Analysis of the effect of photodynamic therapy with Fotoenticine on gliosarcoma cells.

Gliosarcoma is a highly aggressive malignant neoplasm and a histopathological variant of wild-type glioblastoma multiforme isocitrate dehydrogenase (HDI). The current standard treatment consists of chemotherapy, radiotherapy and surgical resection, however, despite advances in these techniques, the patient's prognosis remains unfavorable. Photodynamic therapy (PDT) is a noninvasive technique that has been highlighted as an alternative form of cancer treatment because it does not present the side effects associated with systemic treatments. The objective of this study was to evaluate the cell viability and the intracellular localization of photosensitizer (PS) chlorin e6 Fotoenticine in 9L/lacZ cells. Therefore, tests of cytotoxicity, morphology, and location of PS were performed. The viability test showed no cytotoxicity in the dark at all concentrations and 100 % cell death at the highest concentrations after PDT. The mitochondrial activity test showed a reduction in all groups after PDT. The production of reactive oxygen species (ROS) was higher in the PDT groups and dependent on the PS concentration. Morphological analysis after PDT showed apparent cytoplasmic destruction in all the tested concentrations, with the presence of rounded cells due to the loss of their extensions and absence of nuclear alterations. The PS accumulation in the mitochondria and cytoskeleton was observed by the confocal microscopy; however, there is no evidence of its internalization in the lysosomes. It was concluded that PDT with Fotoenticine is a promising alternative therapy showing decreased cell viability, increased ROS production and adequate localization to trigger cell death.

The effect of low-level laser therapy (660 nm) on the gene expression involved in tissue repair.

The effect of low-level laser therapy (LLLT) on the healing of skin lesions has been evaluated in many studies; however, the molecular mechanisms involved in the biostimulatory effects resulting from this treatment need to be better understood. The paper aims to analyze the effects of LLLT (660 nm) at doses of 1 and 5 J/cm2 on cell viability and expression of vascular endothelial growth factor (VEGF) and interleukin (IL6) genes in L929 fibroblast cells. The dose-response curve was performed with the GaInAlAs (660 nm) laser-treated cells at energy rates of 1 and 5 J/cm2. Cell viability was quantified at 24, 48, and 72 h after irradiation and the effects of TLBP on the cytoskeleton and endoplasmic reticulum were evaluated by fluorescence microscopy and the RT-qPCR method was used for the analysis of gene expression. It was observed that the 72 h group had a statistically significant increase in cell viability compared to the 48 h group (p < 0.01) and when compared to the 72 h control (p = 0.03). In 72 h, a greater distribution of the cytoskeleton filaments and the more evident endoplasmatic reticulum was verified, indicating an increase in the protein synthesis when compared with the control group. In the expression of the VEGF gene, a significant increase of 1.98 times (p < 0.05) in the number of transcripts was observed; whereas for the IL6 gene, a decrease of the transcripts was 4.05 times (p < 0.05), both occurring within 72 h after irradiation at 5 J/cm2. The LLLT (660 nm) at the dose of 5 J/cm2 should modulate cellular viability, upregulated VEGF, and downregulated IL6 expression of messenger RNA in culture of L929 fibroblast cells.

Thymus vulgaris L. extract has antimicrobial and anti-inflammatory effects in the absence of cytotoxicity and genotoxicity.

OBJECTIVES: This study evaluated the biological effects of the T. vulgaris L. extract., such as antimicrobial activity on planktonic cultures and mono- and polymicrobial biofilms, cytotoxicity, anti-inflammatory activity and genotoxicity. METHODS: Monomicrobial biofilms of Candida albicans, Staphylococcus aureus, Enterococcus faecalis, Streptococcus mutans and Pseudomonas aeruginosa and polymicrobial biofilms composed by C. albicans with each bacterium were formed for 48h and exposed for 5min to the plant extract. Murine macrophages (RAW 264.7), human gingival fibroblasts (FMM-1), human breast carcinoma cells (MCF-7) and cervical carcinoma cells (HeLa) were also exposed to the plant extract for 5min and the cell viability were analyzed by MTT, neutral red (NR) and crystal violet (CV) assays. Interleukin-1 beta (IL-1ß) and tumor necrosis factor alpha (TNF-α) produced by RAW 264.7 was quantified by ELISA, after 24h exposure to the plant extract, both in the absence and presence of lipopolysaccharide (LPS) from Escherichia coli. Genotoxicity of the plant extract was evaluated by micronucleus formation (MN) in 1000 cells. The results were analyzed by T-Test or ANOVA and Tukey's Test (P≤0.05). RESULTS: All biofilms showed significant reductions in CFU/mL (colony-forming units per milliliter). Cell viability was above 50% for all cell lines. Anti-inflammatory effect on the synthesis of IL-1ß and TNF-α was observed. The MN was similar or lower than the control group in all cells. CONCLUSIONS: T. vulgaris L. extract was effective against all biofilms, promoted high cell viability, anti-inflammatory effect and presented no genotoxicity.

