Methylene blue for treatment of hospitalized Covid-19 patients: a randomized, controlled, open-label clinical trial, phase 2

ABSTRACT Background: There is no pharmacological intervention on the treatment of hypoxemia and respiratory distress in COVID-19 patients. Objective: The objective of the study was to study the effect of the reduced form of methylene blue (MB) on the improvement of oxygen saturation (SpO2) and respiratory rate (RR). Methods: In an academic medical center, 80 hospitalized patients with severe COVID-19 were randomly assigned to receive either oral MB along with standard of care (SOC) (MB group, n = 40) or SOC only (SOC group, n=40). The primary outcomes were SpO2 and RR on the 3rd and 5th days. The secondary outcomes were hospital stay and mortality within 28 days. Results: In the MB group, a significant improvement in SpO2 and RR was observed on the 3rd day (for both, p < 0.0001) and also the 5th day (for both, p < 0.0001). In the SOC group, there was no significant improvement in SpO2 (p = 0.24) and RR (p = 0.20) on the 3rd day, although there was a significant improvement of SpO2 (p = 0.002) and RR (p = 0.01) on the 5th day. In the MB group in comparison to the SOC group, the rate ratio of increased SpO2 was 13.5 and 2.1 times on the 3rd and 5th days, respectively. In the MB group compared with the SOC group, the rate ratio of RR improvement was 10.1 and 3.7 times on the 3rd and 5th days, respectively. The hospital stay was significantly shortened in the MB group (p = 0.004), and the mortality was 12.5% and 22.5% in the MB and SOC groups, respectively. Conclusions: The addition of MB to the treatment protocols significantly improved SpO2 and respiratory distress in COVID-19 patients, which resulted in decreased hospital stay and mortality. ClinicalTrials.gov: NCT04370288

Methylene Blue for the Treatment of Health Conditions: a Scoping Review

HIGHLIGHTS This scoping review summarizes the findings of clinical trials using methylene blue (MB) for the treatment of various health conditions. This research method allowed mapping main findings, clarifying research topics, and identifying gaps in the literature.

Abstract studies evaluating effective drugs for health conditions are of crucial importance for public health. Methylene blue (MB) is an accessible synthetic drug that presents low toxicity and has been used in several health areas due to its effectiveness. Objective: this scoping review aims to provide a comprehensive overview of relevant research regarding the use of MB for the treatment of health conditions. Methods: a five-stage framework Arksey and O'maley scoping review was conducted. The literature was searched in Cochrane Library database using Mesh term "methylene blue". Data were collected by two independent reviewers and submitted to descriptive synthesis. Results: The search resulted in 429 records, from which 16 were included after exclusion criteria were applied. The therapeutic use of MB was identified for acute conditions (malaria and septic shock), chronic conditions (discogenic back pain, bipolar disorder, refractory neuropathic pain, and post-traumatic stress disorder), and postoperative care (vasoplegic syndrome, and pain after haemorrhoidectomy, lumbar discectomy, and traumatic thoracolumbar fixation). Conclusion: there is much evidence emerging from clinical trials about the therapeutic use of MB for acute, chronic, and postoperative conditions; however, many gaps were identified, which open further avenues for future research.

Microfiltración apical entre dos cementos de obturación: biocerámico y resinoso en premolares unirradiculares preparadas con protaper, y obturadas con condensación lateral / Apical microfiltration between two filling cements: bioceramic and resinous in unirradicular premolars prepared with protaper and sealed with lateral condensation

Resumen Objetivo: Comparar in vitro la microfiltración apical en premolares unirradiculares obturados con cemento a base de resina epóxica y cemento biocerámico. Método: 40 piezas dentales unirradiculares fueron seleccionadas y divididas en dos grupos para ser obturadas esperando el tiempo de fraguado de acuerdo con las instrucciones del fabricante. El grupo A se obturó con cemento biocerámico Endosequence y, el grupo B, con cemento a base de resina epóxica AH-Plus: posteriormente los dos grupos fueron sellados con barniz de uñas hasta 3mm de la parte apical, luego se colocaron las muestras en la incubadora a 37ºC. Ambos fueron sometidos a 750 ciclos de termociclado, para reproducir el ambiente parecido a la cavidad oral. Para análisis de microfiltración se utilizó el método de difusión del colorante, se sumergieron en azul de metileno al 2 % por 6 días, posteriormente los especímenes fueron sometidos en una bomba de vacío. Los dientes fueron cortados longitudinalmente para ser evaluadas mediante el estereomicroscopio. El análisis estadístico fue hecho mediante T-Student y Chi-Cuadrado. Resultados: El grupo A corresponde a Endosequence obtuvo una medida de 0,55mm y el grupo B corresponde a AH-Plus 1,20mm con un estimado de significancia de p=0,013. Conclusión: El análisis de ambos cementos de obturación demostró que Endosequence posee menor microfiltración apical que AH-Plus.

