TGF-β1 on induced osteogenic differentiation of human dermal fibroblast

PURPOSE:To evaluate the role of transforming growth factor beta 1 (TGF-β1) on the induced osteogenic differentiation of human dermal fibroblasts.METHODS:We performed four groups with cultured dermal fibroblasts according to the culture medium: CONTROL (DMEM culture medium); TGF-β1 (DMEM culture medium with 10 ng/ml of TGF-β1); OSTEOG (DMEM culture medium with 0.5 µg/ml of ascorbic acid, 10 mmol/l of β-glycerophosphate and 10 nmol/L of dexamethasone); and OSTEOG/TGF-β1 (osteogenic medium with 10 ng/ml of TGF-β1). Alkaline phosphatase (ALP) activity and the amount of osteocalcin (OC) in the supernatant, as well as the capability to form calcium phosphate deposits, were analysed for 28 dayRESULTS:There were significant differences (p<0.05) between CONTROL and TGF-β1 groups in comparison with OSTEOG and OSTEOG/TGF-β1 groups in the ALP activity and OC amount. Although, both osteogenic groups had the same behavior with regard the expression curve during the experimental time, the OSTEOG/TGF-β1 group achieved significantly higher ALP and OC levels and showed no significant difference in the levels of mineralized deposits and in comparison with the levels found in the OSTEOG group.CONCLUSION:The addition of transforming growth factor beta 1 to the osteogenic culture medium increased the activity of alkaline phosphatase and the amount of osteocalcin, but TGF-β1 did not alter the presence of mineralized calcium phosphate deposits.Federal University of São Paulo Department of SurgeryUniversidade Federal de São Paulo (UNIFESP) Department of SurgeryUNIFESP, Department of SurgeryUNIFESP, Department of SurgerySciEL

Adipose-derived stem cells (ADSC) in the viability of a random pattern dorsal skin flap in rats

PURPOSE:To evaluate the viability of random pattern dorsal skin flaps in rats after injection of adipose-derived stem cells (ADSC).METHODS: Thirty five adult male Wistar EPM rats (weight 250-300 g) were distributed, at random, in two groups. I- Control (flap elevation with injection of saline solution) with fifteen animals and II- Experimental (flap elevation with injection of ADSC ) with fifteen animal. The ADSC were isolated from others five adult male rats. A dorsal skin flap measuring 10x4 cm was raised and a plastic barrier was placed between the flap and its bed in both groups and the injection (cells or saline solution) were perfomed immediately after the surgery. The percentage of flap necrosis was measured on the seventh postoperative day.RESULTS:The ADSC were able to replicate in our culture conditions. We also induced their adipogenic, osteogenic and chondrogenic differentiation to verify their mesenchymal stem cells potentiality in vitro. The results were statistically significant showing that the ADSC decreased the area of necrosis (p<0.05).CONCLUSIONS:The cells demonstrated adipogenic, osteogenic and chondrogenic differentiation potential in vitro. The administration of adipose-derived stem cells was effective to increase the viability of the random random pattern dorsal skin flaps in rats.Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Federal University of São Paulo Department of SurgeryUniversidade Estadual de Santa Cruz Department of SurgeryUNIFESPUNIFESP, Department of SurgeryUNIFESPCNPq: 312356/2009-9SciEL

Use of Bone Marrow Aspirate Concentrate (BMAC) Associated with Hyperbaric Oxygenation Therapy in Maxillary Appositional Bone Reconstruction. A Randomized Clinical Trial

