Potencial alelopático e identificação de compostos secundários em extratos de calopogônio (Calopogonium mucunoides) utilizando eletroforese capilar

In this paper we describe the assessment of the possible allelopathic potential of organic extracts obtained from leaves of Calopogonium mucunoides under laboratory conditions, after that some secondary compounds were identified and quantified using capillary electrophoresis. After the identification and quantification of the compounds, we studied the effects of these compounds on the germination of some common weed, which are actually becoming a real problem in pastures in the state of Pará - Brazil. Calopogônio presented allelopathic potential. The organic crude extracts obtained from solvents with high dielectric constants (dichloromethane and ethyl acetate) were the most efficient in the inhibition of the weed germination. Seeds of Mimosa pudica were more affected by the extracts; this fact reveals the specificity of the organic extracts obtained. Capillary electrophoresis protocols were highly specific, which makes it possible to identify 5 classes of compounds using the same crude extract samples and analyze them fartly (up to 20min). Many of identified compounds show inhibitory effect in the weeds germination, seeds of malicia were the most sensible, the bioassays with the misture of compounds indicated the possibility of synergic effect.Neste trabalho avaliou-se o potencial alelopático de extratos orgânicos obtidos a partir das folhas de Calopogonium mucunoides sobre a germinação de sementes de algumas plantas daninhas comumente encontradas em áreas de pastagens cultivadas da Amazônia brasileira, as quais causam grandes danos à produtividade: Cassia tora (mata-pasto), Mimosa pudica (malícia) e Cassia occidentalis (fedegoso). Compostos secundários foram identificados e quantificados nos extratos brutos utilizando eletroforese capilar. Após identificar e quantificar os compostos presentes nos extratos realizaram-se novos bioensaios com os padrões dos compostos identificados a fim de verificar se os mesmos poderiam atuar como inibidores na germinação das sementes das plantas daninhas em estudo. Calopogonium mucunoides apresentou potencial alelopático o qual variou com a espécie de planta daninha estudada. Os protocolos desenvolvidos utilizando eletroforese capilar se mostraram eficientes e bastante específicos, sendo possível a separação e identificação de 5 classes de compostos nos extratos brutos sem necessidade de clean up ou fracionamento dos mesmos, com análises rápidas (em menos de 20 minutos) e baixas quantidades de solventes utilizadas quando comparadas aos métodos tradicionais de análises. Vários dos compostos identificados apresentaram potencial de inibição de germinação nas sementes estudadas, sendo malícia a mais sensível, os bioensaios também indicaram certo efeito sinérgico ao utilizar a mistura de compostos.Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)Universidade de São Paulo Instituto de Química de São CarlosUniversidade Federal de São Paulo (UNIFESP) Departamento de Ciências Exatas e da TerraEmpresa Brasileira de Pesquisa AgropecuáriaUNIFESP, Depto. de Ciências Exatas e da TerraSciEL

In Vitro Wound Healing Improvement By Low-level Laser Therapy Application In Cultured Gingival Fibroblasts