Abstract Aim: To compare in vitro by the stereomiscroscope the lesser apical microfiltration in uniradicular premolars, between the cement based on Epoxy Resin and Bioceramic cement. Method: The study was performed on 40 uniradicular dental pieces divided into two groups, the samples were prepared using the protaper system and the obturation was performed using the lateral condensation technique, the first group was sealed withEndosequence bioceramic cement and the second group with Cement based on AH-Plus epoxy resin, then sealed with nail varnish up to 3mm from the apical part, then the samples were placed in the incubator at 37ºC, waiting for the set time corresponding to each group according to the manufacturer's instructions . The two groups of the present study were submitted to 750 cycles of thermocycling, to provide an environment similar to the oral cavity, then longitudinal cuts were made to the samples. The microfiltration was evaluated using the dye diffusion method, immersed in 2% methylene blue for 6 days, then the specimens were subjected to a vacuum pump. The teeth were cut longitudinally to be evaluated by the stereomicroscope. The statistical analysis was through T-Student and Chi-Square. Results: Group A corresponds to Endosequence obtained a mean of 0,55mm and Group B corresponds to AH-Plus 1,20mm with an estimated significance of p = 0,013. Conclusion: Both obturation cements have significant differences, therefore Endosequence has less apical microfiltration than AHPlus.

Evaluation of the therapeutic effect of methylene blue on the liver of rats submitted to ischemia and reperfusion

Abstract Purpose: To analyze the effect of methylene blue (MB) therapy during the liver ischemia-reperfusion injury (I/R) process. Methods: Thirty-five male Wistar rats were used, (70%) submitted to partial ischemia (IR) or not (NIR) (30%) were obtained from the same animal. These animals were divided into six groups: 1) Sham (SH), 2) Sham with MB (SH-MB); 3) I/R, submitted to 60 minutes of partial ischemia and 15 minutes of reperfusion; 4) NI/R, without I/R obtained from the same animal of group I/R; 5) I/R-MB submitted to I/R and MB and 6) NI/R-MB, without I/R. Mitochondrial function was evaluated. Osmotic swelling of mitochondria as well as the determination of malondialdehyde (MDA) was evaluated. Serum (ALT/AST) dosages were also performed. MB was used at the concentration of 15mg/kg, 15 minutes before hepatic reperfusion. Statistical analysis was done by the Mann Whitney test at 5%. Results: State 3 shows inhibition in all ischemic groups. State 4 was increased in all groups, except the I/R-MB and NI/R-MB groups. RCR showed a decrease in all I/R and NI/R groups. Mitochondrial osmotic swelling showed an increase in all I/R NI/R groups in the presence or absence of MB. About MDA, there was a decrease in SH values in the presence of MB and this decrease was maintained in the I/R group. AST levels were increased in all ischemic with or without MB. Conclusions: The methylene blue was not able to restore the mitochondrial parameters studied. Also, it was able to decrease lipid peroxidation, preventing the formation of reactive oxygen species.

Does methylene blue attenuate inflammation in nonischemic lungs after lung transplantation? / O azul de metileno atenua a inflamação em pulmões não isquêmicos após transplante pulmonar?

ABSTRACT Objective: To evaluate whether methylene blue (MB) could minimize the effects of ischemia-reperfusion injury in the nonischemic lung on a lung transplantation rodent model. Methods: Forty female Sprague-Dawley rats were divided into 20 donors and 20 recipients. The 20 recipient rats were divided into two groups (n = 10) according to the treatment (0.9% saline vs. 1% MB solutions). All animals underwent unilateral lung transplantation. Recipients received 2 mL of saline or MB intraperitoneally prior to transplantation. After 2 h of reperfusion, the animals were euthanized and histopathological and immunohistochemical analyses were performed in the nonischemic lung. Results: There was a significant decrease in inflammation-neutrophil count and intercellular adhesion molecule-1 (ICAM-1) expression in lung parenchyma were higher in the saline group in comparison with the MB group-and in apoptosis-caspase-3 expression was higher in the saline group and Bcl-2 expression was higher in MB group. Conclusions: MB is an effective drug for the protection of nonischemic lungs against inflammation and apoptosis following unilateral lung transplantation in rats.

