Modificaciones en el nivel de anión superóxido en leche materna, según la ingesta de flavonoides y carotenoides / Modifications of the superoxide anion level in breast milk by the intake of flavonoids and carotenoids

Resumen: Objetivo: Asociar la ingesta de flavonoides y carotenoides con el nivel en leche materna del anión superóxido, como marcador de estrés oxidativo. Material y métodos: Durante el periodo 2013-2015 se estudió a 100 mujeres lactantes de Córdoba (Argentina), dentro los primeros seis meses posparto; se evaluaron sus datos sanitarios, ingesta alimentaria y nivel lácteo del anión con regresión logística múltiple. Resultados: La ingesta de flavonoides, carotenoides provitamínicos y carotenoides no provitaminas fue de 72 (61) mg/día, 1 813 (1657) µg/día y 5 427 (3664) µg/día, respectivamente. El anión se asoció con la ingesta de flavanoles (RM=1.081; IC95 1.001-1.167) y flavanonas (RM=1.025; IC95 1.001-1.048). No se observó este efecto con otros flavonoides ni con los carotenoides. Conclusiones: La ingesta de flavanoles y flavanonas aumenta el riesgo de oxidación láctea, lo cual es relevante para realizar recomendaciones dietéticas.

Abstract: Objective: To associate the intake of flavonoids and carotenoids with the breast milk level of superoxide anion, as an oxidative stress marker. Materials and methods: 100 women from Cordoba (Argentina), who breastfed within the first postpartum 6 months, were studied during the 2013-2015 period, by evaluating their sanitary data, food intake and anion level in milk with multiple logistic regression. Results: The intake of flavonoids, provitamin A carotenoids and non-provitamin carotenoids was 72 (61) mg/d, 1813 (1 657) µg/d y 5427 (3 664) µg/d, respectively. The anion was associated with the intake of flavanols (OR=1.081; CI95 1.001-1.167) y flavanones (OR=1.025; CI95 1.001-1.048). This effect was not seen with other flavonoids and carotenoids. Conclusions: Intake of flavanols and flavanones increases milk oxidation risk, which is relevant to develop diet recommendations.

Aerobic Swim Training Restores Aortic Endothelial Function by Decreasing Superoxide Levels in Spontaneously Hypertensive Rats

OBJECTIVE: We aimed to determine whether aerobic training decreases superoxide levels, increases nitric oxide levels, and improves endothelium-dependent vasodilation in the aortas of spontaneously hypertensive rats. METHODS: Spontaneously hypertensive rats (SHR) and Wistar Kyoto rats (WKY) were distributed into 2 groups: sedentary (SHRsd and WKYsd, n=10 each) and swimming-trained (SHRtr, n=10 and WKYtr, n=10, respectively). The trained group participated in training sessions 5 days/week for 1 h/day with an additional work load of 4% of the animal’s body weight. After a 10-week sedentary or aerobic training period, the rats were euthanized. The thoracic aortas were removed to evaluate the vasodilator response to acetylcholine (10-10 to 10-4 M) with or without preincubation with L-NG-nitro-L-arginine methyl ester hydrochloride (L-NAME; 10-4 M) in vitro. The aortic tissue was also used to assess the levels of the endothelial nitric oxide synthase and nicotinamide adenine dinucleotide oxidase subunit isoforms 1 and 4 proteins, as well as the superoxide and nitrite contents. Blood pressure was measured using a computerized tail-cuff system. RESULTS: Aerobic training significantly increased the acetylcholine-induced maximum vasodilation observed in the SHRtr group compared with the SHRsd group (85.9±4.3 vs. 71.6±5.2%). Additionally, in the SHRtr group, superoxide levels were significantly decreased, nitric oxide bioavailability was improved, and the levels of the nicotinamide adenine dinucleotide oxidase subunit isoform 4 protein were decreased compared to the SHRsd group. Moreover, after training, the blood pressure of the SHRtr group decreased compared to the SHRsd group. Exercise training had no effect on the blood pressure of the WKYtr group. CONCLUSIONS: In SHR, aerobic swim training decreased vascular superoxide generation by nicotinamide adenine dinucleotide oxidase subunit isoform 4 and increased nitric oxide bioavailability, thereby improving endothelial function.