The aim of this study was to determine adequate energy doses using specific parameters of LLLT to produce biostimulatory effects on human gingival fibroblast culture. Cells (3 10 4 cells/cm 2) were seeded on 24-well acrylic plates using plain DMEM supplemented with 10 fetal bovine serum. After 48-hour incubation with 5 CO2 at 37C, cells were irradiated with a InGaAsP diode laser prototype (LASERTable; 780 3 nm; 40mW) with energy doses of 0.5, 1.5, 3, 5, and 7J/cm 2. Cells were irradiated every 24h totalizing 3 applications. Twenty-four hours after the last irradiation, cell metabolism was evaluated by the MTT assay and the two most effective doses (0.5 and 3J/cm 2) were selected to evaluate the cell number (trypan blue assay) and the cell migration capacity (wound healing assay; transwell migration assay). Data were analyzed by the Kruskal-Wallis and Mann-Whitney nonparametric tests with statistical significance of 5. Irradiation of the fibroblasts with 0.5 and 3J/cm 2 resulted in significant increase in cell metabolism compared with the nonrradiated group (P 0.05). Both energy doses promoted significant increase in the cell number as well as in cell migration (P 0.05). These results demonstrate that, under the tested conditions, LLLT promoted biostimulation of fibroblasts in vitro. Copyright © 2012 Fernanda G. Basso et al.Hkkinen, L., Uitto, V.J., Larjava, H., Cell biology of gingival wound healing (2000) Periodontology 2000, 24 (1), pp. 127-152Kreisler, M., Christoffers, A.B., Al-Haj, H., Willershausen, B., D'Hoedt, B., Low level 809-nm diode laser-induced in vitro stimulation of the proliferation of human gingival fibroblasts (2002) Lasers in Surgery and Medicine, 30 (5), pp. 365-369. , DOI 10.1002/lsm.10060Posten, W., Wrone, D.A., Dover, J.S., Arndt, K.A., Silapunt, S., Alam, M., Low-level laser therapy for wound healing: Mechanism and efficacy (2005) Dermatologic Surgery, 31 (3), pp. 334-340Saygun, I., Karacay, S., Serdar, M., Ural, A.U., Sencimen, M., Kurtis, B., Effects of laser irradiation on the release of basic fibroblast growth factor (bFGF), insulin like growth factor-1 (IGF-1), and receptor of IGF-1 (IGFBP3) from gingival fibroblasts (2008) Lasers in Medical Science, 23 (2), pp. 211-215. , DOI 10.1007/s10103-007-0477-3Skopin, M.D., Molitor, S.C., Effects of near-infrared laser exposure in a cellular model of wound healing (2009) Photodermatology Photoimmunology and Photomedicine, 25 (2), pp. 75-80Hakki, S.S., Bozkurt, S.B., Effects of different setting of diode laser on the mRNA expression of growth factors and type i collagen of human gingival fibroblasts (2012) Lasers in Medical Science, 27 (2), pp. 325-331Peplow, P.V., Chung, T.Y., Baxter, G.D., Laser photobiomodulation of proliferation of cells in culture: A review of human and animal studies (2010) Photomedicine and Laser Surgery, 28, pp. 3-S40. , supplement 1Basso, F.G., Oliveira, C.F., Kurachi, C., Hebling, J., Costa, C.A., Biostimulatory effect of low-level laser therapy on keratinocytes in vitro Lasers in Medical Science, , In pressOliveira, C.F., Basso, F.G., Lins, E.C., Kurachi, C., Hebling, J., Bagnato, V.S., De Souza Costa, C.A., In vitro effect of low-level laser on odontoblast-like cells (2011) Laser Physics Letters, 8 (2), pp. 155-163Mosmann, T., Rapid colorimetric assay for cellular growth and survival: Application to proliferation and cytotoxicity assays (1983) Journal of Immunological Methods, 65 (1-2), pp. 55-63Wiegand, C., Hipler, U., Methods for the measurement of cell and tissue compatibility including tissue regeneration process (2008) GMS Krankenhaushygiene Interdisziplinr, 3 (1), pp. 1863-5245Hoang, A.M., Oates, T.W., Cochran, D.L., In vitro wound healing responses to enamel matrix derivative (2000) Journal of Periodontology, 71 (8), pp. 1270-1277Liang, C.-C., Park, A.Y., Guan, J.-L., In vitro scratch assay: A convenient and inexpensive method for analysis of cell migration in vitro (2007) Nature Protocols, 2 (2), pp. 329-333. , DOI 10.1038/nprot.2007.30, PII NPROT.2006.30Cceres, M., Romero, A., Copaja, M., Daz-Araya, G., Martnez, J., Smith, P.C., Simvastatin alters fibroblastic cell responses involved in tissue repair (2011) Journal of Periodontal Research, 46 (4), pp. 456-463Chor, A., De Azevedo, A.M., Maiolino, A., Nucci, M., Successful treatment of oral lesions of chronic lichenoid graft-vs.-host disease by the addition of low-level laser therapy to systemic immunosuppression (2004) European Journal of Haematology, 72 (3), pp. 222-224. , DOI 10.1046/j.0902-4441.2003.00202.xAbramoff, M.M.F., Lopes, N.N.F., Lopes, L.A., Dib, L.L., Guilherme, A., Caran, E.M., Barreto, A.D., Petrilli, A.S., Low-level laser therapy in the prevention and treatment of chemotherapy-induced oral mucositis in young patients (2008) Photomedicine and Laser Surgery, 26 (4), pp. 393-400Woodruff, L.D., Bounkeo, J.M., Brannon, W.M., Dawes Jr., K.S., Barham, C.D., Waddell, D.L., Enwemeka, C.S., The efficacy of laser therapy in wound repair: A meta-analysis of the literature (2004) Photomedicine and Laser Surgery, 22 (3), pp. 241-247. , DOI 10.1089/1549541041438623Damante, C.A., De Micheli, G., Miyagi, S.P.H., Feist, I.S., Marques, M.M., Effect of laser phototherapy on the release of fibroblast growth factors by human gingival fibroblasts (2009) Lasers in Medical Science, 24 (6), pp. 885-891Almeida-Lopes, L., Rigau, J., Zangaro, R.A., Guidugli-Neto, J., Jaeger, M.M.M., Comparison of the low level laser therapy effects on cultured human gingival fibroblasts proliferation using different irradiance and same fluence (2001) Lasers in Surgery and Medicine, 29 (2), pp. 179-184. , DOI 10.1002/lsm.1107Alghamdi, K.M., Kumar, A., Moussa, N.A., Low-level laser therapy: A useful technique for enhancing the proliferation of various cultured cells (2011) Lasers in Medical Science, 27 (1), pp. 237-249Gao, X., Xing, D., Molecular mechanisms of cell proliferation induced by low power laser irradiation (2009) Journal of Biomedical Science, 164Karu, T.I., Pyatibrat, L.V., Kolyakov, S.F., Afanasyeva, N.I., Absorption measurements of a cell monolayer relevant to phototherapy: Reduction of cytochrome c oxidase under near IR radiation (2005) Journal of Photochemistry and Photobiology B: Biology, 81 (2), pp. 98-106. , DOI 10.1016/j.jphotobiol.2005.07.002, PII S1011134405001302Eells, J.T., Henry, M.M., Summerfelt, P., Wong-Riley, M.T.T., Buchmann, E.V., Kane, M., Whelan, N.T., Whelan, H.T., Therapeutic photobiomodulation for methanol-induced retinal toxicity (2003) Proceedings of the National Academy of Sciences of the United States of America, 100 (6), pp. 3439-3444. , DOI 10.1073/pnas.0534746100Zhang, L., Xing, D., Gao, X., Wu, S., Low-power laser irradiation promotes cell proliferation by activating PI3K/Akt pathway (2009) Journal of Cellular Physiology, 219 (3), pp. 553-562Azevedo, L.H., De Paula Eduardo, F., Moreira, M.S., De Paula Eduardo, C., Marques, M.M., Influence of different power densities of LILT on cultured human fibroblast growth: A pilot study (2006) Lasers in Medical Science, 21 (2), pp. 86-89. , DOI 10.1007/s10103-006-0379-9Lagan, K.M., Alyson Clements, B., McDonough, S., David Baxter, G., Low intensity laser therapy (830nm) in the management of minor postsurgical wounds: A controlled clinical study (2001) Lasers in Surgery and Medicine, 28 (1), pp. 27-32. , DOI 10.1002/1096-9101(2 001)28:13.0.CO;2-

