In Vitro effect of low-level laser therapy on typical oral microbial biofilms

The aim of this study was to evaluate the effect of specific parameters of low-level laser therapy (LLLT) on biofilms formed by Streptococcus mutans, Candida albicans or an association of both species. Single and dual-species biofilms - SSB and DSB - were exposed to laser doses of 5, 10 or 20 J/cm2 from a near infrared InGaAsP diode laser prototype (LASERTable; 780 ± 3 nm, 0.04 W). After irradiation, the analysis of biobilm viability (MTT assay), biofilm growth (cfu/mL) and cell morphology (SEM) showed that LLLT reduced cell viability as well as the growth of biofilms. The response of S. mutans (SSB) to irradiation was similar for all laser doses and the biofilm growth was dose dependent. However, when associated with C. albicans (DSB), S. mutans was resistant to LLLT. For C. albicans, the association with S. mutans (DSB) caused a significant decrease in biofilm growth in a dose-dependent fashion. The morphology of the microorganisms in the SSB was not altered by LLLT, while the association of microbial species (DSB) promoted a reduction in the formation of C. albicans hyphae. LLLT had an inhibitory effect on the microorganisms, and this capacity can be altered according to the interactions between different microbial species.

O objetivo deste estudo foi avaliar o efeito de parâmetros específicos de irradiação com laser de baixa intensidade sobre biofilmes formados por Streptococcus mutans (S. mutans), Candida albicans (C. albicans) ou associação de ambas as espécies. Biofilmes isolados ou associados destes microrganismos foram irradiados com um dispositivo laser infra-vermelho próximo de diodos InGaAsP (LaserTABLE 780 ±3 nm, 0,04W), utilizando-se para isto o dispositivo LASERTable. Quinze horas após a irradiação, foi demonstrado, por meio da avaliação da viabilidade celular (Teste de MTT), da morfologia das células (MEV) e do crescimento do biofilme (UFC/mL), que esta terapia foi capaz de reduzir o metabolismo celular, número de microrganismos presentes no biofilme, bem como seu crescimento no local. Quanto à viabilidade celular, a resposta à irradiação do biofilme de S. mutans (SSB) foi semelhante para todas as doses de energia, sendo que o crescimento do biofilme foi dose dependente. Porém, quando associado à C. albicans, este microrganismo apresentou resistência à fototerapia. Já a C. albicans associada ao S. mutans apresentou redução de crescimento significativa, sendo este resultado também foi dose dependente. A morfologia dos microrganismos não foi alterada pelas irradiações realizadas quando em biofilmes isolados. A associação entre os microrganismos promoveu redução na formação de hifas pela C. albicans. A laserterapia de baixa intensidade apresentou efeito inibitório sobre microrganismos, sendo que esta capacidade pode ser alterada de acordo com a interação entre diferentes microrganismos.

Cytotoxic effects of White-MTA and MTA-Bio cements on odontoblast-like cells (MDPC-23)

This study evaluated the cytotoxic effects of 2 mineral trioxide aggregate (MTA) cements - White-MTA-Angelus and a new formulation, MTA-Bio - on odontoblast-like cell (MDPC-23) cultures. Twenty-four disc-shaped (2 mm diameter x 2 mm thick) specimens were fabricated from each material and immersed individually in wells containing 1 mL of DMEM culture medium for either 24 h or 7 days to obtain extracts, giving rise to 4 groups of 12 specimens each: G1 - White-MTA/24 h; G2 - White-MTA/7 days; G3 - MTA-Bio/24 h; and G4 - MTA-Bio/7 days. Plain culture medium (DMEM) was used as a negative control (G5). Cells at 30,000 cells/cm² concentration were seeded in the wells of 24-well plates and incubated in a humidified incubator with 5 percent CO2 and 95 percent air at 37ºC for 72 h. After this period, the culture medium of each well was replaced by 1 mL of extract (or plain DMEM in the control group) and the cells were incubated for additional 2 h. Cell metabolism was evaluated by the MTT assay and the data were analyzed statistically by ANOVA and Tukey's test (α=0.05). Cell morphology and the surface of representative MTA specimens of each group were examined by scanning electron microscopy. There was no statistically significant difference (p>0.05) between G1 and G2 or between G3 and G4. No significant difference (p>0.05) was found between the experimental and control groups either. Similar cell organization and morphology were observed in all groups, regardless of the storage periods. However, the number of cells observed in the experimental groups decreased compared to the control group. MTA-Bio presented irregular surface with more porosities than White-MTA. In conclusion, White-MTA and MTA-Bio presented low cytotoxic effects on odontoblast-like cell (MDPC-23) cultures.

