicas já existente para o tratamento do enfisema pulmonar.
Chronic Obstructive Pulmonary Disease is a disease of high prevalence, morbidity and mortality, characterized by a diffuse chronic lung inflammation. In periods of exacerbation, it causes irreversible airflow obstruction and lung remodeling, favoring a progressive reduction in lung function and worsening in the quality of life. No conventional treatment does not reduce the progression of the disease and have side effects. The compounds were extracted from dried leaves of Nectandra leucanta: eugenol, (4-allyl-2-methoxyphenol) and dehydrodieugenol or biseugenol (3,3'-dimethoxy-5,5'-di-2-propenyl-1,1'- biphenyl-2,2'-diol. Both substances have anti-inflammatory and antioxidant therapeutic properties with known beneficial effects. Aim. To evaluate the effects of eugenol and biseugenol treatment on lung changes in an experimental model of elastase-induced pulmonary emphysema. Materials and methods. First, we have made a dose response curve in vitro to evaluate the total antioxidant capacity of the two compounds. C57BL/6 mice received intranasal instillation of elastase on day 0. Treatment with eugenol and its dimer biseugenol was performed by intraperitoneal injection in consecutive days 21 to 28 of the protocol. On the 28th day, two hours after the last injection, it was evaluated lung mechanics, collected the bronchoalveolar lavage, peripheral blood and bone marrow to evaluate inflammatory cells. The lungs were removed, fixed in constant pressure and submitted to histological habitual techniques. By morphometric analyses, medium linear intercept was evaluated in lung parenchyma (H&E stain) and the collagen volume proportion (picrosirius stain). We also performed immunohistochemistry to measure cytokines and oxidative stress. Statistical analyzes were performed using the GraphPad Prism program and P<0.05 was considered significant. Results. Biseugenol showed greater antioxidant capacity compared to eugenol. Elastase induced an increase in medium linear intercept, in inflammatory cells and those positive to IL1β, IL-6, and IL-17, iNOS, and transcription factor (NF-κB and Nrf2), in the lung when compared to control animals. Eugenol and biseugenol treatments reduced medium linear intercept, and the positive cells to Nrf2, NFκB, cytokines and NOS in the lung. There were no differences among the experimental groups in peripheral blood cell counts and lung function (Gtis, Htis, Ers, Rsr and Raw) or in the expression of SOD 1 and 2 positive cells in lung tissue. For data analysis, one-factor analysis of variance and Student-Newman-Keuls analysis was used, using the GraphPad Prisma program, considering p<0.05 significant. Conclusion: These data indicate that eugenol and biseugenol were effective in controlling emphysema, inflammation and iNOS in the lung of mice, and these compounds may be important and effective tools, combined with existing pharmacological terapys for the treatment of lung emphysema.
Chronic Obstructive Pulmonary Disease is a disease of high prevalence, morbidity and mortality, characterized by a diffuse chronic lung inflammation. In periods of exacerbation, it causes irreversible airflow obstruction and lung remodeling, favoring a progressive reduction in lung function and worsening in the quality of life. No conventional treatment does not reduce the progression of the disease and have side effects. The compounds were extracted from dried leaves of Nectandra leucanta: eugenol, (4-allyl-2-methoxyphenol) and dehydrodieugenol or biseugenol (3,3'-dimethoxy-5,5'-di-2-propenyl-1,1'- biphenyl-2,2'-diol. Both substances have anti-inflammatory and antioxidant therapeutic properties with known beneficial effects. Aim. To evaluate the effects of eugenol and biseugenol treatment on lung changes in an experimental model of elastase-induced pulmonary emphysema. Materials and methods. First, we have made a dose response curve in vitro to evaluate the total antioxidant capacity of the two compounds. C57BL/6 mice received intranasal instillation of elastase on day 0. Treatment with eugenol and its dimer biseugenol was performed by intraperitoneal injection in consecutive days 21 to 28 of the protocol. On the 28th day, two hours after the last injection, it was evaluated lung mechanics, collected the bronchoalveolar lavage, peripheral blood and bone marrow to evaluate inflammatory cells. The lungs were removed, fixed in constant pressure and submitted to histological habitual techniques. By morphometric analyses, medium linear intercept was evaluated in lung parenchyma (H&E stain) and the collagen volume proportion (picrosirius stain). We also performed immunohistochemistry to measure cytokines and oxidative stress. Statistical analyzes were performed using the GraphPad Prism program and P<0.05 was considered significant. Results. Biseugenol showed greater antioxidant capacity compared to eugenol. Elastase induced an increase in medium linear intercept, in inflammatory cells and those positive to IL1β, IL-6, and IL-17, iNOS, and transcription factor (NF-κB and Nrf2), in the lung when compared to control animals. Eugenol and biseugenol treatments reduced medium linear intercept, and the positive cells to Nrf2, NFκB, cytokines and NOS in the lung. There were no differences among the experimental groups in peripheral blood cell counts and lung function (Gtis, Htis, Ers, Rsr and Raw) or in the expression of SOD 1 and 2 positive cells in lung tissue. For data analysis, one-factor analysis of variance and Student-Newman-Keuls analysis was used, using the GraphPad Prisma program, considering p<0.05 significant. Conclusion: These data indicate that eugenol and biseugenol were effective in controlling emphysema, inflammation and iNOS in the lung of mice, and these compounds may be important and effective tools, combined with existing pharmacological terapys for the treatment of lung emphysema.
Descrição
Citação
TAGUCHI, Laura. Efeito do tratamento com eugenol e deidrodieugenol em modelo experimental de enfisema pulmonar induzido por elastase. 2022. 77 f. Tese (Doutorado Interdisciplinar em Ciências da Saúde) - Instituto de Saúde e Sociedade, Universidade Federal de São Paulo, Santos, 2022.