Associations between movement behaviours and obesity markers among preschoolers compliant and non-compliant with sleep duration:A latent profile analysis

This study identifies physical activity (PA) and sedentary behaviour (SB) clusters in preschoolers compliant (C) or non-compliant (NC) with sleep recommendations; and associates these clusters with obesity markers. PA and SB were objectively assessed (Actigraph WGT3-X) in 272 preschoolers (4.4 ± 0.7 years old). Sleep duration was parent-reported, and preschoolers were classified as C (3–4 years old: 600–780 min/day; 5 years old: 540–660 min/day) or NC with sleep recommendations. Body mass index (BMI) and waist circumference (WC) were assessed according to international protocols. Moderate to vigorous physical activity (MVPA) and light physical activity (LPA) were categorized as low/high (60 min/day or <180 min/180 min/day, respectively). SB was defined according to mean values between clusters. Latent profile analysis was performed. Associations between the observed clusters and obesity markers were determined using linear regression (RStudio; 1.3.1073). Four cluster solutions for C and NC preschoolers were identified. A negative association between C/Low MVPA cluster and BMI, and a positive association between NC/Low MVPA and BMI (β = −0.8, 95%CI = −1.6;−0.1, and β = 0.9, 95%CI = 0.1;1.7, respectively) were observed. No association was seen for SB clusters. Adequate sleep duration may have a protective role for preschoolers’ BMI, even if the children do not comply with MVPA recommendations

Cytotoxic Effects Of Zoledronic Acid On Human Epithelial Cells And Gingival Fibroblasts