Plant extracts: initial screening, identification of bioactive compounds and effect against Candida albicans biofilms.

AIM: This study screened plants from Brazilian Pantanal for Candida albicans antibiofilm activity. MATERIAL & METHODS: Sixty extracts were obtained from ten plants using different extraction methods. Antifungal activity was assessed. Effects on biofilm inhibition and disruption and cytotoxicity were also evaluated. The most active extract was chemically characterized. RESULTS: Buchenavia tomentosa ethanolic extract showed noticeable antifungal activity and was selected for biofilm experiments. Subinhibitory concentration of extract inhibited fungal adhesion. Maximum killing reached 90% of C. albicans cells in suspension and 65% of cells in biofilms. The active extract was noncytotoxic. Chemical characterization showed the presence of phenols. Ellagic and gallic acids showed activity on C. albicans. CONCLUSION: B. tomentosa extract and its isolated compound, ellagic acid, presented antibiofilm activity and low toxicity.

Comparative analysis of different doses of coherent light (laser) and non-coherent light (light-emitting diode) on cellular necrosis and apoptosis: a study in vitro

Introduction Threshold doses of electromagnetic radiation can initiate necrosis and apoptosis in cells. The purpose of this study was to evaluate cellular apoptosis and necrosis immediately (t0) and 24 hours (t24) after irradiation with different doses of coherent light (laser) or non-coherent light (LED). Methods CHO-K1 lineage cells were irradiated with laser (810nm) or LED (945±20nm), with 24mW, contact area of 1cm2 and doses of 10, 20, 30, 40 and 50J/cm2 for 300, 660, 960, 1230 and 1620s, respectively, at both wavelengths. Cells were evaluated by fluorescence microscopy, differentiating viable, apoptotic and necrotic cells immediately and 24 hours after irradiation. Results The number of necrotic cells at t0 was higher in the LED 40 and 50J/cm2 groups (86±14 and 84±16% respectively, p <0.05), than in the 10 and 20J/cm2 laser (5±2 and 5±3%, p<0.05) and LED (5±3 and 4±1%, p<0.05) conditions. At t24, the LED 40J/cm2 (80±20%, p<0.05) group also showed more necrosis than the control and lower dose groups (laser 10, 20, and 30J/cm2 percentage of 6±4, 10±3 and 7±3%, p<0.05; LED 10 and 20J/cm2 percentage of 3±1 and 17±10%, p<0.05). A decrease in apoptotic cells was observed in the laser group with doses of 10, 40, and 50J/cm2 (6±4, 3±1 and 1±1% respectively, not significant), as well as in the LED 40J/cm2 (2±2%, not significant) group versus control. The cells had a higher percentage of apoptosis cells in the control group and with laser doses of 10 and 30J/cm2 (percentage of 20±1 and 20±4%, not significant), while only the LED 40J/cm2 (10±10%, not significant) had a lower percentage compared the control group. Conclusion Laser or LED stimulation promoted an increase in cell necrosis in a high energy density condition as characterized in a dose-dependent inhibition therapy. Laser or LED infrared irradiation in low doses (up to 20J/cm2) reduced the percentage of apoptosis in CHO-K1 cells, while high doses (30J/cm2) elevated apoptosis.

Efeito da terapia ultrassônica de baixa intensidade em cultura celular de fibroblastos / Efecto del ultrasonido de baja intensidad en cultivo celular de fibroblastos / Effect of low-intensity pulsed ultrasound therapy on a fibroblasts cell culture