The antioxidative defense system is involved in the premature senescence in transgenic tobacco (Nicotiana tabacum NC89)

BACKGROUND: α-Farnesene is a volatile sesquiterpene synthesized by the plant mevalonate (MVA) pathway through the action of α-farnesene synthase. The α-farnesene synthase 1 (MdAFS1) gene was isolated from apple peel (var. white winterpearmain), and transformed into tobacco (Nicotiana tabacum NC89). The transgenic plants had faster stem elongation during vegetative growth and earlier flowering than wild type (WT). Our studies focused on the transgenic tobacco phenotype. RESULTS: The levels of chlorophyll and soluble protein decreased and a lower seed biomass and reduced net photosynthetic rate (Pn) in transgenic plants. Reactive oxygen species (ROS) such as hydrogen peroxide (H2O2) and superoxide radicals (O2._) had higher levels in transgenics compared to controls. Transgenic plants also had enhanced sensitivity to oxidative stress. The transcriptome of 8-week-old plants was studied to detect molecular changes. Differentially expressed unigene analysis showed that ubiquitin-mediated proteolysis, cell growth, and death unigenes were upregulated. Unigenes related to photosynthesis, antioxidant activity, and nitrogen metabolism were downregulated. Combined with the expression analysis of senescence marker genes, these results indicate that senescence started in the leaves of the transgenic plants at the vegetative growth stage. CONCLUSIONS: The antioxidative defense system was compromised and the accumulation of reactive oxygen species (ROS) played an important role in the premature aging of transgenic plants.

Superoxide overproduction and kidney fibrosis: a new animal model / Produção aumentada de superóxido e fibrose renal: novo modelo animal

Objective To establish whether the mutation in the Immp2L gene induces renal fibrosis and whether aging exacerbates renal morphology in mice. Methods Female mutant mice with mutation in the inner mitochondrial membrane peptidase 2-like protein at 3 and 18 months of age were used. Renal fibrosis was analyzed using classic fibrosis score, Masson’s trichrome staining, and analysis of profibrotic markers using real time polymerase chain reaction (superoxide dismutase 1, metalloproteinase-9, erythropoietin, transforming growth factor beta), and immunostaining (fibroblasts and Type IV collagen). Oxidative stress markers were determined by immunohistochemistry. The number of renal apoptotic cells was determined. Renal function was estimated by serum creatinine. Results Young mutant mice had significantly more glomerulosclerosis than age-matched mice (p=0.034). Mutant mice had more tubular casts (p=0.025), collagen deposition (p=0.019), and collagen type IV expression (p<0.001). Superoxide dismutase 1 expression was significantly higher in young mutants (p=0.038). Old mutants exhibited significantly higher expression of the fibroblast marker and macrophage marker (p=0.007 and p=0.012, respectively). The real time polymerase chain reaction of metalloproteinase-9 and erythropoietin were enhanced 2.5- and 6-fold, respectively, in old mutants. Serum creatinine was significantly higher in old mutants (p<0.001). Conclusion This mutation altered renal architecture by increasing the deposition of extracellular matrix, oxidative stress, and inflammation, suggesting a protective role of Immp2L against renal fibrosis. .

Objetivo Estabelecer se a mutação no gene Immp2L induz à fibrose renal e se o envelhecimento exacerba a morfologia renal em camundongos. Métodos Foram usadas fêmeas de camundongos mutantes para proteína semelhante à peptidase 2 da camada interna da mitocôndria, com 3 e 18 meses de idade. Para analisar a fibrose renal, foram usados o escore clássico de fibrose, a coloração com tricrômio de Masson, e a análise de marcadores profibróticos, por meio da reação em cadeia de polimerase em tempo real (superóxido dismutase 1, metalonoproteinase-9, eritropoietina e fator transformador de crescimento beta), e a imunocoloração (fibroblastos e colágeno IV). Marcadores de estresse oxidativo foram determinados por imuno-histoquímica. O número de células apoptóticas renais foi analisado. A função renal foi estimada por creatinina sérica. Resultados Camundongos mutantes jovens apresentaram glomeruloesclerose em quantidade significativamente maior que animais da mesma idade (p=0,034). Os mutantes mostraram maior formação de cilindros tubulares (p=0,025), deposição de colágeno (p=0,019) e maior expressão de colágeno do tipo IV (p<0,001). A expressão de superóxido dismutase 1 foi maior em mutantes jovens (p=0,038). Mutantes idosas exibiram maior expressão dos marcadores de fibroblastos e macrófagos (p=0,007 e p=0,012, respectivamente). As reações da cadeia de polimerase em tempo real da metalanoproteinase-9 e da eritropoietina estavam aumentadas em 2,5 e 6 vezes, respectivamente, em mutantes idosas. A creatinina sérica foi significantemente maior em animais idosos mutantes (p<0,001). Conclusão Essa mutação alterou a arquitetura renal pelo aumento da deposição de matriz extracelular, estresse oxidativo e inflamação, sugerindo papel de proteção de Immp2L contra a fibrose renal. .