Bisphosphonate-induced osteonecrosis has been related to the cytotoxicity of these drugs on oral mucosa cells. A previous study showed that 5 μM of zoledronic acid (ZA), a nitrogen-containing bisphosphonate, is the highest concentration of this drug found in the oral cavity of patients under treatment. Therefore, in order to simulate an osteonecrosis clinical condition, the aim of this study was to evaluate the highest concentration of ZA applied on human epithelial cells (HaCaT) and gingival fibroblasts. For this purpose, cells (3x104 cells/cm2) were seeded in wells for 48 h using complete culture medium (cDMEM). After 48 h incubation, the cDMEM was replaced by fresh serum-free culture medium (DMEM-FBS) in which the cells were maintained for additional 24 h. Then, 5 μM ZA were added to the DMEM-FBS and the cells incubated in contact with the drug for 48 h. After this period, the number of viable cells (trypan blue), cell viability (MTT assay), total protein (TP) production and cell morphology (SEM analysis) were assessed. Data were analyzed statistically by Mann-Whitney, ANOVA and Tukey's test (α=0.05). ZA caused a significant reduction in the number of viable cells and decreased the metabolic activity of both cell lines. However, decrease of TP production occurred only in the epithelial cell cultures. Morphological alterations were observed in both cell types treated with ZA. In conclusion, ZA (5 μM) was cytotoxic to human epithelial cells and gingival fibroblast cultures, which could be associated, clinically, with the development of bisphosphonateinduced osteonecrosis.246551558Civitelli, R., Napoli, N., Armamento-Villareal, R., Use of intravenous bisphosphonates in osteoporosis (2007) Curr Osteoporos Rep, 5, pp. 8-13Cohen, S.B., An update on bisphosphonates (2004) Curr Rheumatol Rep, 6, pp. 59-65Rogers, M.J., Watts, D.J., Russel, R.G., Overview of bisphosphonates (1997) Cancer, 80, pp. 1652-1660Rogers, M.J., Gordon, S., Benford, H.L., Coxon, F.P., Luckman, S.P., Monkkonen, J., Cellular and molecular mechanisms of action of bisphosphonates (2000) Cancer Supl, 88, pp. 2961-2978Lawson, M.A., Xia, Z., Barnett, B.L., Triffitt, J.T., Phipps, R.J., Dunford, J.E., Differences between bisphosphonates in binding affinities for hydroxyapatite (2010) J Biomed Mater Res Part B: Appl Biomater, 92, pp. 149-155Allen, M.R., Burr, D.B., The pathogenesis of bisphosphonate-related osteonecrosis of the jaw: So many hypotheses, so few data (2009) J Oral Maxillofac Surg, 67, pp. 61-70Otto, S., Pautke, C., Opelz, C., Wesphal, I., Drosse, I., Swager, J., Osteonecrosis of the jaw: Effects of bisphosphonate type, local concentration, and acidic milieu on the pathomechanism (2010) J Oral Maxillofac Surg, 68, pp. 2837-2845Reid, I.R., Booland, M.J., Is bisphosphonate-associated osteonecrosis of the jaw caused by soft tissue toxicity? (2007) Bone, 41, pp. 318-320Scheper, M.A., Badros, A., Chausparat, R., Cullen, K.J., Meiller, T.F., Effect of zoledronic acid on oral fibroblasts and epithelial cells: A potential mechanism of bisphosphonate-associated osteonecrosis (2009) Br J Haematol, 144, pp. 667-676Scheper, M.A., Badros, A., Salama, A.R., Wartburton, G., Cullen, K.J., Weikel, D.S., A novel bioassay model to determine clinically significant bisphosphonate levels (2009) Support Care Cancer, 17, pp. 1553-1557Ruggiero, S.L., Mehrotra, B., Rosenberg, T.J., Engroff, S.L., Osteonecrosis of the jaws associated with the use of bisphosphonates: A review of 63 cases (2004) J Oral Maxillofac Surg, 62, pp. 527-534Walter, C., Klein, M.O., Pabst, A., Al-Nawas, B., Duscher, H., Ziebart, T., Influence of bisphosphonates on endothelial cells, fibroblasts, and osteogenic cells (2010) Clin Oral Investig, 14, pp. 35-41Kumar, S.K.S., Gorur, A., Schaauddin, C., Shuler, C.F., Costerton, J.W., Sedghizadeh, P.P., The role of microbial biofilms in osteonecrosis of the jaw associated with bisphosphonate therapy (2010) Curr Osteoporos Rep, 8, pp. 40-48Aas, J.A., Paster, B.J., Stokes, L.N., Olsen, I., Dewhirst, F.E., Defining the normal bacterial flora of the oral cavity (2005) J Clin Microbiol, 43, pp. 5721-5732Basso, F.G., Pansani, T.N., Turrioni, A.P.S., Bagnato, V.S., Hebling, J., de Souza Costa, C.A., In vitro wound healing improvement by low-level laser therapy application in cultured gingival fibroblasts (2012) Int J Dent, , [Epub ahead of print. DOI: 10.1155/2012/719452]Wiegand, C., Hipler, U., Methods for the measurement of cell and tissue compatibility including tissue regeneration process (2008) GMS Krankenhhyg Interdiszip, 3, pp. 1863-5245Basso, F.G., Oliveira, C.F., Kurachi, C., Hebling, J., de Souza Costa, C.A., Biostimulatory effect of low-level laser therapy on keratinocytes in vitro (2013) Lasers Med Sci, 28, pp. 367-374De Souza Costa, C.A., Duarte, P.T., de Souza, P.P., Giro, E.M., Hebling, J., Cytotoxic effects and pulpal response caused by a mineral trioxide aggregate formulation and calcium hydroxide (2008) Am J Dent, 21, pp. 255-261Oliveira, C.F., Basso, F.G., Lins, E.C.C., Kurachi, C., Hebling, J., Bagnato, V.S., Increased viability of odontoblast-like cells subjected to low-level laser irradiation (2010) Laser Phys, 20, pp. 1659-1666Read, S.M., Northcote, D.H., Minimization of variation in the response to different proteins of the Coomassie blue G dye-binding assay for protein (1981) Anal Biochem, 116, pp. 53-64Oliveira, C.F., Basso, F.G., Lins, E.C., Kurachi, C., Hebling, J., Bagnato, V.S., In vitro effect of low-level laser on odontoblast-like cells (2011) Laser Phys Lett, 8, pp. 155-163Simon, M.J.K., Niehoff, P., Kimming, B., Wiltfang, J., Açil, Y., Expression profile and synthesis of different collagen types I, II III and V of human gingival fibroblasts, osteoblasts, ans SaOs-2 cells after bisphosphonate treatment (2010) Clin Oral Investig, 14, pp. 51-58Migliorati, C.A., Siegel, M.A., Elting, L.S., Bisphosphonate-associated osteonecrosis: A long-term complication of bisphosphonate treatment (2006) Lancet Oncol, 7, pp. 508-514Werner, S., Krieg, T., Smola, H., Keratinocyte-fibroblast interactions in wound healing (2007) J Investigative Dermatol, 127, pp. 998-1008Ravosa, M.J., Ning, J., Liu, Y., Stack, M.S., Bisphosphonate effects on the behavior of oral epithelial cells and oral fibroblasts (2011) Arch Oral Biol, 56, pp. 491-49

