A Preliminary Research Study on the Cytotoxicity of Expired and Non-expired Composite Resins: In Vitro Study / Estudio preliminar in vitro de la citotoxicidad de resinas compuestas caducadas y no caducadas

Astracts Objective: Studies have focused on use of non-expired composites. Unfortunately some clinicians still use expired composite resins without considering their effects. The objective of this in vitro preliminary research was to investigate cytotoxicity of expired(6-months) and non-expired composite resins. Materials and methods: Expired (E) and non-expired (NE) samples of one bulk-fill (Tetric N-Ceram Bulk-fill (TNB), Ivoclar Vivadent), two nano-hybrid (Tetric N-Ceram (TN), Ivoclar Vivadent; Clearfil Majesty ES-2 (CM), Kuraray) composite resins were tested on L929 fibroblast cells. Medium covering cells was removed then plastic rings (2-mm height) were filled with non-polymerized composite resins, placed in direct contact with cells and polymerized with LED light curing unit (LCU). Three samples were prepared for each group. After polymerization, removed medium was added to the cells. Cells that were left without medium (WOM) and cells that were exposed to LCU were used as positive control groups. Cells without any treatment were used as negative control group (C). Cells were incubated with tested materials for 7-days to evaluate cytotoxicity. Cell viability was calculated by sulforhodamine B test as a percentage (%). One-way ANOVA and post-hoc Tukey tests were used for statistical analyses (p0.05), except between TN NE and TN E (p0.05). All experimental groups compared with C group showed statistically significant cytotoxicity (p<0.05). A statistically significant difference existed between LCU and C groups (p<0.05). Conclusions: In clinical practice, expired composite resins should never be used. Although a correlation was found between expiration dates of nano-hybrid composite resins and cell viability, opposite data were obtained for bulk- fill composite resin. Researches are still required to evaluate biocompatibility of bulk- fill composite resins at various thicknesses with current LCUs.

Resumen Objetivo: Los estudios se han concentrado en el uso de resinas compuestas no vencidos. Desafortunadamente, algunos clínicos aún usan resinas caducadas sin considerar sus efectos. El objetivo de este estudio preliminar in vitro fue investigar la citotoxicidad de resinas compuestas caducadas (6 meses) y no caducadas. Materiales y métodos: muestras caducadas (E) y no caducadas (NE) de una resina bulk-fill (Tetric N-Ceram Bulk-fill (TNB), Ivoclar Vivadent) y dos resinas nanohíbridas (Tetric N-Ceram (TN) Ivoclar Vivadent) (Clearfil Majesty ES-2 (CM), Kuraray), se probaron en células de fibroblastos L929. Se retiraron las células que cubrían el medio, luego se llenaron anillos de plástico (2 mm de altura) con resinas no polimerizadas, se colocaron en contacto directo con las células y se polimerizaron con una unidad de fotocurado LED (LCU). Se prepararon tres muestras para cada grupo. Después de la polimerización, se añadió el medio eliminado a las células. Las células que quedaron sin medio (WOM) y las células que se expusieron a LCU se usaron como grupos de control positivo. Las células sin ningún tratamiento se utilizaron como grupo de control negativo (C). Las células se incubaron con las resinas durante 7 días para evaluar la citotoxicidad. La viabilidad celular se calculó mediante la prueba de sulforodamina B como un porcentaje (%). ANOVA unidireccional y pruebas post-hoc de Tukey se utilizaron para los análisis estadísticos (p 0.05), excepto entre TN NE y TN E (p 0.05). Todos los grupos experimentales en comparación con el grupo C mostraron citotoxicidad estadísticamente significativa (p <0,05). Existió una diferencia estadísticamente significativa entre LCU y grupos C (p <0.05). Conclusiones: En la práctica clínica, las resinas compuestas caducadas nunca deben usarse. Aunque se encontró una correlación entre las fechas de vencimiento de las resinas compuestas nano-híbridas y la viabilidad celular, se obtuvieron datos opuestos para la resina bulk-fill. Se requieren nuevas investigaciones para evaluar la biocompatibilidad de las resinas bulk-fill en distintos espesores con las LCU actuales.