RESUMO Objetivo: Avaliar se o azul de metileno (AM) pode minimizar os efeitos da lesão de isquemia-reperfusão sobre o pulmão não isquêmico em um modelo de transplante pulmonar em roedores. Métodos: Quarenta ratas Sprague-Dawley foram divididas em 20 doadoras e 20 receptoras. As 20 ratas receptoras foram divididas em dois grupos (n = 10) de acordo com o tratamento (solução salina a 0,9% vs. AM a 1%). Todos os animais foram submetidos a transplante pulmonar unilateral. As receptoras receberam 2 ml de solução salina ou de AM por via intraperitoneal antes do transplante. Após 2 h de reperfusão, os animais foram sacrificados, e foram realizadas análises histopatológicas e imuno-histoquímicas no pulmão não isquêmico. Resultados: Houve diminuição significativa da inflamação - a contagem de neutrófilos e a expressão de intercellular adhesion molecule-1 (ICAM-1, molécula de adesão intercelular-1) foram maiores no grupo salina em comparação com o grupo AM - e da apoptose - a expressão de caspase-3 foi maior no grupo salina, e a expressão de Bcl-2 foi maior no grupo AM. Conclusões: O AM é uma droga eficaz para a proteção de pulmões não isquêmicos contra inflamação e apoptose após transplante pulmonar unilateral em ratos.

Use of photodynamic inactivation for in vitro reduction of prevalent bacteria in Fournier's Gangrene

ABSTRACT Fournier's Gangrene (FG) is an infectious disease caused by several synergic microbes, with high morbidity and mortality rates; therefore, the search for new less invasive and mutilating treatments, with faster recovery, has been proposed. Surgical intervention, the use of several systemic and topic antibiotics, and hyperbaric oxygen therapy are currently the best approach for the treatment of these patients. The use of Photodynamic Inactivation (PDI) aims to lower morbidity and mortality, by reducing bacterial microbiota and speeding wound healing. In the present study, viable bacteria were separated in four groups: Group L-/F- (no irradiation with red laser and absence of methylene blue photosensitizer), Group L-/F+ (no irradiation with red laser and presence of methylene blue), Group L+/F- (irradiation with red laser and absence of methylene blue) and L+/F+ (irradiation with red laser associated to methylene blue). In all groups, exposure time to treatment was 5, 10 and 15 minutes. The concentration of methylene blue photosensitizer was 0.1mg/L, and the dose of red laser (660nm wave length) was 176.9mW/cm2. Following irradiation, the reduction of number of bacteria was evaluated, and the results were expressed in colony forming units (CFU) and as exponential reduction. As the main results, in the L+/F+ group, there were no Clostridium perfringens and Staphylococcus aureus CFUs and there was a reduction of Escherichia coli that was not observed in the other groups.

Metahemoglobinemia relacionada con ingesta de puré de acelgas / Methemoglobinemia due to ingestion of chard puree

La metahemoglobinemia es un síndrome clínico dado por la presencia de una forma aberrante de hemoglobina, ocasionada por diversos agentes oxidantes. Se describe un caso clínico de metahemoglobinemia severa asociada a la ingesta de puré de acelgas con alto contenido en nitratos y nitritos. Paciente de un año, con antecedentes de comunicación interauricular (CIA), que presentó un cambio en coloración de la piel 7 h antes, en forma progresiva, acompañado de vómitos. Ingresó al Departamento de Emergencia con cianosis generalizada que no mejoró con oxigenoterapia, taquicardia y tendencia a hipotensión arterial. En cuidados intensivos se realizó ecocardiograma que evidenció CIA sin repercusión hemodinámica. Metahemoglobinemia 37%. Se realizó dosis de azul de metileno al 1% por vía intravenosa, con franca mejoría clínica a la hora de la administración del antídoto y descenso de niveles de metahemoglobina. Alta médica a las 36 horas del ingreso. Existía una relación cronológica entre la exposición a nitratos por ingesta de un puré de acelgas y la aparición del cuadro. Los niveles de nitratos hallados en dicho alimento fueron muy elevados considerando estándares internacionales, lo que sumado a una inadecuada conservación del alimento cocido los días previos, permitió confirmar el planteo etiológico realizado. Resulta importante sospechar esta entidad patológica poco frecuente frente a cianosis que no mejora con oxígeno, y prevenir cuadros similares al descrito mediante una adecuada manipulación y conservación de las verduras con alto contenido en nitratos.

