Marcadores de superfície de Células-tronco: estudo de reação cruzada inter-espécies

Mesenchymal stem cells (MSCs) are adult multipotent cells with fibroblastoid morphology and adherent to plastic. Furthermore, they can be obtained from different sources. Besides bone marrow, these cells are taken from umbilical cord blood, umbilical vein, saphenous vein, peripheral blood, arteries, liver and fetal pancreas, placenta, dental pulp and adipose tissue. MSCs derived from adipose tissue are important because of the abundant number of cells that can be obtained from this tissue, easy access and little discomfort to the patient. This study compared two techniques for obtaining MSCs from adipose tissue: mechanical dissociation (MD) and enzymatic digestion (ED). We also analyzed the inter-species cross-reactions using commercial monoclonal antibodies directed against surface antigens of stem cells from different species: mouse, horse, rabbit, monkey and human. We found that MD technique is favorable in relation to ED within 15 days of culture, and ED is more efficient in the first days of culture. The data also showed that MD causes less damage to cellular DNA. About inter-species cross-reactions, the monoclonal antibody A69 directed against stem cells from rabbits, which can be used in veterinary medicine, particularly in research involving horses

Efeito da opsonização de células leveduriformes de Paracoccidioides brasiliensis com soro humano sobre a atividade fungicida de monócitos

Monócitos e macrófagos são células da imunidade inata que desempenham importante papel na defesa contra infecções fúngicas, através do reconhecimento dos fungos, ativação e desenvolvimento de atividade fungicida. A função das células fagocitárias depende do seu estado de ativação, induzido principalmente pelo ambiente de citocinas, presentes durante a interação com o microrganismo. O presente projeto teve por objetivo avaliar o efeito da opsonização de células leveduriformes de Paracoccidioides brasiliensis com soro humano sobre a produção de peróxido de hidrogênio (H2O2) e atividade fungicida de monócitos humanos, estimulados ou não com Interferon-gama (IFN-γ), contra P. brasiliensis. Monócitos de sangue periférico, obtidos de indivíduos saudáveis, foram cultivados na ausência ou presença de IFN-γ por 24h a 37oC. A seguir, essas células foram desafiadas com a amostra Pb18 de P. brasiliensis por 60 min para determinação da produção de H2O2 e por 4h para a avaliação da atividade fungicida nas proporções fungomonócito de 1:50. A suspensão fúngica foi previamente incubada por 30 min a 37oC na ausência de soro (SS) ou na presença de pool de soro humano normal, inativado (SI) por aquecimento a 56oC ou soro humano fresco (SF) não submetido à inativação do sistema complemento, nas concentrações de 5%, 10%, 20% e 30% em meio de cultura RPMI. A atividade fungicida de monócitos contra a amostra Pb18 foi avaliada por plaqueamento dessas co-culturas em meio de cultivo BHIágar e determinação da recuperação de fungos viáveis após 14 dias de incubação a 36oC. A produção de H2O2 por monócitos foi determinada através da técnica de redução do vermelho de fenol. Os resultados mostraram que a adição de SF ou SI em diferentes concentrações a culturas de P. brasiliensis não interfere com a viabilidade... (Resumo completo, clicar acesso eletrônico abaixo)

Influência da luz LED na cultura celular de Mieloma múltiplo murino–NS1 in vitro

A monoclonal antibody (mAb) is an important tool in medical biotechnology and the production of biopharmaceuticals, especially for disease diagnosis and treatment of infections, because the antibodies have a significant advantage over chemical agents used in conventional therapies . The last thirty years the technology of production of monoclonal antibodies developed mainly the technique of obtaining in vitro, but also of their production is laborious, the cost is high. A major element of the high cost of production is the fact that the long-term culture consumes a large amount of imported inputs with high added value. A major contribution of this work is to promote cell growth more quickly and efficiently. Currently, a great race to discover new technologies and techniques to synthesize new antibodies and significantly increase the production of murine mAbs. New technologies such as laser and LED are innovations and widespread in modern life, so much so that its use has proliferated worldwide, primarily in the medical field. Recent studies show a series of results from the influence of the LED light in biological tissues such as: increasing the rate of cell proliferation, increased production rate of fibroblasts, increasing the rate of synthesis of RNA and DNA synthesis of ATP, etc. To assess the contribution of the LED in the culture of Myeloma NS1murino compared to the standard procedure. - NS1 cells were provided and followed the criteria of culture medium of the Laboratory of Cellular Engineering Center of Botucatu (POPs). The same amount of cells was grown in bottles of 25 cm2 polystyrene Tissue Culture Treated, specifically marked and kept in special medium RPMI 1640 Gibco BRL  supplemented with fetal bovine serum 10%, essential amino acids and non-essential, glucose, insulin and antibiotics. It was used in LEDs Cromatek wavelength of 630nm, 475nm and 530nm. The groups were... (Complete abstract click electronic access below)

Produção de PGE2 por células dendríticas mediante fagocitose de células apoptóticas infectadas