Dentro da prática fisioterápica verifica-se a ampla utilização do ultrassom terapêutico para tratamento das diversas afecções musculoesqueléticas. O objetivo deste estudo foi avaliar o efeito da irradiação Ultrassônica de Baixa Intensidade, com diferentes regimes de pulsos e intensidade, em cultura celular de fibroblastos L929 (ATCC CCL-1 NCTC), de modo a verificar a viabilidade celular e definir parâmetros de dosimetria. Para isso, utilizou-se a aplicação de ultrassom pulsado, com frequência de 1Mhz, em cultura de células fibroblásticas, divididas em cinco grupos (controle e com intensidade instantâneas de 0,3W/cm2-10%; 0,3W/cm2 -20 %; 0,5W/cm2 -10% e US 0,5W/cm2 -20 % - 100Hz). A irradiação ocorreu com intervalos de 24, 48 e 72 horas, por dois minutos, e após 24 horas de cada irradiação foi realizado teste de MTT Brometo de [3-(4,5-dimetiltiazol)-2,5-difeniltetrazólio]. Os resultados revelaram que ao compararem-se os valores de células viáveis pelo método MTT nos cinco grupos, não foi possível encontrar diferença estatisticamente significativa em nenhum deles, nos três momentos avaliados (24, 48 e 72 horas); enquanto que, ao se realizar a análise de medida repetida nos diferentes grupos, encontrou-se diferença estatisticamente significativa apenas no grupo irradiado com ultrassom a 0,5W/ cm2com regime de pulso de 10% (p=0,003). Com base nesses resultados, conclui-se que a irradiação Ultrassônica de Baixa Intensidade em cultura celular de fibroblastos L929, somente no grupo com intensidade de 0,5W/cm2-10% obteve o crescimento numérico, com significância estatística em todos os períodos de avaliação.

En la práctica fisioterápica se utiliza bastante el ultrasonido como terapia para el tratamiento de diversos trastornos muscoloesqueléticos. Este artículo tiene por objetivo evaluar el efecto de la irradiación ultrasónica de baja intensidad, con diferentes regímenes de pulsos y de intensidades, en cultivo celular de fibroblastos L929 (ATCC CCL-1 NCTC), para verificar la viabilidad celular y establecer los parámetros de dosimetría. Se utilizó el ultrasonido pulsado, con frecuencia de 1Mhz, en un cultivo de células fibroblásticas, divididas en cinco grupos (con control y con la intensidad instantánea del 0,3W/cm2-10%; 0,3W/cm2 -20%; 0,5W/cm2 -10% y US 0,5W/cm2-20 % - 100Hz). La irradiación se llevó a cabo en intervalos de 24, 48 y 72 horas, durante dos minutos y después de las 24 horas de cada irradiación se realizó la prueba de MTT {Bromuro de [3-(4,5-dimetiltiazol)-2,5-difeniltetrazólio]}. Los resultados mostraron que en la comparación entre los valores de células viables por el método MTT en los cinco grupos evaluados no ha sido posible encontrar ninguna diferencia estadísticamente significativa en los tres momentos evaluados (24, 48 y 72 horas). En cambio, al llevar a cabo el análisis de medida repetida en los diferentes grupos, se obtuvo una diferencia estadísticamente significativa solamente en el grupo irradiado con ultrasonido a 0,5W/cm2 con el régimen de pulso del 10% (p=0,003). Basándose en estos resultados se concluyó que la irradiación ultrasónica de baja intensidad en cultivo celular de fibroblastos L929 obtuvo el aumento sólo en el grupo con intensidad de 0,5W/cm2-10%, con significancia en todos los periodos de evaluación.

Within the physiotherapy practice there is a wide use of therapeutic ultrasound for the treatment of various musculoskeletal disorders. The aim of this study was to evaluate the effect of Low Intensity Ultrasonic irradiation with differents forms of pulse and intensity in cell culture of L929 fibroblasts (ATCC CCL-1 NCTC), in order to check cell viability and define parameters of dosimetry. For this purpose, it was used the application of pulsed ultrasound with a frequency of 1MHz in cultured fibroblast cells divided into five groups (control and instantaneous intensity of 0.3W/cm2-10%, 0.3W/cm2 -20%, 0.5W/cm2-10% and US 0.5W/cm2-20% - 100Hz). Irradiation occurred at intervals of 24, 48 and 72 hours for two minutes and 24 hours after each irradiation test MTT [3-(4,5-dimethylthiazol)-2,5-diphenyltetrazolium bromide] was performed. Results showed that when comparing the values of viable cells by MTT method in the five groups, we could not find statistically significant difference in any of them, in these three conditions (24, 48 and 72 hours); while. Whereas, when performing the analysis of repeated measures in the different groups, it was found a statistically significant difference only in the group irradiated with ultrasound at 0.5W/cm2 with pulse regime of 10% (p=0.003). Based on these results, it is concluded that the Low Intensity Ultrasonic irradiation in L929 fibroblast cell culture, only in the group with an intensity of 0.5W/cm2 -10% obtained numerical growth, with statistical significance in all periods evaluation.