Protective Effect of Ischemic Preconditioning on Myocardium Against Remote Tissue Injury Following Transient Focal Cerebral Ischemia in Diabetic Rats / Efeito Protetor do Pré-Condicionamento Isquêmico no Miocárdio Contra Lesão Remota de Tecidos após Isquemia Cerebral Focal Transitória em Ratos Diabéticos

Abstract Background: Remote ischemic preconditioning (IPreC) could provide tissue-protective effect at a remote site by anti-inflammatory, neuronal, and humoral signaling pathways. Objectives: The aim of the study was to investigate the possible protective effects of remote IPreC on myocardium after transient middle cerebral artery occlusion (MCAo) in streptozotocin- induced diabetic (STZ) and non-diabetic rats. Methods: 48 male Spraque Dawley rats were divided into eight groups: Sham, STZ, IPreC, MCAo, IPreC+MCAo, STZ+IPreC, STZ+MCAo and STZ+IPreC+MCAo groups. We induced transient MCAo seven days after STZ-induced diabetes, and performed IPreC 72 hours before transient MCAo. Remote myocardial injury was investigated histopathologically. Bax, Bcl2 and caspase-3 protein levels were measured by Western blot analysis. Total antioxidant status (TAS), total oxidant status (TOS) of myocardial tissue were measured by colorimetric assay. Oxidative stress index(OSI) was calculated as TOS-to-TAS ratio. For all statistical analysis, p values < 0.05 were considered significant. Results: We observed serious damage including necrosis, congestion and mononuclear cell infiltration in myocardial tissue of the diabetic and ischemic groups. In these groups TOS and OSI levels were significantly higher; TAS levels were lower than those of IPreC related groups (p < 0.05). IPreC had markedly improved histopathological alterations and increased TAS levels in IPreC+MCAo and STZ+IPreC+MCAo compared to MCAo and STZ+MCAo groups (p < 0.05). In non-diabetic rats, MCAo activated apoptotic cell death via increasing Bax/Bcl2 ratio and caspase-3 levels. IPreC reduced apoptotic cell death by suppressing pro-apoptotic proteins. Diabetes markedly increased apoptotic protein levels and the effect did not reversed by IPreC. Conclusions: We could suggest that IPreC attenuates myocardial injury via ameliorating histological findings, activating antioxidant mechanisms, and inducing antiapoptotic activity in diabetic rats.

Resumo Fundamentos: O pré-condicionamento isquêmico remoto (IPreC) poderia fornecer efeito protetor de tecido em um local remoto por vias de sinalização anti-inflamatórias, neuronais e humorais. Objetivos: O objetivo do estudo foi investigar os possíveis efeitos protetores do IPreC remoto no miocárdio após a oclusão transitória da artéria cerebral média (MCAo) em ratos com diabetes induzida por estreptozotocina (STZ) e ratos não diabéticos. Métodos: 48 ratos Spraque Dawley machos foram divididos em oito grupos: grupos Sham, STZ, IPreC, MCAo, IPreC + MCAo, STZ + IPreC, STZ + MCAo e STZ + IPreC + MCAo. Induzimos MCAo sete dias após a diabetes induzida por STZ e realizamos IPreC 72 horas antes do MCAo. A lesão miocárdica remota foi investigada histopatologicamente. Os níveis de proteína Bax, Bcl2 e caspase-3 foram medidos pela análise Western Blot. O estado de antioxidante total (TAS), e o estado de oxidação total (TOS) do tecido miocárdico foram medidos por meio de um estudo colorimétrico. O índice de estresse oxidativo (OSI) foi calculado como a relação TOS-TAS. Para todas as análises estatísticas, os valores de p < 0,05 foram considerados significativos. Resultados: Observamos danos graves, incluindo necrose, congestão e infiltração de células mononucleares no tecido miocárdico dos grupos diabético e isquêmico. Nesses grupos os níveis de TOS e OSI foram significativamente maiores; os níveis de TAS foram inferiores aos dos grupos relacionados com IPreC (p < 0,05). O IPreC melhorou marcadamente as alterações histopatológicas e aumentou os níveis de TAS em IPreC + MCAo e STZ + IPreC + MCAo em comparação com os grupos MCAo e STZ + MCAo (p < 0,05). Em ratos não diabéticos, MCAo activou a morte celular apoptótica através do aumento da relação Bax / Bcl2 e dos níveis de caspase-3. IPreC reduziu a morte celular apoptótica pela supressão de proteínas pró-apoptóticas. O diabetes aumentou acentuadamente os níveis de proteína apoptótica e o efeito não foi revertido pelo IPreC. Conclusões: Podemos sugerir que o IPreC atenua a lesão miocárdica através da melhora dos achados histológicos, ativando mecanismos antioxidantes e induzindo atividade antiapoptótica em ratos diabéticos.