Methemoglobinemia is a clinical syndrome due to the presence of an aberrant form of hemoglobin, caused by various oxidizing agents. The study reports a case of severe methemoglobinemia associated with the ingestion of chard puree with high levels of nitrates and nitrites. A 1-year-old patient with a history of atrial septal defect (ASD), who progressively showed change of skin color 7 hours earlier, accompanied by vomiting. She was admitted to the Emergency Department with generalized cyanosis not improving with oxygen therapy, tachycardia and tendency to hypotension. In the intensive care unit, an echocardiogram showed ASD without hemodynamic complications. Methemoglobinemia 37%. A 1% methylene blue dose was administered intravenously, with clinical improvement one hour after antidote administration and decrease in methemoglobin levels. Medical discharge at 36 hours of admission. There was a chronological relationship between nitrates exposure by ingestion of chard puree and the clinical onset of methemoglobinemia. The toxic cause was confirmed after high nitrates levels were found in this vegetable considering international standards, and an inadequate preservation of the cooked chard on previous days. It is important to suspect this rare pathological entity when cyanosis fails to improve with oxygen, and to prevent poisonings similar to those described by an adequate manipulation and preservation of vegetables with high nitrate levels.

Terapia fotodinámica como adyuvante en el tratamiento periodontal no quirúrgico: evaluación clínica / Photodynamic therapy as an adjuvant in nonsurgical periodontal treatment:A clinical evaluation

La terapia fotodinámica es considerada como un procedimiento no invasivo dentro de la práctica periodontal, reduciendo la morbilidad e incrementando la comodidad del paciente. Objetivo: el objetivo del presente estudio fue evaluar la eficacia clínica del uso de terapia fotodinámica como adyuvante al raspado y alisado radicular en periodontitis crónica. Material y métodos: se realizó un estudio comparativo, correlacional, aplicado a 20 pacientes con periodontitis crónica moderada-avanzada (pérdida de inserción clínica >- 6 mm) durante el raspado y alisado radicular no quirúrgico, con la adición de azul de metileno, donde se administró la terapia fotodinámica en dos sesiones, evaluando los parámetros clínicos (profundidad de bolsa, nivel de inserción clínico, índice de placa, sangrado al sondeo y recesión gingival). Resultados: tras cuatro semanas de evaluación postratamiento, no se observaron diferencias significativas en los parámetros clínicos entre el grupo experimental y control, aunque se encontró un mayor aumento de recesión gingival con el uso de la terapia fotodinámica (p=0,353). Conclusión: el uso de la terapia fotodinámica como adyuvante durante el raspado y alisado radicular convencional no proporciona beneficios adicionales al tratamiento periodontal convencional.

Photodynamic therapy is considered a non-invasiveprocedure in periodontal practice, one which reduces morbidity andincreases patient comfort. Objective: The aim of this study was toevaluate the clinical effi cacy of photodynamic therapy as an adjuvantto scaling and root planing in chronic periodontitis. Material andmethods: A comparative and correlational study was carried out on 20patients with moderate to advanced chronic periodontitis (attachmentloss ≥ 6 mm) during nonsurgical scaling and root planing, including theaddition of methylene blue. The photodynamic therapy was applied intwo sessions, during which the clinical parameters (pocket depth, clinicalattachment level, plaque index, bleeding on probing, and gingivalrecession) were assessed. Results: After four weeks of post-treatmentassessment, no signifi cant diff erences were found between the clinicalparameters of the experimental and control groups, although a greaterincrease in gingival recession was found when photodynamic therapywas used (p = 0.353). Conclusion: The use of photodynamic therapyas an adjuvant in conventional scaling and root planing provides noadditional benefi ts to conventional periodontal treatmen.

Direct intraperitoneal resuscitation with lidocaine, methylene blue and pentoxiphylline combination does not decreases inflammation after intestinal ischemia-reperfusion injury in rats

ABSTRACT PURPOSE: To evaluate the effects of an intraperitoneal solution of methylene blue (MB), lidocaine and pentoxyphylline (PTX) on intestinal ischemic and reperfusion injury METHODS: Superior mesenteric artery was isolated and clamped in 36 adult male Sprague Dawley rats. After 60 minutes, clamp was removed and a group received intraperitoneally UNITO solution (PTX 25mg/kg + lidocaine 5mg/kg + MB 2mg/kg), while the other group was treated with warm 0.9% NaCl solution. Rats were euthanized 45 min after drug administration. Lung and bowel were collected for histological evaluation (using Park's score) and determination of myeloperoxidase (MPO) and malondialdehyde (MDA) levels. RESULTS: Control samples showed lymphoplasmocytic infiltrate and crypt necrosis of villi. MPO and MDA measurements shown no differences between treated and control groups. CONCLUSION: The combination of lidocaine, methylene blue and pentoxyphylline administered intraperitoneally at the studied dose, did not decreased histological lesion scores and biochemical markers levels in intestinal ischemia/reperfusion injury.