Wound healing properties and mucilage content of Pereskia aculeata from different substrates

Physiologic growth parameters Wound healing Pereskia aculeata Mill., Cactaceae, is a cactus with high mucilage production, well-known for its nutritional properties. Folk use consists on skin injuries, and mucilage is probably involved in the wound healing activity. This work studied some aspects of its cultivation, specifically regarding soil (substrate), to correlate the effects of nutritional content to mucilage production and to the wound-healing property. Plants were grown under five different soil treatment (sand, crude soil, sand and soil, sand and cattle manure, soil and cattle manure), and after eight months extracts were prepared by turbo-extraction to obtain a crude hydroethanolic extract. We evaluated the effects of these extracts on swelling index, cytotoxicity, and in vitro wound healing property. The results show that the substrate used in cultivation may interfere with mucilage production, but not with cytotoxicity and wound healing, this shows the safety of its use, despite the soil treatment received along the various biomes where P. aculeata is cultivated. Furthermore, morphological studies demonstrated the beneficial effect of the mucilage-containing extract on the fibroblast cell culture, corroborating its folk use for wound healing.

Effect of photodynamic therapy supplemented with quercetin in HEp-2 cells.

Photodynamic therapy (PDT) is a technique that can be used as a complementary therapy in cancer treatment combined with other therapeutic modalities. Quercetin (QCT) is known to be effective in the treatment of cancer, by reducing the cell viability of different cancer cell lines. This study aimed to evaluate the influence of different concentrations of QCT in PDT on the viability, mitochondrial membrane potential and induction of apoptosis/necrosis in the human larynx carcinoma cells (HEp-2). The HEp-2 cells were treated with aluminum phthalocyanine tetrasulfonate (AlPcS4) and QCT and subsequently irradiated with a diode laser light (685 nm, 35 mW, 4.5 J/cm(2)). The results demonstrated that treatment of HEp-2 cells with high concentrations of QCT (at least 50 µM) reduced cell viability. This response was enhanced in cells subjected to PDT supplemented with high concentrations of QCT. In addition, was observed decrease in the mitochondrial membrane potential and characteristics of late apoptosis and/or initial necrosis process. QCT at concentrations from 50 µM improves PDT-induced cytotoxicity by significantly reducing cell viability by apoptosis and/or necrosis, and mitochondrial membrane potential of Hep-2 cells.

Avaliação da atividade mitocondrial no processo de morte celular em células tumorais de mama após tratamento com Ciclosporina A e Photosan3® / Evaluation of mitochondrial activity in the cell death process in breast tumor cells after treatment with Cyclosporine A and Photosan3®

INTRODUÇÃO: A terapia fotodinâmica (PDT) é uma modalidade terapêutica para o tratamento de doenças neoplásicas e não neoplásicas tendo como alvo a mitocôndria, organela que tem atraído maior atenção devido ao seu envolvimento direto no processo de morte celular. Inibindo a atividade mitocondrial é possível estudar outras organelas envolvidas no processo de morte celular. O objetivo deste estudo foi avaliar a importância da mitocôndria no processo de morte celular na linhagem celular M3, induzida após a fotossensibilização com Photosan3®. MÉTODOS: Para os experimentos foram utilizados os seguintes grupos: grupo controle I, células sem nenhum tratamento; grupo II PDT, células incubadas com Photosan3®; grupo III PDT, células incubadas com CsA e Photosan3®; grupo IV células tratadas somente com Estaurosporina (STS). Após a incubação com o fotossensibilizador os grupos II e III foram irradiados com diodo laser semicondutor (λ 670 nm). Todos os grupos foram incubados a 37 ºC em estufa com atmosfera de 5% de CO2, por 24 h e 48 h. No final destes períodos todos os grupos foram submetidos ao ensaio de citotoxicidade, pelo teste de MTT (brometo de 3-[4,5-dimetil-tiazol-2-il]-2,5-difeniltetrazólio) e corados com anexina V e iodeto de propídio para determinar a proporção de morte celular, sendo as análises realizadas por microscopia de fluorescência. RESULTADOS: Os resultados mostraram que as células realizaram apoptose por via independente de mitocôndria. A CsA apresentou-se eficiente na inativação da mitocôndria no processo apoptótico durante a fotossensibilização com Photosan3®. CONCLUSÃO: A associação de CsA e Photosan3® na terapia fotodinâmica demonstrou a presença de morte celular por apoptose independente da participação mitocondrial.

