Avaliação dos efeitos do azul de metileno sobre as arritmias cardíacas e letalidade em ratos normotensos e hipertensos submetidos à isquemia e reperfusão cardíaca
Data
2024-04-30
Autores
Araújo, Erisvaldo Amarante de 
Orientadores
Rodrigues, Francisco Sandro Menezes
Tipo
Dissertação de mestrado
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ISSN da Revista
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Resumo
Introdução: O azul de metileno (AM) tem sido proposto em alguns estudos como uma importante ferramenta farmacológica em experimentos de isquemia e reperfusão de diversos órgãos, como rins, fígado e pulmões, além de sua utilização no tratamento de doenças cardiovasculares como a Síndrome Vasoplégica, devido à sua atividade como bloqueador da Guanilato Ciclase (GC) ativada por Oxido Nítrico (NO) e adicional redução da produção de radicais livres, cujos mecanismos celulares e moleculares envolvidos nestes efeitos permanecem ainda sob investigação. Objetivos: Avaliar os efeitos do AM no coração de ratos das linhagens Wistar normotensos (NWR) e Wistar geneticamente hipertensos (SHR) submetidos à isquemia e reperfusão cardíaca (IRC) e também investigar, utilizando métodos computacionais, os possíveis alvos farmacológicos do AM em cardiomiócitos. Materiais e métodos: Ratos machos, NWR e SHR (12 a 16 semanas de idade), foram anestesiados e mantidos sob ventilação mecânica por um período de estabilização de 15 min, isquemia cardíaca (IC) por 10 min e reperfusão cardíaca (RC) por 75 min. A atividade cardíaca dos animais foi monitorada e registrada durante todo o protocolo experimental por eletrocardiograma (ECG), para avaliação das incidências de arritmias ventriculares (AV), bloqueio atrioventricular (BAV) e letalidade (LET). Amostras de sangue foram coletadas e processadas para a determinação da concentração sérica de marcadores bioquímicos de lesão cardíaca, creatina quinase fração MB (CK-MB) e troponina I (TnI). Os animais NWR e SHR foram distribuídos em 3 grupos experimentais: (1)Grupo SS+IRC: tratados com solução salina (SS) 0,9% administrada (i.v) antes da IRC; (2)Grupo AM+IRC: tratados com AM (2 mg/kg, i.v) antes da IRC; (3)Grupo AM+RC: tratados com AM (2 mg/kg, i.v) antes da RC. Para avaliar possíveis alvos farmacológicos do AM em cardiomiócitos foi utilizada a metodologia computacional de Docagem molecular (DMol). Resultados: Nos grupos de animais NWR, os tratados com SS antes da IRC apresentaram elevadas incidências de AV (90%), BAV (70%) e LET (60%). O tratamento com o AM antes da IRC causou um aumento significativo nas incidências de BAV (100%) e de LET (100%), enquanto o AM antes da reperfusão reduziu significativamente as incidências de AV (50%), BAV (28,5%) e LET (21,4%). Já em SHR, o grupo tratado com SS apresentou baixas incidências de AV (40%), BAV (30%) e LET (20%). A administração do AM antes da IRC aumentou significativamente as incidências de AV (100%), BAV (85,7%) e LET (78,5%), enquanto o grupo tratado com AM antes da RC apresentou incidências de AV (42,8%), BAV (28,5%) e LET (21,4%) similares ao controle. Não se observou diferenças importantes em relação às concentrações séricas dos marcadores de lesão cardíaca estudados em NWR e SHR, exceto os níveis de CK-MB dos animais NWR tratados com AM antes da IRC. A DMol apresentou ligações classificadas entre boa e ótima do AM para a Monoaminaoxidase-B (MAO-B), Calmodulina (CaM), Receptor Muscarínico M2 e Guanilato Ciclase (GC). Conclusão: Estes resultados mostraram que a administração do AM antes da RC reduziu ou preveniu o aumento das incidências de AV, BAV e LET decorrentes da IRC, tanto em NWR como em SHR. Em contraste, a administração do AM antes da IRC aumentou significativamente as incidências de AV, BAV e LET decorrentes da IRC, tanto em NWR como em SHR, sugerindo que este efeito cardiotóxico pode comprometer severamente a função cardíaca em resposta à IRC.