Incidência de fistula após palatoplastia à Von Langenbeck com veloplastia intravelar estendida / Incidence of cleft palate fistula after von Langenbeck palatoplasty with extended intravelar veloplasty

Introdução: A fissura labiopalatina é a deformidade craniofacial mais frequente e sua incidência é estimada em 1:600 nascidos vivos no Brasil. O objetivo desse estudo é avaliar a incidência de fístulas para os pacientes submetidos à palatoplastia com veloplastia intravelar estendida. Métodos: Trata-se de estudo descritivo retrospectivo com 25 pacientes que foram operados pelo mesmo cirurgião no período de setembro de 2011 a setembro de 2012. A técnica de Von Langenbeck, juntamente com a veloplastia intravelar estendida, foi realizada em todos os pacientes. Foram excluídos do estudo os pacientes com fístulas palatinas, portadores de síndromes ou outras malformações. A idade média da realização da palatoplastia foi de 30,6 meses, variando de 12 meses a 159 meses. Foram selecionados 19 pacientes: onze (58%) do gênero masculino e oito (42%) do gênero feminino. A fissura palatal isolada foi a mais comum, encontrada em nove (47%) pacientes. A fissura transforame esquerda estava presente em sete (37%) pacientes e três (16%) pacientes eram portadores de fissura transforame bilateral. Resultados: Somente dois (11%) pacientes evoluíram com fístula palatina até o acompanhamento pós-operatório de 6 meses. Conclusão: A técnica de Von Langenbeck associada à veloplastia estendida mostrou-se com baixa incidência de fístulas palatinas (11%) quando comparada ao índice encontrado na literatura mundial (7% a 42%).

Introduction: Cleft lip and palate is the most common craniofacial deformity, with an estimated incidence of 1 case per 600 live births in Brazil. The aim of this study was to determine the incidence of fistula among patients undergoing palatoplasty with extended intravelar veloplasty. Methods: This retrospective descriptive study evaluated 25 patients operated between September 2011 and September 2012 by the same surgeon. The von Langenbeck technique combined with extended intravelar veloplasty was performed in all patients. The study excluded patients with palatal fistulas, syndromes, or other malformations. The age at palatoplasty varied between 12 and 159 months, and the average age was 30.6 months. Nineteen patients were selected, comprising 11 (58%) male patients and 8 (42%) female patients. Isolated cleft palate was the most common deformity, found in 9 (47%) patients. Unilateral (left) trans-foramen cleft was present in 7 (37%) patients, and bilateral trans-foramen cleft was seen in 3 (16%) patients. Results: Only 2 (11%) patients had palatal fistula in a postoperative follow-up period of 6 months. Conclusion: The von Langenbeck technique associated with extended veloplasty resulted in a low incidence of cleft palate fistulas (11%) compared with the rate found in previous studies (7%-42%).

Uso da terapia fotodinâmica empregando azul de metileno a 2% como agente fotossensibilizante para o controle de crescimento tumoral em calopsita (Nynphicus hollandicus) portador de sarcoma / Use of photodynamic therapy using methylene blue 2% as photosensitizing agent to control tumor growthin a cockatiel (Nynphicus hollandicus) carrier sarcoma

Os sarcomas são neoplasias de origem mesenquimal. Os sarcomas são neoplasias de origem mesenquimal, sendo considerados tumores agressivos que frequentemente gerammetástases e rápida debilidade. A indicação terapêutica é a exérese tumoral enquanto ainda não houver metástases. A terapiafotodinâmica tem sido utilizada para o tratamento de neoplasias tanto em humanos quanto em animais e muitos estudos têm sidodesenvolvidos de maneira a aprimorar e avaliar a efetividade da técnica. Uma calopsita (Nynphycus hollandicus) foi levada paraatendimento no Hospital Veterinário Professor Firmino Mársico Filho (HUVET-UFF) apresentando nódulo em região escapuloumeral.O diagnóstico citopatológico concluiu sarcoma. Devido à localização e impossibilidade de exérese cirúrgica, optou-sepela realização da terapia fotodinâmica utilizando-se o azul de metileno a 2% como agente fotossensibilizador e fonte de luz laserdiodo (Vetlight®). A terapia fotodinâmica empregando o azul de metileno como agente fotossensibilizante foi eficaz no controle docrescimento tumoral até os 30 dias posteriores observados.