Adubos verdes e seus efeitos no rendimento da cana-de-açúcar em sistema de plantio direto.

O objetivo deste trabalho foi avaliar adubos verdes e seus efeitos no rendimento da cana-de-açúcar em sistema de plantio direto (SPD). O trabalho foi realizado em Campos dos Goytacazes (RJ), no período de dezembro de 2003 a julho de 2005. O delineamento experimental foi em blocos casualizados, com quatro repetições. Os tratamentos foram: feijão-de-porco (Canavalia ensiformis), mucuna-preta(Mucuna aterrimum), crotalária (Crotalária juncea) em plantio direto e vegetação espontânea em preparo convencional (testemunha). Com crotalária aos 35 dias após emergência (DAE) houve maior taxa de cobertura do solo – 87% – e, aos 92 DAE produziu 17.852 kg ha-1 de matéria seca, respectivamente, 41%, 78% e 407% superior ao feijão-de-porco, mucuna e vegetação espontânea, além de superá-las em acúmulos de K, Mg, S, Zn e Fe. O feijão-de-porco e a mucuna proporcionaram o maior teor de N na parte aérea. Com feijão-de-porco, os teores de P e Ca foram maiores que a crotalária e a mucuna. Com vegetação espontânea, o maior teor de K foi na parte aérea. As leguminosas acumularam maiores quantidades de N e Cu do que a vegetação espontânea. A crotalária e o feijão-de-porco acumularam 66% a mais de P na parte área que a mucuna. O SPD utilizando a adubação verde contribuiu significativamente para a maior produtividade de cana-de-açúcar, 135.863 kg ha-1, sendo 37% superior ao PC com a vegetação espontânea