Este estudo avaliou o efeito citotóxico de dois cimentos MTA - MTA Branco-Angelus e uma nova formulação, MTA-Bio - sobre células odontoblastóides (MDPC-23) mantidas em cultura. Vinte e quatro espécimes padronizados (2 mm diâmetro x 2 mm largura) foram confeccionados de cada material e imersos individualmente em compartimentos contendo 1 mL de meio de cultura DMEM por 24 h ou 7 dias para obtenção dos extratos, formando 4 grupos de 12 espécimes cada: G1 - MTA-Branco/24 h; G2 - MTA-Branco/7 dias; G3 - MTA-Bio/24 h; e G4 - MTA-Bio/7 dias. Meio de cultura puro (DMEM) foi utilizado como controle negativo (G5). Células na concentração de 30.000 células/cm² foram semeadas nas placas de 24 compartimentos e incubadas em incubadora com 5 por cento CO2 e 95 por cento ar a 37ºC por 72 h. Após esse período, o meio de cultura de cada compartimento foi substituído por 1 mL do extrato (ou DMEM puro no grupo controle) e as células foram incubadas pelo período adicional de 2 h. O metabolismo celular foi avaliado pelo teste do MTT e os dados foram analisados estatisticamente pelo teste de ANOVA e Tukey (α=0,05). A morfologia celular e da superfície dos espécimes de MTA representativos de cada grupo foram avaliados em microscopia eletrônica de varredura. Não houve diferença estatisticamente significante (p>0,05) entre os gurpos G1 e G2 ou entre G3 e G4. Não foi encontrada diferença estatística (p>0,05) entre os grupos experimentais e controle. Morfologia e organização celular semelhante foram observadas em todos os grupos, independente do período de extração. Entretanto, o número de células observado nos grupos experimentais diminui quando comparado ao grupo controle. MTA-Bio apresentou uma superfície irregular com mais porosidades que o MTA-Branco. Pode-se concluir que os cimentos MTA-Branco e MTA-Bio apresentaram reduzido efeito citotóxico sobre células odontoblastóides (MDPC-23) mantidas em cultura.

Application of electron spin resonance for evaluation of the level of free radicals in the myometrium in full-term pregnancy with normal labour and uterine inertia.

In order to identify and quantify free radicals in the tissues of patients with normal physiological and pathological states of births, we developed a method to evaluate the amount of free radicals in myometrium of subplacental area and from body of uterus, using electron spin resonance spectroscopy. Analysis of the concentration of free radicals in the myometrium in full-term pregnancy with normal labour and during uterine inertia was studied. The activities of Ca2+-ATPase, cytochrome c oxidase and succinate dehydrogenase in samples of these tissues were tested too. Low free radical concentrations in these tissues were associated with disturbances in contractile activity of myometrium along with reduction of Ca2+-ATPase, cytochrome c oxidase and succinate dehydrogenase activity. There proved to be an association between the level of free radicals in the tissues and alteration in the physiological processes.

Effect of thyroidectomy on fast and slow muscle fibres of rat gastrocnemius muscle.