Protective effect of 4,4'-diaminodiphenylsulfone against paraquat-induced mouse lung injury

Although 4,4'-diaminodiphenylsulfone (DDS, dapsone) has been used to treat several dermatologic conditions, including Hansen disease, for the past several decades, its mode of action has remained a topic of debate. We recently reported that DDS treatment significantly extends the lifespan of the nematode C. elegans by decreasing the generation of reactive oxygen species. Additionally, in in vitro experiments using non-phagocytic human fibroblasts, we found that DDS effectively counteracted the toxicity of paraquat (PQ). In the present study, we extended our work to test the protective effect of DDS against PQ in vivo using a mouse lung injury model. Oral administration of DDS to mice significantly attenuated the lung tissue damage caused by subsequent administration of PQ. Moreover, DDS reduced the local expression of mRNA transcripts encoding inflammation-related molecules, including endothelin-1 (ET-1), macrophage inflammatory protein-1alpha (MIP-1alpha), and transforming growth factor-beta (TGF-beta). In addition, DDS decreased the PQ-induced expression of NADPH oxidase mRNA and activation of protein kinase Cmicro (PKCmicro). DDS treatment also decreased the PQ-induced generation of superoxide anions in mouse lung fibroblasts. Taken together, these data suggest the novel efficacy of DDS as an effective protective agent against oxidative stress-induced tissue damages.

Antioxidant isolated from Schisandra propinqua (Wall. ) Baill

Schisandra propinqua (Wall.) Baill.(Schisandraceae) is widely used as a Chinese folk medicine. In this study, activity-guided fractionation of the ethanol extract from the stem of Schisandra propinqua led to the isolation of four extracts. Subsequently, a neolignan 4,4-di(4-hydroxy-3-methoxyphenly)-2,3-dimethylbutanol was isolated from the EtOAc part of the stem of Schisandra propinqua, the free radical scavenging activities of which were researched in vitro. The present work demonstrated that extracts and pure compound possessed scavenging activities to DPPH, superoxide anions and hydroxy radical, and could depress lipid peroxidation reaction induced by oxygen radical produced by the Fe2+/cysteine system in vitro. This suggests that the traditional application of Schisandra propinqua in China may be related to its antioxidant activities, and the EtOAc part of the stems of Schisandra propinqua can be utilized as an effective source of antioxidants.

Efeito da desnutrição neonatal sobre o recrutamento celular e a atividade oxidante-antioxidante de macrófagos em ratos adultos endotoxêmicos / Effect of neonatal malnutrition on cell recruitment and oxidant-antioxidant activity of macrophages in endotoxemic adult rats

OBJETIVO: Avaliar se a desnutrição no período neonatal produz prejuízos no recrutamento celular para o pulmão e na atividade oxidante-antioxidante de macrófagos alveolares em ratos adultos endotoxêmicos. MÉTODOS: Ratos machos Wistar (n=48) foram alimentados por mães cuja dieta, durante a lactação, continha 23 por cento de proteína no grupo nutrido e 8 por cento no grupo desnutrido. Após o desmame todos os animais foram recuperados com dieta normoprotéica. Entre 90 e 120 dias, a metade de cada grupo foi submetida à endotoxemia por meio da administração por via intraperitonial (v.i) de lipopolissacarídio na dose de 1mg/kg de peso corporal. Após 24 horas desse procedimento coletou-se o sangue para contagem total e diferencial de leucócitos e para a dosagem de óxido nítrico. Além do sangue coletou-se também o lavado broncoalveolar para contagem total e diferencial de leucócitos e, a partir de macrófagos isolados deste lavado, foram realizadas as dosagens de superóxido, óxido nítrico e superóxido dismutase. RESULTADOS: A desnutrição acarretou um déficit ponderal que persistiu até a idade adulta, além disso, reduziu a contagem total de leucócitos sangüíneos e o número de neutrófilos após o estímulo com lipopolissacarídio. A atividade oxidante-antioxidante foi alterada havendo diminuição da produção de superóxido, óxido nítrico e superóxido dismutase antes e após a indução da endotoxemia. CONCLUSÃO: Esses resultados sugerem que a desnutrição neonatal, mesmo após a recuperação nutricional, compromete o recrutamento celular para o pulmão e a atividade oxidante-antioxidante dos macrófagos alveolares em ratos adultos. A endotoxemia contribui para evidenciar essas seqüelas da resposta do hospedeiro frente a este modelo de desnutrição.