INTRODUCTION: Photodynamic therapy (PDT) is a therapeutic modality for the treatment of neoplastic and non-neoplastic diseases. Mitochondria have attracted great attention due to their direct involvement in the cell death process. By inhibiting the mitochondrial activity, it is possible to study other organelles involved in the cell death process. The objective of this study was to evaluate the involvement of mitochondria in induced cell death process in M3 cell line after photosensitization with Photosan3®. METHODS: The experiments involved the following groups: control group I, cells with no treatment; group II PDT, cells incubated with Photosan3®; group III PDT, cells incubated with CsA and Photosan3®; group IV, cells treated only with treated only Staurosporine (STS). After incubation with the photosensitizer, the groups II and III were irradiated using a semiconductor laser diode (λ 670 nm). All groups were incubated at 37 ºC in an atmosphere of 5% CO2 for 24 and 48 h. After this period, all groups were subjected to the MTT (3 - [4,5-dimethyl-thiazol-2-yl] -2,5-diphenyltetrazolium bromide) cytotoxicity assay and labeled with Annexin V and Iodide Propidium to determine the rate of cell death. The analyses were performed by fluorescence microscopy. RESULTS: The results show that PDT Photosan leads to apoptosis of breast cell line M3 by a route independent of the mitochondria. CONCLUSION: The association of CsA and Photosan3® in photodynamic therapy showed the occurrence of cell death (apoptosis) independent of mitochondrial participation.

Effect of low-intensity pulsed ultrasound on l929 fibroblasts.

INTRODUCTION: Ultrasound has proven to be an important therapeutic resource regarding musculoskeletal disease and is routinely used in physical therapy and medicine both therapeutically and diagnostically. The aim of the present study was to analyse the effects with different ultrasound intensities in order to establish the ideal radiation level in cell cultures. MATERIAL AND METHODS: FIBROBLAST CELL CULTURES WERE DIVIDED INTO FIVE GROUPS: group I - control (did not receive irradiation); group II - 0.2 W/cm(2) in pulsed mode at 10% (1 : 9 duty cycle); group III - 0.6 W/cm(2) in pulsed mode at 10% (1 : 9 duty cycle); group IV - 0.2 W/cm(2) in pulsed mode at 20% (2 : 8 duty cycle); and group V - 0.6 W/cm(2) in pulsed mode at 20% (2 : 8 duty cycle). Each group was irradiated with 24-h intervals, observing the following post-irradiation incubation times: 24, 48, 72 and 96 h; after 24 h of each irradiation, cultures were analysed using the MTT method. RESULTS: Analysis of the results following ultrasound irradiation demonstrated that the effect of ultrasound with 0.6 W/cm(2) in pulsed mode at 10% (1 : 9 duty cycle) was statistically significant in relation to ultrasonic irradiation in pulsed mode at 20% (2 : 8 duty cycle) (p < 0.05). CONCLUSIONS: According to parameters used in the irradiation of cultivated fibroblasts, the pulse mode regime and the control of intensity are of fundamental importance for the optimal use of therapeutic ultrasound. Furthermore, low and medium intensities decreased cell damage, which establishes that acoustic pulsed energy induces the proliferation of fibroblast cells.

Effects of a polarized light source (400-2000 nm) on Hep.2 and L929 cell lines: a spectroscopic in vitro study.

OBJECTIVE: The aim of this study was to evaluate the effects on Hep.2 cells originating from laryngeal carcinomas, and L929 cells originating from a fibroblast line, subjected to polarized light at a wavelength of 400-2000 nm. BACKGROUND DATA: Recently there has been increased interest in the propagation of polarized light in randomly scattering media such as biological tissues, because of its potential applications in medicine. MATERIALS AND METHODS: Irradiation was performed at two time points: T0 (24 h after cell culture) and T48 (48 h after the first irradiation). Cellular viability was assessed using an MTT assay at the following times: T0 (first irradiation), T6 (6 h after the first irradiation), T12 (12 h after the first irradiation), T24 (24 h after the first irradiation), T48 (48 h after the first irradiation), and T72 (72 h after the first irradiation). The results were analyzed using Graphpad Prism software. RESULTS: The results showed that time influenced the cellular viability of L929 cells of both control (p = 0.0014) and illuminated cultures (p = 0.0035). Significant differences between control cells (p = 0.0001) and illuminated Hep.2 cells (p = 0.0001) were observed. There was a significant difference between the proliferation of the two types of cells illuminated compared to their controls: Hep.2 (p = 0.0001) and L929 (p = 0.0002). CONCLUSION: The use of polarized light on Hep.2 and L929 cells resulted in photobiological effects that need further investigation, as this is the first study using this methodology.

Assessment of cytoskeleton and endoplasmic reticulum of fibroblast cells subjected to low-level laser therapy and low-intensity pulsed ultrasound.