A fagocitose de células apoptóticas é um processo dinâmico e de fundamental importância para homeostase dos tecidos após uma injúria. A fagocitose de células apoptóticas promove a síntese de mediadores anti-inflamatórios como PGE2, TGF-β e IL-10, podendo resultar na supressão da resposta imune do hospedeiro contra agentes infecciosos. Entretanto, um elegante estudo utilizando células apoptóticas infectadas demonstrou que a fagocitose destas células promove a geração não apenas de citocinas anti-inflamatórias como TGF-β mas também de IL-6 e IL-23, promovendo um efeito imunoestimulador, a diferenciação de células Th17. A atuação da PGE2 na imunidade adaptativa vem sendo investigada quanto à diferenciação e ativação de linfócitos Th1, Treg e Th17. Os resultados aqui apresentados demonstram que o protocolo de diferenciação de células dendríticas utilizado foi capaz de gerar em torno de 85% de CD imaturas evidenciado pela expressão de um perfil fenotípico CD11c+CD11b+MHCIIlowCD80lowCD86low. Quanto à produção de PGE2, a fagocitose de AC+PAMP por células dendríticas foi capaz de induzir níveis elevados deste mediador lipídico nas diferentes proporções de células apoptóticas utilizadas. Os níveis de PGE2 encontrados no sobrenadante de cultura foi proporção dependente evidenciando uma relação direta entre fagocitose de AC+PAMP e a produção de PGE2. A fim de mimetizar a cinética da carga bacteriana durante uma infecção, ou seja, inicialmente uma menor carga bacteriana que tende a aumentar conforme ocorre a colonização, os animais foram inoculados com alta (high - 106 UFC de E. coli) e baixa (low - 105 UFC de E. coli) cargas bacterianas, gerando desta forma o que chamamos de AC+PAMPhigh e AC+PAMPlow, respectivamente. Os resultados aqui apresentados demonstram que diante de uma alta carga bacteriana há uma maior produção dos mediadores

Expansão ex vivo de células uroteliais em scaffolds 3D

Malformations and possible damages to the urogenital system can be originated in the embryonic period. Moreover, fire guns, knives and accidents, where there is the disruption of the urethra, also cause these lesions. The objective was to analyze the contribution of tissue engineering in the construction of neo-urethra, developed by bioengineering. We performed an urothelial ex vivo expansion of cells in 3D scaffolds (platelet gel matrix and acellular porcine aorta) to assess the contribution of this technique in the construction of a neo-urethra. Mechanical dissociation was made of the inner wall of 10 North Folk rabbit’s bladder, weighing 2.5 to 3.0 kg. After dissociation the cell content was centrifuged and obtained a pellet of urothelial cells. The pellet was ressuspended in culture medium DMEM F12 and cells were maintained in culture for 15 days. Immunohistochemical analysis characterized the urothelial culture. The cells were then implanted in the scaffold - platelet gel. In a second experiment using aortic porcine acellular matrix were implanted urothelial cells alone and urothelial cells on platelet gel, on the inner wall of the scaffold - aorta, with space for setting bordered by a urethral probe. The complex probe - cells - aorta and probe - cells in platelet gel - aorta, were sealed with suture material and culture were maintained in a humidified 37ºC incubator with 5% CO2 in air for 12 days to subsequent histological analysis of urothelium cell adhesion to the scaffolds. By observation under an optical microscope, we could see the growth of cells in the scaffold platelet gel, from a monolayer in to a three-dimensional structure. In the acellular porcine aortic matrix containing the platelet gel, we could observe a few quantity of urothelial cells adhered. However with the acellular porcine aortic matrix in which was implanted only the urothelial cells, we have obtained adhesion to the wall

Influência da luz LED em culturas de células-tronco mesenquimais in-vitro

Stem cells are defined as cells capable of self-renewal and differentiation into specialized cells when submited to external signalings in the enviroment. Among adult stem cells, mesenchymal cells occupy an important position because they can differentiate into mesodermal cells such as osteoblasts, adipocytes and chondrocytes. Cell therapy consists in the use of mesenchymal stem cells (MSC) in the treatment of degenerative diseases and harmed tissue reconstruction. Due to the longstanding and costly procedure for cultivation of MSC, it was proposed the use of low power light sources, such as light emitting diodes (LED), to optimize these factors. Recent works have shown a series of results from the influence of LED light on biological tissues such as increased rate of cell proliferation, increased RNA, DNA and ATP synthesis rate. The purpose of this study is to compare the biomodulator effect of LED light set at wavelengths 630nm ± 10nm and 805nm ± 10nm on the mesenchymal stem cells proliferation. For this, the mesenchymal stem cells culture adopted the procedure used in the Departament of Animal Reproduction and Veterinary Radiology of the Faculty of Veterinary Medicine and Animal Sciences of Botucatu. MSC were obtained from an adult horse bone marrow, and isolated by density gradient separation, with the FICOLL reagent and by centrifugation. The pellet containing the stem cells was removed and these were placed in low glucose DMEM culture medium, containing 10% fetal calf serum and antibiotics. The material was observed daily by inverted microscopy for monitoring the progression of the cells and subsequently the amount of cells were counted in a Neubauer counting chamber. The amount of MSC was obtained by cell culture seeded in 24 wells culture plate and segregated into three distinct groups: Group 1 was irradiated with wavelength set at 630nm ± 10 nm, Group... (Complete abstract click electronic access below)