OBJECTIVE: The objective of this study was to assess if neonatal malnutrition impairs cell recruitment to the lungs and the oxidant-antioxidant activity of alveolar macrophages in adult endotoxemic rats. METHODS: Male Wistar rats (n=48) were divided into two groups and suckled by dams fed experimental diets containing a normal protein content of 23 percent (nourished group) and a low protein content of 8 percent (undernourished group) during lactation. After weaning, all animals received a normal protein diet. Between 90 and 120 days, half of each group was submitted to endotoxemia by intraperitoneal administration of 1mg/kg of body weight of lipopolysaccharide. Blood was collected 24 hours after this procedure for total and differential leukocyte count and measurement of nitric oxide. Bronchoalveolar lavage was also done to determine total and differential leukocyte count and measure superoxide, nitric oxide and superoxide dismutase in the macrophages isolated from this lavage. RESULTS: Malnourished animals remained underweight until adulthood. Furthermore, the following also decreased: total blood leukocyte count, number of neutrophils after lipopolysaccharide administration and production of superoxide, nitric oxide and superoxide dismutase before and after induced endotoxemia. CONCLUSION: These results suggest that neonatal malnutrition, even after nutritional recovery, compromises cell recruitment to lungs and the oxidant-antioxidant activity of alveolar macrophages of adult rats. Endotoxemia contributes to evidence these sequelae to the host response before this model of malnutrition.

Cytotoxicity of catechol towards human glioblastoma cells via superoxide and reactive quinones generation

It is known that the exposure to benzene in the petroleum industry causes lympho-haematopoietic cancer among workers. However, there is little data concerning the toxicity of benzene to the central nervous system. Benzene easily penetrates the brain where it is metabolized to catechol. Since catechol autoxidizes in physiological phosphate buffer, we hypothesized that it could be toxic towards glial cells due to the generation of reactive oxygen species and quinones. In this work we studied the cytotoxic properties of catechol towards human glioblastoma cells. We found that catechol was toxic towards these cells after 72 hours and this toxicity was related to the formation of quinones. Catechol at 230µM killed 50% of cells. The catechol-induced cytotoxicity was prevented by the addition of 100U superoxide dismutase, which also inhibited the formation of quinones. These data suggest that catechol induces cytotoxicity via the extracellular generation of superoxide and quinones.

Sabe-se que a exposição de trabalhadores ao benzeno na indústria petrolífera é uma causa de câncer do sistema linfo-hematopoiético. Pouco se sabe, contudo, a respeito da toxicidade do benzeno no sistema nervoso central. O benzeno penetra facilmente no cérebro, onde é metabolizado a catecol. Como o catecol se auto-oxida em tampão fosfato no pH fisiológico, supôs-se que esse composto poderia ser tóxico para células gliais por gerar espécies reativas do oxigênio e quinonas. Nesse trabalho estudou-se a citotoxicidade do catecol para células de glioblastoma humano. O catecol foi tóxico após 72 horas e essa toxicidade relacionou-se com a formação de quinonas. O catecol a 230mM matou metade das células em cultura. A toxicidade do catecol e a produção de quinonas foram inibidas por 100U de superóxido dismutase. Esses dados sugerem que a toxicidade induzida pelo catecol deve-se à produção extracelular de superóxido e quinonas reativas.

Aderência e atividade microbicida de monócitos em portadores de esquistossomose mansônica na forma hepatoesplênica cirúrgica / Adherence and microbicidal activity of monocytes in patients with schistosomisis mansoni in surgical hepatosplenic form

A cirurgia nas crianças portadoras de esquistossomose mansônica inclui esplenectomia, ligadura da veia gástrica esquerda e o auto-implante de tecido esplênico no omento maior. A eficácia desse procedimento pode ser responsável pelo desaparecimento da sepse fulminante pós-esplenectomia (SFPE) neste tipo de paciente. Esta condição é atribuída à diminuição de IgM, de linfócitos circulantes, de properdina e ausência de tuftsina, o que conduz a deficiência da atividade das células macrófágicas, que são responsáveis pela aderência à bactéria, fagocitose e destruição das mesmas. OBJETIVO: Analisar os aspectos funcionais dos monócitos destes pacientes, operados quando crianças, no Serviço de Cirurgia Geral da Criança MÉTODOS: Foram analisados os índices de aderência in vitro dos monócitos e a geração do ânion superóxido (O2-), em três grupos. O 1§, auto-implante (AI), constituído por 18 portadores de esquistossomose mansônica na forma hepatoesplênica, submetidos a esplenectomia, ligadura da veia gástrica esquerda e auto-implante de tecido esplênico no omento maior; o 2§, (ESP), formado por nove pacientes similares, submetidos a esplenectomia e desconexão ázigo-portal, e o 3§,(CT), constituído por 12 adolescentes sadios, oriundos da mesma condição sócio-econômica-geográfica. RESULTADOS: Não houve diferença no índice de aderência entre os três grupos. Os monócitos dos pacientes do grupo AI tiveram a geração de O2- semelhante à dos indivíduos do grupo CT, e significantemente maior do que os pacientes do grupo ESP. do Hospital das Clínicas da UFPE, entre 1991 a 2000. CONCLUSÕES: Os monócitos dos portadores de esquistossomose hepatoesplênica submetidos a esplenectomia, ligadura da veia gástrica esquerda e auto-implante de tecido esplênico no omento maior se mostram funcionalmente similares aos de indivíduos normais da mesma condição sócio-econômica-geográfica.