Sarcoma are tumors of mesenchymal origin. They are aggressive tumors that frequently metastasize and rapid weakness. Thetherapeutic indication is the resection while there is still no metastasis. Photodynamic therapy has been used for the treatment ofcancer in both humans and animals and many studies have been developed in order to improve and enhance the effectivenessof the techniques. A Nynphycus hollandicus was brought to HUVET-UFF clinical service presenting a lump in scapulohumeralregion. The citological exam had a diagnosis of sarcoma. Due to its location and inability to surgical excision, was decided bythe completion of photodynamic therapy using methylene blue at 2% as a phosensitizing agent and laser diode light source(Vetlight®). Photodynamic therapy using methylene blue as photosensitizing agent was effective in controlling growth up to 30days later observed.

Avaliação dos efeitos da terapia fotodinâmica utilizando compostos fenotiazínicos sobre melanoma in vitro / Assessment of the effects of photodynamic therapy using phenothiazine compounds on in vitro melanoma

O câncer é considerado a segunda maior causa de morte em países ocidentais. Nos Estados Unidos da América (EUA) as mortes por câncer anualmente superam a soma das mortes provocadas em guerras como Vietnã, Coreia e as Grandes Guerras Mundiais. Apresentando-se como a mais agressiva das neoplasias dermatológicas, o melanoma cutâneo está associado à cerca de 75% das causas de morte por câncer de pele. Um dos tratamentos estudados para aplicação em pacientes com esta e outras patologias é a Terapia Fotodinâmica (TFD), que é baseada no uso de corantes de baixa toxicidade, que tem seletividade por alguns tecidos ou células e quando ativados por baixas doses de luz visível induzam alterações celulares como a produção ERO. Os fenotiazínicos são moléculas catiônicas com, absorção de luz na região entre 620-660nm, espectro que permite maior penetração nos tecidos, promovendo maior ação do composto em tecidos mais profundos. A irradiação foi feita com luz lazer com λ= 660nm, com potência de 100 mW, densidade de energia de 150 J/cm2 por 1 min. As quantificações de viabilidade celular foram feitas por método colorimétrico, e realizada a leitura em leitor de microplaca a 655 nm. A determinação das IC50 do Azul de Metileno e Azul de Toluidina O foi 2,5 μM e 8,4 μM, respectivamente. Utilizando a TFD observamos que os efeitos citotóxicos dos compostos fenotiazínicos apresentam significância estatística (p<0,05) tanto em AM quanto em ATO. Em microscopia óptica com coloração por May-Grunwald-Giemsa observamos que existe uma aparente diminuição do conteúdo citoplasmático e preservação nuclear nas células tratadas com TFD, além da diminuição da quantidade de células por campo. A avaliação ultraestrutural por Microscopia de varredura mostra células com visível extração citoplasmática após a TFD, apontando uma lesão membranar. Novas avaliações precisam ser feitas como estabelecer um melhor protocolo para aumentar os efeitos da TFD.

Cancer is considered the second leading cause of death in Western countries. In the United States of America (USA) cancer deaths annually exceed the total of deaths in wars like Vietnam, Korea and the Great World Wars. Presenting as the most aggressive of the skin neoplasms, cutaneous melanoma is associated with circa 75% of the deaths from skin cancer. One of the therapies used in patients with melanoma and other pathologies is Photodynamic Therapy (PDT), which is based upon the use of dyes of low toxicity, which has selectivity for certain tissues or cells and low doses when activated by visible light induce cellular changes such as ROS production. The phenothiazic dyes are cationic molecules, absorption of light in the region between 620-660 nm, allowing greater spectrum tissue penetration, promoting higher activity of the compound in deeper tissues. The irradiation was performed with laser light with λ = 660 nm with 100 mW, energy density of 150 J/cm2 for 1 min. Quantification of cell viability was performed by the colorimetric method, and performed a reading in a microplate reader at 655nm. Determination of IC50 of Methylene Blue and Toluidine Blue O was 2.5 μM and 8.4 μM respectively. Using the protocol of PDT observed that there is one of the cytotoxic effects of phenothiazine compounds with statistical significance (p <0.05) in both MB and in TBO. Through optical microscopy by staining with fast Panotic was observed that there is an apparent decrease in the cytoplasm in cells treated with MB and TBO as well as reduced number of cells per field. The Scanning Electron Microscopy, shows cells with cytoplasm extraction, after PDT, indicated possibility, membrane damage. New assessments need to be made to establish a better protocol to potentiate the effects of PDT.