OBJECTIVE: The aim of the present study was to compare the effect of low-level laser therapy (LLLT) and low-intensity pulsed ultrasound (LIPUS) on the cytoskeleton and endoplasmic reticulum of L929 cells. Thermal and non-thermal physical mechanisms such as LLLT and LIPUS induce clinically significant responses in cells, tissues, and organs. MATERIALS AND METHODS: L929 fibroblast cell cultures were irradiated with LLLT and subjected to LIPUS. Cultures irradiated with the laser (904 nm) were divided into three groups: group I, control (no irradiation); group II, irradiated at 6 J/cm(2); and group III, irradiated at 50 mJ/cm(2). Cultures subjected to ultrasound were divided into five groups: group I, control (no LIPUS); group II, LIPUS at 0.2 W/cm(2) in pulsed mode at 10% (1:9 duty cycle); group III, LIPUS at 0.6 W/cm(2) in pulsed mode at 10% (1:9 duty cycle); group IV, LIPUS at 0.2 W/cm(2) in pulsed mode at 20% (2:8 duty cycle); and group V, LIPUS at 0.6 W/cm(2) in pulsed mode at 20% (2:8 duty cycle). Each group was irradiated at 24-h intervals, with the following post-treatment incubation times: 24, 48, and 72 h. The effects of LLLT and LIPUS on the cytoskeleton and endoplasmic reticulum was evaluated by the use of fluorescent probes and with fluorescence microscopy analysis. RESULTS: The results following LLLT and LIPUS demonstrate that ultrasound was more effective than laser on fibroblast cell cultures when the endoplasmic reticulum was assessed, whereas there was a better distribution of the filaments of the cytoskeleton in the cells subjected to laser irradiation. CONCLUSION: The study demonstrated that both LLLT and LIPUS promote changes on the cellular level. However, LIPUS was more effective than LLLT at the doses used here, as assessed by fluorescence microscopy, which revealed increased reticulum activity and increased protein synthesis. However, when the organization of actin filaments was assessed, LLLT achieved a better result.

Cytotoxicity of octal-bromide zinc phthalocyanine after photodynamic therapy with different light sources.

OBJECTIVE: The objective of this study was to investigate the cytotoxicity of octal-bromide zinc phthalocyanine (ZnPcBr8) at different concentrations (0.25, 0.5, and 1 microM) after irradiating HEp-2 cell cultures with two different light sources: a diode semiconductor laser (660 nm, 30 mW) or an LED (640 nm, 70 mW). In order to obtain comparative results, the irradiation parameters of both light sources were adjusted so that the amount of energy density delivered would be the same (4.5 J/cm2). BACKGROUND DATA: Numerous photosensitizers and light sources used in the treatment of human disease have been studied. Based on these studies, a comparative evaluation of two light sources used in photodynamic therapy (PDT) with ZnPcBr8 was proposed. MATERIALS AND METHODS: HEp-2 cells were incubated with ZnPcBr8 at different concentrations (0.25, 0.5, or 1 microM) for 1 h, irradiated with the diode semiconductor laser (660 nm at 30 mW for 300 sec; 4.5 J/cm2) or the LED laser (640 nm at 70 mW for 128 sec; 4.5 J/cm2), and then incubated in MEM medium for 1 or 24 h. The cells were analyzed using the MTT and trypan blue dye exclusion tests. RESULTS: The results demonstrated that the concentration of 1 microM of ZnPcBr8 was the most effective after PDT administered by both light sources. According to the MTT results, HEp-2-cell viability decreased by 97.96% 1 h after, and by 99.87% 24 h after irradiation with the diode semiconductor laser, and decreased by 94.03% 1 h after, and by 99.21% 24 h after irradiation with the LED. The results obtained using the trypan blue dye exclusion test confirmed the photodynamic efficacy of ZnPcBr8 employed with both light sources. With regard to HEp-2-cell viability, the following results were observed: a decrease of 98.73% 1 h after, and of 99.49% 24 h after irradiation with the diode semiconductor laser; and a decrease of 98.76% 1 h after, and of 99.23% 24 h after irradiation with the LED. CONCLUSIONS: According to our results with the irradiation parameters studied here, both the LED and diode semiconductor laser can be used for PDT in vitro, since both light sources had excellent photodynamic efficacy.

Evaluation of low-level laser therapy of osteoblastic cells.

OBJECTIVE: The purpose of the present study was to evaluate the effect of biomodulation on osteoblastic cells using a gallium-aluminium-arsenide diode laser. BACKGROUND DATA: Low-level laser therapy (LLLT) is a non-pharmacological therapeutic resource to which biological tissues respond well, producing such effects as the acceleration of bone formation and bone repair. MATERIALS AND METHODS: Osteoblastic cell cultures (OFCOL II) were irradiated with a gallium-aluminium-arsenide diode laser (GaAlAs lambda = 830 nm; 50 mW; 3 J/cm(2); 600-microm-diameter optical fiber) and divided into two groups: group 1--irradiated cells, and group 2--non-irradiated cells. Irradiation occurred at 24-h intervals for a total of 3 d. After each interval, the cells were marked with Mito Tracker Orange dye to assess the biostimulatory effect on mitochondrial activity and cell proliferation using an MTT assay. RESULTS: Intense grouping of mitochondria in the perinuclear region was observed at 24 h and 48 h following irradiation. Changes from a filamentous to a granular appearance in mitochondrial morphology and mitochondria distributed throughout the cytoplasm were observed 72 h following proliferation. Such changes led to an in vitro proliferation process, as confirmed by the MTT assay. CONCLUSION: LLLT has shown itself capable of altering mitochondrial activity and the population of OFCOL II cells.