Cytotoxic, genotoxic and apoptotic effects of naringenin-oxime relative to naringenin on normal and cancer cell lines

Objective: To assess and compare the cytotoxic, genotoxic, apoptotic and reactive oxygen species (ROS) generating effects of naringenin (NG) and its new derived compound naringenin-oxime (NG-Ox) on MCF-7, HT-29, PC-12 cancer and L-929 normal cell lines. Methods: The cells were incubated with different doses of NG-Ox and NG (50–1 000 mmol/L) for 24 h. The cell viability was assessed based on ATP cell viability assay. Intracellular accumulation of ROS was determined using the fluorescent probes 2070-dichlorodihydrofluorescin diacetate. Genotoxic effects were evaluated by alkaline single cell gel electrophoresis assay (comet assay) and, the apoptotic effect was evaluated by acridine orange staining at below the IC50 levels. Results: Both NG-Ox and NG exhibited cytotoxic, genotoxic and apoptotic effects and resulted in increased ROS values in a dose-dependent manner. The effects were more pronounced on cancer cell lines. The cytotoxic, genotoxic and apoptotic effects of NG-Ox were higher than that of NG in all cell lines. Significant correlations were observed be-tween cell viability, DNA damage, apoptosis and ROS, in all cell lines exposed to either NG-Ox or NG. Conclusions: This study showed that both NG-Ox and NG possess cytotoxic, genotoxic and apoptotic activities through the production of ROS on cells, NG-Ox being the more effective one. Therefore, derived compound of NG might be used as antiproliferative agents for the treatment of cancer.

DNA Damage in Children Exposed to Secondhand Cigarette Smoke and its Association with Oxidative Stress.

Objective: To compare oxidative status, total antioxidant capacity and values of DNA damage in peripheral blood lymphocytes in children exposed to secondhand cigarette smoke with healthy controls. Design: Analytical, Observational. Participants: 54 children without any chronic diseases, attending the healthy child monitoring polyclinic. These comprised 27 children who had been exposed to passive cigarette smoke and 27 children who had not been exposed to cigarette smoke. Main Outcome Measures: Urine cotinine levels by the chemiluminescent technique; DNA damage by alkaline comet assay; and total oxidant status (TOS) using a novel automated measurement method. Results: The mean urine cotinine, TOS, Oxidative Stress Index (OSI) and DNA damage values of the group exposed to cigarette smoke were determined to be at significantly higher level compared to the group not exposed to cigarette smoke (P<0.001). No statistically significant difference was determined in the TAS level between the two groups (P=0.1) Conclusions: The results showed that TOS levels, OSI index and DNA damage in peripheral blood lymphocytes were significantly higher in children exposed to secondhand cigarette smoke than in those not exposed to secondhand cigarette smoke.

Aumento do estresse oxidativo em pré-escolares expostos ao tabagismo passivo / Increased oxidative stress in preschool children exposed to passive smoking

OBJETIVOS: Estudar o efeito do fumo passivo sobre o estado plasmático oxidativo e antioxidativo em pré-escolares fumantes passivos e compará-los com controles. MÉTODOS: Trinta e quatro pré-escolares fumantes passivos (cinco a 50 cigarros/dia) (grupo de estudo) e 32 controles que nunca estiveram expostos à fumaça de cigarro foram escolhidos aleatoriamente entre crianças de 4 a 6 anos. Foram determinados os níveis de cotinina urinária e de indicadores do estado oxidativo e antioxidativo, isto é, estado oxidante total (EOT), capacidade antioxidante total (CAT) e índice de estresse oxidativo (IEO). RESULTADOS: A média do consumo ambiental de cigarros foi de 22±13 cigarros por dia nas crianças fumantes passivas. Os níveis médios de cotinina urinária foram 77,6±41,4 ng/mL e 11,9±2,3 ng/mL nos grupos de estudo e controle, respectivamente (p < 0,001). Os níveis médios da CAT plasmática foram 0,95±0,13 mmol equivalente de Trolox/L e 1,01±0,09 mmol equivalente de Trolox/L, respectivamente (p = 0,039). Os níveis médios de EOT plasmático foram 28,6±7,9 µmol H2O2 equivalente/L e 18,5±6,3 µmol H2O2 equivalente/L, respectivamente (p < 0,001). Os níveis médios de IEO foram 3,08±0,98 unidade arbitrária e 1,84±0,64 unidade arbitrária, respectivamente (p < 0,001). Uma pequena quantidade de fumaça de cigarro (cinco a 10 cigarros/dia) causa estresse oxidativo considerável. Não houve correlações significativas entre o número de cigarros consumidos e os níveis de estado oxidante e de IEO. CONCLUSÕES: O tabagismo passivo é um potente oxidante em pré-escolares. Seus efeitos deletérios não se limitam apenas tabagismo passivo pesado, mas também ocorrem com a exposição a pequenas quantidades de fumaça.