Inibição da guanilato ciclase pelo azul de metileno no choque circulatório causado por pancreatite aguda necrosante: uma palavra de cuidado embasada em modelo suíno / Guanylate cyclase inhibition by methylene blue in circulatory shock caused by acute necrotizing pancreatitis: a word of caution based on a porcine model

OBJETIVO: estudar o uso terapêutico do bloqueio da guanilato ciclase pelo azul de metileno em um modelo experimental de pancreatite aguda grave em suínos. MÉTODOS: a pancreatite aguda necrotizante foi induzida em porcos anestesiados por infusão ductal pancreática retrógrada de 1ml/kg de taurocolato de sódio a 5% e 8U/kg de enteroquinase. Três grupos foram estudados (n=5): controle (C), pancreatite (PA), "bolus" de azul seguido por pancreatite (AM+PA). Os dados incluíram enzimas séricas e do líquido abdominal, variáveis hemodinâmicas, hemogasometria arterial, volume de líquido abdominal, marcadores inflamatórios plasmáticos, nitrito/nitrato e mieloperoxidase e malondialdeído plasmático. Aplicou-se a análise de variância seguida do pós-teste de Bonferroni (p<0,05). RESULTADOS: os valores de amilase e lipase foram três e dez vezes mais elevados no grupo PA. A atividade da mieloperoxidase foi 50% superior no grupo PA. Os dados hemodinâmicos indicaram choque hipovolêmico precoce seguido de choque cardiogênico. Observou-se grave translocação de líquidos para a cavidade peritoneal. A nitrito/nitrato plasmática permaneceu inalterada. O grupo AM+PA teve aumento de cinco vezes do mieloperoxidase em comparação com o grupo C. CONCLUSÕES: a utilização de azul de metileno em suínos com pancreatite não demonstrou efeitos significativos sobre variáveis hemodinâmicas e inflamatórias. Seu uso terapêutico na pancreatite necro-hemorrágica pode ser inadequado e extremo cuidado deve ser tomado dado o aumento da peroxidação lipídica evidenciado pelo aumento dos valores do malondialdeído.

OBJECTIVE: To study the therapeutic application of guanylate cyclase inhibition by methylene blue in an experimental model of acute pancreatitis in pigs. METHODS: acute necrotizing pancreatitis was induced in anesthetized pigs by the retrograde infusion of 1 ml/kg of 5% sodium taurocholate and 8 U/kg enterokinase in the pancreatic duct. Three groups were studied (n = 5): control (C), pancreatitis (AP), and MB bolus followed by pancreatitis (MB+P). The data included serum and abdominal fluid enzymes, hemodynamic variables, arterial hemogasometry, abdominal fluid volume, inflammatory markers, plasma nitrite/nitrate (NOx), plasma myeloperoxidase (MPO) and plasma malondialdehyde (MDA). One- and two-way analysis of variance (ANOVA) was performed, followed by the Bonferroni test (p < 0.05). RESULTS: amylase and lipase were three and 10-fold higher in the AP group. Myeloperoxidase activity was 50% higher in the AP group. The hemodynamic data indicated early hypovolemic shock followed by cardiogenic shock. Severe fluid translocation to the peritoneal cavity was observed. Plasma NOx remained unchanged. The MB+P group had a five-fold increase in MDA compared with the C group. CONCLUSION: preemptive application of MB in pigs with AP demonstrated no significant effects on hemodynamic and inflammatory variables. The use of MB is inadequate in cases of exponential NO release, and extreme caution must be exercised, given the increase in lipid peroxidation based on the malondialdehyde dosage.

Ionic imbalance and lack of effect of adjuvant treatment with methylene blue in the hamster model of leptospirosis

Leptospirosis in humans usually involves hypokalaemia and hypomagnesaemia and the putative mechanism underlying such ionic imbalances may be related to nitric oxide (NO) production. We previously demonstrated the correlation between serum levels of NO and the severity of renal disease in patients with severe leptospirosis. Methylene blue inhibits soluble guanylyl cyclase (downstream of the action of any NO synthase isoforms) and was recently reported to have beneficial effects on clinical and experimental sepsis. We investigated the occurrence of serum ionic changes in experimental leptospirosis at various time points (4, 8, 16 and 28 days) in a hamster model. We also determined the effect of methylene blue treatment when administered as an adjuvant therapy, combined with late initiation of standard antibiotic (ampicillin) treatment. Hypokalaemia was not reproduced in this model: all of the groups developed increased levels of serum potassium (K). Furthermore, hypermagnesaemia, rather than magnesium (Mg) depletion, was observed in this hamster model of acute infection. These findings may be associated with an accelerated progression to acute renal failure. Adjuvant treatment with methylene blue had no effect on survival or serum Mg and K levels during acute-phase leptospirosis in hamsters. .