A combined histochemical, biochemical and electrophoretic study with respect to the enzymes succnic dehydrogenase(SDH), myofibrillar adenosine triphosphatase (m-ATPase), lactate dehydrogenase (LDH) isozymes and myosin light chains was carried out to investigate the response of rat gastrocnemius muscle (medial head). Twelve weeks after thyroidectomy, the results indicated a shift from fast to slow type pattern of LDH isozymes, fibre type transformation from Type II to Type I and a decrease in SDH and m-ATPase activity. The results suggest, possible thyroidal involvement in determining the phenotypic properties of skeletal muscle.

Regenerative ability of gastrocnemius muscle under diabetic condition with special reference to SDH & m-ATPase.

The present study analyzes the regeneration of skeletal muscle in streptozotocin (STZ) induced diabetic rats. Two weeks later the gastrocnemius muscle from diabetic rats were transplanted into diabetic and normal host to initiate regeneration and the normal gastrocnemius muscle was transplanted in normal and diabetic hosts for comparison. The regenerates were analyzed after 15 and 30 days of transplantation for histochemical (with respect to SDH and m-ATPase) and 7, 14, 21 and 28 days after transplantation for biochemical studies (with respect to SDH and m-ATPase). Least enzymatic activity and the poorest regenerative ability in case of normal muscle in diabetic host (NM-DbH) and comparatively higher enzymatic activity and better regenerative ability in diabetic muscle in normal host (DbM-NH) was observed. The result of this study strongly supports that the normal host environment is crucial for the muscle recovery.

Caracterizaçäo de miopatias mitocondriais através da avaliaçäo das atividades enzimáticas envolvidas no metabolismo energético / Characterization of mitochondrial myopathies through the evaluation of the enzymatic activities involved in the energetic metabolism

Foi determinada a atividade das enzimas NADH desidrogenase, NADH citocromo e redutase, succinato desidrogenase, succinato citocromo e redutase, citocromo e oxidase e citrato sintase em mitocôndrias de músculo esquelético humano normal e doente (suspeito de miopatia mitocondrial). O grupo controle foi constituído de 13 indivíduos normais e que nao faziam uso contínuo de fármacos. O grupo doente era constituído de 10 pacientes cujo diagnóstico anatomopatológico indicava suspeita de miopatia mitocondrial. Observou-se reduçao na atividade das enzimas em todos os pacientes: 7 com anormalidades em todas as enzimas ensaiadas; 2 com deficiências em todas as enzimas exceto na citocromo e oxidase; e 1 paciente com disfunçao apenas na atividade da succinato desidrogenase e succinato citocromo e redutase. Este perfil possibilitou caracterizar múltiplas deficiências ou deficiência combinada da cadeia respiratória, além da disfunçao na citrato sintase em 9 pacientes. Um dos casos constituiu exceçao, sendo a deficiência enzimática restrita ao complexo II. Foi possível concluir que a metodologia usada é adequada e facilmente aplicável aos objetivos clínicos. Os resultados obtidos possibilitam a caracterizaçao dos complexos enzimáticos mitocondriais deficientes, mostrando que tais enfermidades sao originadas de disfunçao no metabolismo energético.

Metabolic studies on mycobacteria-I. Demonstration of key enzymes of glycolysis and tricarboxylic acid cycle on polyacrylamide gels.

Polyacrylamide gel electrophoresis (PAGE) technique was standardised to demonstrate some key enzymes of glycolysis, hexose mono phosphate (HMP) pathway and tricarboxylic acid cycle in slow growing mycobacteria (M. avium. M. gastri) as well as in fast growing mycobacteria (M. vaccae, M. phlei). The enzymes studied were lactate dehydrogenase (LDH) glucose-6-phosphate dehydrogenase (G6PD), aconitase, isocitrate dehydrogenase (ICD), succinic dehydrogenase (SDH), fumerase and malate dehydrogenase (MDH). All the three pathways were found to be operative in slow as well as fast growing mycobacteria. Using this technique M. leprae specific MDH activity was demonstrated in the cell free extract of M. leprae. It's (MDH) electrophoretic mobility on gels lies in the range shown by other mycobacterial species studied and was distinct from that of host MDH. It appears that PAGE offers a useful tool for metabolic characterization of M. leprae using infected tissues.