Introduction: Methylene blue (MB) has been proposed in some studies as an important pharmacological tool in ischemia and reperfusion experiments of various organs, such as kidneys, liver, and lungs, in addition to its use in the treatment of cardiovascular diseases such as Vasoplegic Syndrome, due to its activity as a blocker of Nitric Oxide (NO)-activated Guanylate Cyclase (GC) and additional reduction in the production of free radicals. The cellular and molecular mechanisms involved in these effects are still under investigation. Objective: To evaluate the effects of MB on the hearts of normotensive Wistar rats (NWR) and genetically hypertensive Wistar rats (SHR) subjected to cardiac ischemia and reperfusion (IRC), and also to investigate, using computational methods, the possible pharmacological targets of MB in cardiomyocytes. Materials and Methods: Male rats, NWR and SHR (12 to 16 weeks old), were anesthetized and kept under mechanical ventilation for a stabilization period of 15 minutes, cardiac ischemia (IC) for 10 minutes, and cardiac reperfusion (RC) for 75 minutes. The cardiac activity of the animals was monitored and recorded throughout the experimental protocol by electrocardiogram (ECG) to assess the incidence of ventricular arrhythmias (AV), atrioventricular block (BAV), and lethality (LET). Blood samples were collected and processed to determine the serum concentration of biochemical markers of cardiac injury, creatine kinase MB fraction (CK-MB), and troponin I (TnI). The NWR and SHR animals were distributed into 3 experimental groups: (1)SS+IRC group: treated with 0.9% saline solution (SS) administered (i.v) before IRC; (2)MB+IRC group: treated with MB (2 mg/kg, i.v) before IRC; (3)MB+RC group: treated with MB (2 mg/kg, i.v) before RC. To evaluate possible pharmacological targets of MB in cardiomyocytes, the computational method of molecular docking (DMol) was used. Results: In the NWR animal groups, those treated with SS before IRC showed high incidences of AV (90%), BAV (70%), and LET (60%). Treatment with MB before IRC caused a significant increase in the incidences of BAV (100%) and LET (100%), while MB before reperfusion significantly reduced the incidences of AV (50%), BAV (28.5%), and LET (21.4%). In SHR, the SS-treated group showed low incidences of AV (40%), BAV (30%), and LET (20%). Administration of MB before IRC significantly increased the incidences of AV (100%), BAV (85.7%), and LET (78.5%), while the group treated with MB before RC showed incidences of AV (42.8%), BAV (28.5%), and LET (21.4%) similar to the control. No significant differences were observed in the serum concentrations of the studied cardiac injury markers in NWR and SHR, except for the CK-MB levels in NWR animals treated with MB before IRC. DMol showed classified binding from good to excellent for MB with Monoamine Oxidase-B (MAO-B), Calmodulin (CaM), Muscarinic M2 Receptor (M2), and Guanylate Cyclase (GC). Conclusion: These results showed that the administration of MB before RC reduced or prevented the increase in the incidences of AV, BAV, and LET resulting from IRC in both NWR and SHR. In contrast, the administration of MB before IRC significantly increased the incidences of AV, BAV, and LET resulting from IRC in both NWR and SHR, suggesting that this cardiotoxic effect may severely compromise cardiac function in response to IRC.
Introduction: Methylene blue (MB) has been proposed in some studies as an important pharmacological tool in ischemia and reperfusion experiments of various organs, such as kidneys, liver, and lungs, in addition to its use in the treatment of cardiovascular diseases such as Vasoplegic Syndrome, due to its activity as a blocker of Nitric Oxide (NO)-activated Guanylate Cyclase (GC) and additional reduction in the production of free radicals. The cellular and molecular mechanisms involved in these effects are still under investigation. Objective: To evaluate the effects of MB on the hearts of normotensive Wistar rats (NWR) and genetically hypertensive Wistar rats (SHR) subjected to cardiac ischemia and reperfusion (IRC), and also to investigate, using computational methods, the possible pharmacological targets of MB in cardiomyocytes. Materials and Methods: Male rats, NWR and SHR (12 to 16 weeks old), were anesthetized and kept under mechanical ventilation for a stabilization period of 15 minutes, cardiac ischemia (IC) for 10 minutes, and cardiac reperfusion (RC) for 75 minutes. The cardiac activity of the animals was monitored and recorded throughout the experimental protocol by electrocardiogram (ECG) to assess the incidence of ventricular arrhythmias (AV), atrioventricular block (BAV), and lethality (LET). Blood samples were collected and processed to determine the serum concentration of biochemical markers of cardiac injury, creatine kinase MB fraction (CK-MB), and troponin I (TnI). The NWR and SHR animals were distributed into 3 experimental groups: (1)SS+IRC group: treated with 0.9% saline solution (SS) administered (i.v) before IRC; (2)MB+IRC group: treated with MB (2 mg/kg, i.v) before IRC; (3)MB+RC group: treated with MB (2 mg/kg, i.v) before RC. To evaluate possible pharmacological targets of MB in cardiomyocytes, the computational method of molecular docking (DMol) was used. Results: In the NWR animal groups, those treated with SS before IRC showed high incidences of AV (90%), BAV (70%), and LET (60%). Treatment with MB before IRC caused a significant increase in the incidences of BAV (100%) and LET (100%), while MB before reperfusion significantly reduced the incidences of AV (50%), BAV (28.5%), and LET (21.4%). In SHR, the SS-treated group showed low incidences of AV (40%), BAV (30%), and LET (20%). Administration of MB before IRC significantly increased the incidences of AV (100%), BAV (85.7%), and LET (78.5%), while the group treated with MB before RC showed incidences of AV (42.8%), BAV (28.5%), and LET (21.4%) similar to the control. No significant differences were observed in the serum concentrations of the studied cardiac injury markers in NWR and SHR, except for the CK-MB levels in NWR animals treated with MB before IRC. DMol showed classified binding from good to excellent for MB with Monoamine Oxidase-B (MAO-B), Calmodulin (CaM), Muscarinic M2 Receptor (M2), and Guanylate Cyclase (GC). Conclusion: These results showed that the administration of MB before RC reduced or prevented the increase in the incidences of AV, BAV, and LET resulting from IRC in both NWR and SHR. In contrast, the administration of MB before IRC significantly increased the incidences of AV, BAV, and LET resulting from IRC in both NWR and SHR, suggesting that this cardiotoxic effect may severely compromise cardiac function in response to IRC.
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Citação
ARAÚJO, Erisvaldo Amarante. Avaliação dos efeitos do azul de metileno sobre as arritmias cardíacas e letalidade em ratos normotensos e hipertensos submetidos à isquemia e reperfusão cardíaca. 2024. 104 f. Dissertação (Mestrado em Cardiologia) – Escola Paulista de Medicina, Universidade Federal de São Paulo, São Paulo, 2024