The in vivo antimalarial activity of methylene blue combined with pyrimethamine, chloroquine and quinine

The effectiveness of methylene blue (MB) combined with pyrimethamine (PYR), chloroquine (CQ) or quinine (Q) was examined in a classical four-day suppressive test against a causative agent of rodent malaria, Plasmodium berghei. A marked potentiation was observed when MB was administered at a non-curative dose of 15 mg/kg/day in combination with PYR (0.19 mg/kg/day) or Q (25 mg/kg/day). No synergy was found between MB (15 mg/Kg) and CQ (0.75 mg/Kg). Our results suggest that the combination of MB with PYR or Q may improve the efficacy of these currently used antimalarial drugs.

Methylene blue administration in the compound 48/80-induced anaphylactic shock: hemodynamic study in pigs / Administração de azul de metileno no choque anafilático induzido por composto 48/80: estudo hemodinâmico em suínos

PURPOSE: To verify if the methylene blue (MB) administration prevents and/or reverses the compound 48/80 (C48/80)-induced anaphylactic shock in pigs. METHODS: Female Dalland pigs were anesthetized and had the hemodynamic parameters recorded during the necessary time to administer some drugs and observe their effect. The animals were randomly assigned to one of the five groups: 1) control; 2) MB: the animals received a bolus injection of MB (2 mg/kg) followed by continuous infusion of MB (2.66 mg/Kg/h delivered by syringe infusion pump); 3) C48/80: the animals received a bolus injection of C48/80 (4 mg/kg); 4) C48/80+MB: the animals received a bolus injection of C48/80 (4 mg/kg) and 10 minutes after the C48/80 administration the animals received a bolus injection of MB (2 mg/kg) followed by continuous infusion of MB (2.66 mg/Kg/h delivered by syringe infusion pump); 5) MB+C48/80: the animals received a bolus injection of MB (2 mg/kg) and 3 minutes later they received a bolus injection of C48/80 (4 mg/kg). RESULTS: The intravenous infusion of MB alone caused no changes in the mean arterial pressure (MAP) showing that the administered MB dose was safe in this experimental model. The C48/80 was effective in producing experimental anaphylactic shock since it was observed a decrease in both MAP and cardiac output (CO) after its administration. The MB did not prevent or reverse the C48/80-induced anaphylactic shock in this model. In fact, the MAP of the animals with anaphylactic shock treated with MB decreased even more than the MAP of the animals from the C48/80 group. On the other hand, the C48/80-induced epidermal alterations disappeared after the MB infusion. CONCLUSION: Despite our data, the clinical manifestations improvement brings some optimism and does not allow excluding the MB as a possible therapeutic option in the anaphylactic shock.

OBJETIVO: Verificar se a administração de azul de metileno (AM) previne e/ou reverte o choque anafilático induzido por composto 48/80 (C48/80) em suínos. MÉTODOS: Porcos fêmeas Dalland foram anestesiados e tiveram os parâmetros hemodinâmicos registados durante o tempo necessário para administrar algumas drogas e observar seu efeito. Os animais foram aleatoriamente destribuídos em um dos cinco grupos: 1) controle, 2) AM: os animais receberam uma injeção em bolus de AM (2mg/kg), seguido de infusão contínua de AM (2,66mg/Kg /h por bomba de infusão de seringa); 3) C48/80: os animais receberam uma injeção em bolus de C48/80 (4mg/kg); 4) C48/80 + AM: os animais receberam uma injeção em bolus de C48/80 (4mg/kg) e 10 minutos após a administração de C48/80 os animais receberam uma injeção em bolus de AM (2mg/kg), seguido de infusão contínua de AM (2,66mg/kg/h por bomba de infusão de seringa); 5) AM+C48/80: os animais receberam uma injeção em bolus de AM (2mg/kg) e três minutos depois, receberam uma injeção em bolus de C48/80 (4mg/kg). RESULTADOS: A infusão intravenosa de AM não causou mudanças na pressão arterial média (PAM), mostrando que a dose de AM administrada foi segura neste modelo experimental. O C48/80 foi eficaz na indução do choque anafilático experimental, uma vez que foi observada redução na PAM e débito cardíaco (DC), após a sua administração. O AM não preveniu ou reverte o choque anafilático induzido por C48/80 neste modelo. Na verdade, a PAM dos animais com choque anafilático tratados com AM diminuiu mais do que o PAM dos animais do grupo C48/80. Por outro lado, as alterações epidérmicas induzidas pelo C48/80 desapareceu após a infusão do AM. CONCLUSÃO: Apesar dos resultados a melhora clínica das manifestações anafiláticas permite considerar a possibilidade do azul de metileno como opção terapêutica no tratamento do choque anafilático.