Caracterização e infecção de células tronco mesenquimais e células semelhantes a neurônios, derivadas de cordão umbilical bovino pelo herpesvírus bovino Tipo 5 (BoHV-5)

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Avaliação dos efeitos de aloe-emodin em células uroteliais imortalizadas e neoplásicas cultivadas in vitro

Aloe vera (Aloe barbadensis Miller), popularly known in Brazil as babosa, has a long history of use as medicinal plant for different therapeutic purposes. The components of the plant extract are present in various products of human use, mainly for nutritional and cosmetics purposes. However, some studies suggest that this extract might also have carcinogenic activity. The aloe vera extract is a complex mixture of bioactive compounds. The study of isolated compounds may contribute to elucidate contradictory results about the effects related to the consumption of the plant, as well as their mechanisms of action. One of the most important compound from Aloe vera is aloe-emodin, which is a secondary metabolite generated in the intestinal tract. Putative antimicrobial and antitumor effects were previously attributed to aloe-emodin. Although the exposure of urothelial cells to aloe-emodin was already reported in the literature, only one study showed its effects on urothelial cells, suggesting that aloe-emodin inhibits the viability of T24 cancer cells due to apoptosis induction. Since there is no sufficient information about the effects of aloe-emodin on urothelial cells, and low efficiency in the treatment of bladder cancer currently, the present study aims to evaluate the hypothesis that the treatment with aloe-emodin could impact the behavior of other urothelial cell lines in vitro. Therefore, the in vitro IC50 exposure of aloe-emodin to human immortalized neoplastic urothelial cells will be determinated in order to verify possible differences in the behavior of urothelial cells in vitro treated with aloe-emodin in comparison with untreated cells. Furthermore, differences between cell lines will be also evaluated

TLR 2, TLR 4, ROS, óxido nítrico, P38 e IKK em células mononucleares de cães com leishmaniose Visceral tratadas com o imunomodulador P-MAPA: Larissa Martins Melo. -

Pós-graduação em Ciência Animal - FMVA

Efeito citotóxico de agentes clareadores a base de peróxido de hidrogênio a 20% e 38% sobre células odontoblastóides

The aim of this in vitro study was to evaluate the trans-enamel and transdentinal cytotoxic effects of two in-office tooth bleaching techniques that employ bleaching gels containing 20% and 38% of H2 O2 on cultured odontoblast-like cell line (MDPC-23). Sixty enamel/dentin discs were obtained from bovine central incisors and placed individually in artificial pulp chambers. Six groups were formed according to the following enamel treatments: G1- 20% H2 O2 (1 application); G2- 20% H2 O2 (2 applications); G3- 38% H2 O2 (1 application); G4- 38% H2 O2 (2 applications); G5- 38% H2 O2 (3 applications); and G6- control (no treatment). In G1 and G2, the bleaching gel was left in contact with the enamel surface for 45 min in each application. However, in G3, G4, and G5 the bleaching gel was applied for only 10 min per application. After the last application, the extracts were collected and applied on previously cultured cells (30.000 cells/cm2 ) for 24 h. Cell metabolism was evaluated by the MTT assay and cell morphology was analysed by scanning electron microscopy. Cell metabolism decreased by 96.29%; 96.11%; 96.42%; 95.62%; and 97.18% in G1, G2, G3, G4, and G5, respectively. All treated groups differed significantly from non-treated control group (G6) (p < 0.05). However, the difference in cell metabolism among treated groups was not significant statistically. In addition, significant morphological cell alterations were observed in all treated groups. Under the tested experimental conditions, the extracts collected after both tooth bleaching techniques evaluated in this study caused severe toxic effects on cultured odontoblast-like cell MDPC-23.

Citotoxicidade transamelodentinária de uma fita clareadora com 10% de peróxido de hidrogênio sobre células de linhagem odontoblástica

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Interação de fibroblastos associados ao câncer em co-cultura com linguagens tumorias mamárias e a resposta terapêutica em o uso da melatonina

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Análise celular e molecular da ação do peptídeo Ac2-26 da proteína anti-inflamatória Anexina A nas células de carcinoma de colo de útero

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Análise in vitro da proliferação celular e produção de mediadores inflamatórios por células da conjuntiva estimuladas por materiais utilizados na confecção de próteses oculares

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Desenvolvimento do cultivo 3D a partir de células primarias de neoplasias mamárias caninas: estudo da apoptose sobre efeito ou não da carboplatina

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)