Comparison between the effect of low-level laser therapy and low-intensity pulsed ultrasonic irradiation in vitro.

OBJECTIVE: The objective of this study was to compare the effect of low-level laser therapy (LLLT) and low-intensity pulsed ultrasound (LIPUS) on fibroblast cell culture. Several methods, including ultrasound treatment and LLLT, are being used to facilitate tissue repair and healing processes. MATERIALS AND METHODS: L929 fibroblast cell cultures were irradiated with low-level laser energy and LIPUS. Cultures irradiated with ultrasound were divided into five groups: group 1: control (did not receive irradiation); group 2: 0.2 W/cm(2) in pulsed mode at 10% (1:9 duty cycle); group 3: 0.6 W/cm(2) in pulsed mode at 10% (1:9 duty cycle); group 4: 0.2 W/cm(2) in pulsed mode at 20% (2:8 duty cycle); and group 5: 0.6 W/cm(2) in pulsed mode at 20% (2:8 duty cycle). Cultures irradiated with laser energy were divided into three groups: group 1: control (did not receive irradiation); group 2: 6 J/cm(2); and group 3: 50 mJ/cm(2). Each group was irradiated at 24-h intervals, with the following incubation periods post-irradiation: 24, 48, and 72 h; after each irradiation cycle the cultures were analyzed using MTT [3-(4.5-dimethylthiazol-2-yl)-2.5 diphenyltetrazolium bromide]. RESULTS: Analysis of results after LLLT and LIPUS demonstrated that the effect of laser therapy on fibroblast cell culture was greater than that of LIPUS (p < 0.05). CONCLUSION: Results demonstrated that LLLT significantly increased fibroblastic activity more than LIPUS. Therefore, in the first and second phases of tissue repair, laser treatment may be more effective than ultrasound treatment.

Surface morphology of sound deciduous tooth enamel after application of a photo-absorbing cream and infrared low-level laser irradiation: an in vitro scanning electron microscopy study.

OBJECTIVES: The purpose of this descriptive scanning electron microscopic study was to characterize surface alterations in deciduous tooth enamel after in vitro infrared diode laser irradiation, using a photo-absorbing agent alone and also combined with fluoride, before and after laser irradiation. BACKGROUND DATA: Previous investigations have demonstrated increased enamel caries resistance after laser irradiation. METHODS: Seven extracted or exfoliated primary molar teeth underwent soft tissue débridement and fluoride-free prophylaxis. Buccal surfaces were determined to be caries free by macroscopic examination. Sample groups were divided into: (1) control (no treatment); (2) infrared diode laser irradiation (lambda = 810 nm, 68 nm, 60 mW/mm(2), 30 W) using the photo-absorbing agent alone (IRDL + PA; 500 J/cm(2)); and (3) infrared diode laser irradiation using a photo-absorbing agent combined with 2% fluoride (IRDL + PFA; 500 J/cm(2)). Buccal surfaces were evaluated following standard scanning electron microscopy preparation techniques. Control samples of enamel surfaces were relatively smooth but presented occasional enamel prism ends. There were no areas with cavitations or surface defects. RESULTS: After the IRDL + PA treatment, irradiated surfaces became rough and mildly to moderately irregular with scarce enamel cavitations and without exposure of enamel prism ends. The surfaces had adherent granules and only occasional fine cracks and porosities in surface coatings were noted. After the IRDL + PFA treatment, there was a homogenous confluent surface that masked typical enamel surface markings. The surfaces had well-defined globules resulting from the IRDL + PFA treatment, that were not seen after IRDL + PA treatment. CONCLUSIONS: Treatment of deciduous tooth enamel with infrared diode laser irradiation using a photo-absorbing agent and a photo-absorbing agent combined with 2% fluoride created surface coatings that may act as reservoirs for mineral phases during cariogenic activity on enamel, and also provide a certain degree of protection against cariogenic challenge.

Photodynamic therapy with a new photosensitizing agent.