OBJECTIVES: To study the effect of passive cigarette smoking on plasma oxidative and antioxidative status in passive smoking preschool children and to compare them with controls. METHODS: Thirty-four passive smoking (five to 50 cigarettes per day) preschool children (study group) and 32 controls who had never been exposed to cigarette smoke were randomly chosen from children aged from 4 to 6 years. Urinary cotinine and plasma indicators of oxidative and antioxidative status, i.e., total oxidant status (TOS), total antioxidant capacity (TAC), and oxidative stress index (OSI), were determined. RESULTS: Mean environmental cigarette consumption was 22±13 cigarettes per day in passive smoking children. Mean urinary cotinine levels were 77.6±41.4 ng/mL and 11.9±2.3 ng/mL in the study and control groups, respectively (p < 0.001). Mean plasma TAC levels were 0.95±0.13 mmol Trolox equivalent/L and 1.01±0.09 mmol Trolox equivalent/L, respectively (p = 0.039). Mean plasma TOS levels were 28.6±7.9 µmol H2O2 equivalent/L and 18.5±6.3 µmol H2O2 equivalent/L, respectively (p < 0.001). Mean OSI levels were 3.08±0.98 arbitrary units and 1.84±0.64 arbitrary units, respectively (p < 0.001). A small amount of cigarette smoke (five to 10 cigarettes per day) causes considerable oxidative stress. There were significant correlations between number of cigarettes consumed and oxidant status and OSI levels. CONCLUSIONS: Passive smoke is a potent oxidant in preschool children. Its deleterious effects are not limited just to heavy passive smoking, but also occur with exposure to small amounts of smoke.

Serum copper and zinc levels in mothers and cord blood of their newborn infants with neural tube defects: A case-control study.

Objectives: To measure the cord blood and maternal serum levels of folic acid, vitamin B12, zinc, copper, selenium and lead in infants born with neural tube defect (NTD), and to examine a possible relationship between the nutriture of these micronutrients and occurrence of neural tube defect. Design: Case-control study. Methods: Maternal serum and cord blood samples were obtained at delivery from 70 healthy mothers and 74 mothers who had a newborn with NTD. Results: The mean (± SD) maternal serum zinc level in the NTD group was significantly lower than that of the control group (835.6 µg/L ±333.8 µg/L vs. 1035.7 µg/L ± 299.8 µg/L, P=0.004, respectively). The mean maternal and cord serum copper levels in the NTD group were significantly higher when compared to the control group (2831.1 µg/L ± 1017 µg/L vs. 2402 µg/L ± 744.2 µg/L; P=0.03; and 789.8 µg/L vs 517.2 µg/L, P<0.001, respectively). There was a negative correlation between the cord levels of folic acid and copper in the NTD group with the respective maternal serum levels (r=-0.289; P=0.018). Conclusions: High maternal serum levels of copper and lower level of zinc during pregnancy associated with NTD in newborn.