Effects of Hg and Cu on hemocytes-mediated functions in the prawn, Macrobrachium malcolmsonii.

Sublethal effects of Hg (9.6, 14.5 and 24.1 microg/l) and Cu (60.5, 90.8 and 150.5 microg/l) on the hemocytes-mediated functions in the juveniles of the economically important freshwater prawns, Macrobrachium malcolmsonii were investigated. The population of total hemocytes, percentile phagocytosis and superoxide anion production were found to be increased in test prawns exposed to the lowest sublethal concentration of these two metals in comparison to controls. This indicates the fact that a mechanism of host-defence was in an active state to encounter metal toxicity. The total hemocytes population, percentile phagocytosis and superoxide anion production were found to be decreased in test prawns exposed to intermediate as well as the highest sublethal concentrations of Hg and Cu when compared with controls. This suggests that high concentrations of both Hg and Cu have inhibitory effects on the immune functions of the hemocytes in the prawn, M. malcolmsonii.

A microtiter plate assay for superoxide using MTT reduction method.

A simple, rapid and sensitive microtiter plate assay for superoxide using the reduction of tetrazolium dye MTT to its coloured formazan has been developed. The colour formed can be measured using a microtiter plate reader and the extent of reduction of MTT indicates the amount of superoxide generation. A comparison of the sensitivities of different procedures for the quantitation of superoxide generated by X-XO system has been made. The MTT reduction due to superoxide was confirmed by inhibiting the reduction using purified superoxide dismutase. Using this method superoxide generation by mitochondria and microsomes was demonstrated and this procedure is suitable for detection of intracellularly generated superoxide. The proposed method is inexpensive and is suitable for a routine analysis of large number of samples.

Papel do PMN na falencia de multiplos orgaos e sistemas / Role of the polymorphonuclear neutrophil in the failure of multiple organs and systems

Os autores analisam o papel do neutrofilo polimorfonuclear (PMN) na FMOS pos-trauma e cirurgia, papel este atribuido quer a ativacao para producao de radicais superoxidos e enzimas, quer a depressao funcional dos PMNs. Destacam: 1) a investigacao da sequencia de transmissao intracelular de sinais entre receptores de membrana do PMN e a resposta efetora final; 2) a estrutura e funcao do sistema NADPH do PMN; 3) os estados funcionais do PMN (quiescente, sensibilizado, ativado ou responsivo) em termos de ativacao do sistema NADPH; 4) o mecanismo de lesao tecidual pelo PMN. Discutem investigacoes clinicas sobre o estado de ativacao do PMN, e abordam as perspectivas terapeuticas apontadas pelas pesquisas recentes

Produccion del anion superoxido en polimorfonucleares neutrofilos / Production of superoxide anion in polymorphonuclear neutrophils

La estandarizacion de tecnicas que permitan el analisis nuclear de los polimorfonucleares neutrofilos es importante para determinar, en un momento dado, la capacidad de defensa de un individuo. Uno de los parametros susceptibles de medicion y que se correlaciona positivamente con la capacidad funcional del polimorfonuclear, es la produccion de agentes oxidantes. Especificamente, el anion superoxido es una especie reactiva del oxigeno que se genera durante la explosion respiratoria y sirve como medida de la misma. El anion superoxido puede ser determinado a traves de una reaccion colorimetrica que utiliza el citocromo C como substrato. Este trabajo presenta los resultados de la estandarizacion de un nuevo metodo para la deteccion del anion superoxido con el cual se ahorran significativamente tiempo, reactivos y celulas. Con esta prueba encontramos, en 26 individuos normales, que no hay diferencias en la produccion del anion superoxido en los grupos por edades ni por sexo y el promedio de valores para las celulas en reposo y para las celulas estimuladas fue de 0.81 mas o menos 0.53 y 12.28 mas o menos 0.87 manomoles/10 6 polimorfonucleares/ 30 minutos, respectivamente.