OBJECTIVE: The aim of this study was to investigate the cytotoxicity of octal-bromide zinc phthalocyanine (ZnPcBr(8)) before and after irradiation with a low-power laser (AsGaAl) and analyze the effects of photodynamic therapy (PDT) on the nucleus of L929 cells. BACKGROUND DATA: One of the most recent and promising applications of phthalocyanine in medicine is in the detection and cure of tumors. We studied the ZnPcBr(8) in agreement with the development of new photosensitizing agents for curing tumors. METHODS: L929 cells were cultivated at standard conditions, incubated with ZnPcBr(8) for 1 h at different concentrations, irradiated with a semiconductor laser, and incubated in MEM medium for 1, 12, or 24 h. Cells were analyzed using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide (MTT) technique and fluorescence microscopy. RESULTS: The results demonstrated that ZnPcBr(8) at 1 microM was the most effective concentration for PDT, with a decrease of 63% after 1 h, 99% after 12 h, and 100% after 24 h in relation to the control group. The fluorescence microscopy results showed that ZnPcBr(8) was localized in the perinuclear region when analyzed 1 h after incubation. Nucleus staining with DAPI made it possible to observe that nuclear fragmentation occurred 24 h after PDT, cytoplasm retraction at 1, 12, and 24 h after PDT, and vacuoles along the cytoplasm at 12 and 24 h after PDT. CONCLUSION: According to the results obtained in this study, L929 cell death caused by PDT with ZnPcBr(8) possesses characteristics of apoptosis mediated by the mitochondria, due to the decrease in cells viability, the subcellular localization, and the photodamage found.

The effect of laser therapy on the proliferation of oral KB carcinoma cells: an in vitro study.

OBJECTIVE: The aim of this study was to assess the proliferative effect of carcinoma cells, strain KB, submitted to laser therapy with wavelengths of lambda685nm (31 mW; Ø; 0.38 cm(2), 4 J/cm(2)) or lambda830nm (34.5 mW; Ø; 0.38 cm(2), 4 J/cm(2)). BACKGROUND DATA: It is known that the interaction of laser light with living tissues may lead to different results depending upon several factors such as wavelength, dose, potency, and optical properties of the tissue as well as on the condition being treated. The response to the use of laser light may be of stimulation or inhibition. One successful model used to study the effects of laser light on living tissues is the in vitro use of different lineages of cells in culture. METHODS: Cellular viability was assessed using MTT spectroscopy immediately, and 6, 12, 24, and 72 h after treatment. The irradiations were carried out twice, at 24 h after cell seeding and at 48 h after the first irradiation. The dose of 4 J/cm(2) was given by a lambda685 nm (31 mW, Phi 0.8 cm(2)) or lambda830 nm (34.5 mW, Phi 0.8 cm(2)) diode lasers. RESULTS: The results demonstrated that the time influenced significantly both control (p = 0.01) and both cultures irradiated with lambda685-nm laser (p = 0.01) or lambda830-nm laser (p = 0.09). The influence of the treatment (laser therapy) was also significant when comparing the results observed in irradiated groups and the control (p = 0.01). The influence of the wavelength in the final result, in other words, in the cellular viability of cultures irradiated with the two wavelengths was also significant (p = 0.01). CONCLUSIONS: It is concluded that laser therapy had a positive biomodulatory effect on the proliferation of KB cells and that this was influenced by the wavelength.

Laser light is capable of inducing proliferation of carcinoma cells in culture: a spectroscopic in vitro study.

OBJECTIVE: The objective of this work was to evaluate the influence of time, treatment, and wavelength, through the assessment of the cellular viability with MTT, on the proliferation of H.Ep.2 cells subjected to laser irradiation or not (lambda685 and lambda830 nm) with the same energy density (4 J/cm2). BACKGROUND DATA: Although malignant lesions have been studied for some time, there is not yet a definitive cure. Mortality could be reduced if lesions were diagnosed on initial phases of development. MATERIALS AND METHODS: H.Ep.2 cells were cultured in flasks and maintained in DMEN medium (10% FBS, 1% L-glutamine, and 1% antibiotic solution). For irradiation, cells were kept in 24 wells of the 96-well plaques containing DMEM medium (5% FBS, 1% L-glutamine, and 1% antibiotic solution), irradiated with lasers at lambda685- and lambda830-nm wavelength, and stained at 0, 6, 12, 24, and 48 h after irradiation. RESULTS: There was significant differences when the three groups were compared (p = 0.0087). There was significant difference for both irradiated groups, lambda685 nm (p = 0.0202) and lambda830 nm (p = 0.0324). Time of irradiation significantly influenced only the lambda685-nm group (p = 0.04). The wavelength had a significant influence (p = 0.013). CONCLUSION: Time, treatment, and wavelength significantly influenced the proliferation process of H.Ep.2 cells.