Fototerapia causa danos ao DNA de leucócitos mononucleares periféricos em recém-nascidos a termo / Phototherapy causes DNA damage in peripheral mononuclear leukocytes in term infants

OBJETIVO: Determinar se a fita de DNA de leucócitos mononucleares endógenos é alvo de fototerapia. MÉTODOS: O estudo incluiu 65 recém-nascidos a termo com idades entre 3 e 10 dias que haviam sido expostos a fototerapia intensiva (n = 23) ou convencional (n = 23) por pelo menos 48 horas devido à icterícia neonatal, além de um grupo controle (n = 19). Dano ao DNA foi avaliado por eletroforese alcalina em gel de célula única (ensaio cometa). A capacidade antioxidante total plasmática e os níveis de estado oxidativo total também foram medidos, e a correlação entre danos ao DNA e estresse oxidativo foi investigada. RESULTADOS: Os valores médios de escores de danos ao DNA nos grupos de fototerapia intensiva e convencional foram significativamente maiores do que os do grupo controle (p < 0,001). Os valores médios e desvio padrão foram 32 (9), 28 (9), 21 (7) unidades arbitrárias, respectivamente. Os níveis de estado oxidativo nos grupos de fototerapia intensiva e convencional foram significativamente maiores do que os do grupo controle (p < 0,005). Os valores médios (desvio padrão) foram 18, 1 (4,2), 16.9 (4,4), 13,5 (4,2) µmol H2O2 equivalente/L, respectivamente. De maneira semelhante, os níveis de estresse oxidativo nos grupos de fototerapia intensiva e convencional foram significativamente maiores do que os do grupo controle (p = 0,041). A capacidade antioxidante total plasmática e os níveis de bilirrubina total não diferiram entre os grupos (p > 0,05). Não houve correlações significativas entre escores de danos ao DNA e bilirrubina, estado oxidante total e níveis de estresse oxidativo entre os grupos de fototerapia (p > 0,05). CONCLUSÕES: Tanto a fototerapia intensiva quanto a convencional causam danos ao DNA dos leucócitos mononucleares endógenos em recém-nascidos a termo com icterícia.

OBJECTIVE: Our aim was to determine whether endogenous mononuclear leukocyte DNA strand is a target of phototherapy. METHODS: The study included 65 term infants aged between 3-10 days that had been exposed to intensive (n = 23) or conventional (n = 23) phototherapy for at least 48 hours due to neonatal jaundice, and a control group (n = 19). DNA damage was assayed by single-cell alkaline gel electrophoresis (comet assay). Plasma total antioxidant capacity and total oxidant status levels were also measured, and correlation between DNA damage and oxidative stress was investigated. RESULTS: Mean values of DNA damage scores in both the intensive and conventional phototherapy groups were significantly higher than those in the control group (p < 0.001). Mean values and standard deviation were 32 (9), 28 (9), 21 (7) arbitrary unit, respectively. Total oxidant status levels in both the intensive and conventional phototherapy groups were significantly higher than those in the control group (p = 0.005). Mean (standard deviation) values were 18.1 (4.2), 16.9 (4.4), 13.5 (4.2) µmol H2O2 equivalent/L, respectively. Similarly, oxidative stress index levels in both the intensive and conventional phototherapy groups were significantly higher than those in the control group (p = 0.041). Plasma total antioxidant capacity and total bilirubin levels did not differ between the groups (p > 0.05). There were no significant correlations between DNA damage scores and bilirubin, total oxidant status and oxidative stress levels in either phototherapy group (p > 0.05). CONCLUSIONS: Both conventional phototherapy and intensive phototherapy cause endogenous mononuclear leukocyte DNA damage in jaundiced term infants.

Adenosine deaminase activities in sera, lymphocytes and granulocytes in patients with cutaneous leishmaniasis

Adenosine deaminase (ADA) activities in sera, lymphocytes and granulocytes in patients with cutaneous leishmaniasis were investigated and compared with control groups. Fifty patients and 50 healthy individuals were studied. The clinical diagnosis was parasitologically confirmed by culture and Giemsa stain. ADA activities were measured by colorimetric method. Serum ADA activities 37.80 ñ 11.90, 18.28 ñ 6.08 IU/L (p<0.0001), lymphocyte specific ADA activities 14.90 ñ 7.42, 8.38 ñ 7.42 U/mg protein (p=0.04), granulocyte specific ADA activities 1.15 ñ 0.73, 1.09 ñ 0.67 U/mg protein (p>0.05) were found in patients and control groups, respectively. ADA activity increases in some infectious diseases were cell mediated immune mechanisms are dominant. In cutaneous leishmaniasis, lymphokine-mediated macrophage activity is the main effector mechanism. Increase in serum and lymphocyte ADA activities in patients with cutaneous leishmaniasis may be dependent on and reflects the increase in phagocytic activity of macrophages and maturation of T-lymphocytes.