Objectives: The objective of this study was to evaluate bone reconstruction using xenograft alone and associated with bone marrow aspirate concentrate (BMAC) and hyperbaric oxygen therapy. Material and Methods: Twenty-four maxillary edentulous patients were randomly assigned into three groups: Control group (CG)—xenograft bone alone (n = 8); Group 1 (G1)—xenogeneic bone block combined with BMAC (n = 8), and Group 2 (G2)—xenogeneic bone block combined with BMAC and hyperbaric oxygenation (n = 8). Bone biopsies were harvested 6 months after grafting. Vital Mineralized Tissue (VMT), Non-vital Mineralized Tissue (NVMT), and Non-Mineralized Tissue (NMT) were measured. Computed tomography was also performed on three occasions T0 (preoperative), T4 (4 months postoperative), and T8 (8 months postoperative). The difference between T4 and T8 values with respect to T0 was used to determine the thickness level gain after 4 and 8 months, respectively. Results: The tomographic evaluation did not show significant differences between the groups either at 4 or at the 8 months postoperatively. Regarding the histomorphometric analysis, CG had the lowest percentages of VMT (36.58 ± 9.56%), whereas G1 and G2 had similar results (55.64 ± 2.83% and 55.30 ± 1.41%, respectively). Concerning NMT and NVMT levels, the opposite was observed, with CG levels of 51.21 ± 11.54% and 11.16 ± 2.37%, G1 of 39.76 ± 11.48% and 3.65 ± 0.87%, and G2 of 40.3 ± 11.48% and 4.10 ± 0.87%, respectively. Conclusions: The use of bone block xenograft associated with BMAC resulted in a significant increase of bone neoformation when compared to the xenograft alone, though hyperbaric oxygenation did not enhance the results.Odontologí

Development of experimental in vitro burn model

PURPOSE:To propose an experimental burn model in NIH-3T3 cell line.METHODS: Induction of thermal injury in cultures of mouse fibroblast - NIH-3T3- cell line and determination of cell viability by MTT and imunofluorescence.RESULTS: The heating of the Petri dish increased proportionally to the temperature of the base and the time of exposure to microwave. In this in vitro burn model, using the cell line NIH-3T3 was observed drastic cellular injury with significant changes in cell viability and activity. It showed drastically modified cell morphology with altered membrane, cytoskeleton and nucleus, and low cellularity compared to the control group.CONCLUSION: The burn model in vitro using the cell line NIH-3T3 was reproductive and efficient. This burn model was possible to determine significant changes in cell activity and decreased viability, with drastic change in morphology, cell lysis and death.Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Universidade Federal de São Paulo (UNIFESP)Universidade Estadual de Santa Cruz Department of Biological SciencesUNIFESPUNIFESPSciEL

Relação entre a densidade mineral óssea e a resistência à compressão de blocos ósseos xenógenos / Relationship between bone mineral density and compressive strength of xenogenous bone blocks

O objetivo deste estudo foi avaliar a densidade mineral óssea (DMO), a resistência final à compressão (RFC), e a eventual correlação entre os valores dessas variáveis, de quatro blocos ósseos xenógenos comercialmente disponíveis no Brasil para aplicações em implantodontia. Trinta e dois espécimes dos materiais de enxertia analisados foram divididos em 4 grupos de estudo (n = 8): Grupo 1 (G1), Biocollagen (Bioteck, Arcugnano, Itália); Grupo 2 (G2), Bio-Graft, (Geistilich, Wolhusen, Suíça); Grupo 3 (G3), OrthoGen (Baumer, Mogi Mirim, SP, Brasil); e Grupo 4 (G4), Bonefill (Bionnovation, Bauru, SP, Brasil). A DMO foi determinada por meio da análise da densidade óptica dos materiais, observada em imagens tomográficas de alta definição, e expressa em unidades Hounsfield (HU). A RFC foi determinada por meio de um ensaio mecânico de compressão e expressa em Newton (N). Os valores de DMO encontrados para os grupos G1, G2, G3 e G4 foram significativamente diferentes, a saber, 354,3 HU, 317,7 HU, 206,5 HU e 145,6 HU, respectivamente (p ? 0,05). Os valores de RFC encontrados para os grupos G1, G2, G3 e G4 também foram significativamente diferentes, a saber, 685,18 N, 563,18 N, 915,20 N e 1399,70 N, respectivamente (p ? 0,05). Observou-se uma correlação positiva e moderada entre a DMO e a RFC apenas no G4 (p = 0,015; r2 = 0,655). A análise dos resultados demonstrou que os blocos de enxerto ósseo xenógenos com uma menor DMO tendem a ter uma maior RFC

TGF-beta1 in human dermal fibroblasts cultured in a collagen sponge

Introduction: The research of new sources of cells for Tissue Engineering of the human bone, it is necessary because the use of the primary choice for this therapy (bone marrow cell), can result in a morbidity of the donor area and poor expansion in vitro. Therefore, it is important to seek other cellular sources to contribute to this therapy. Objective: To evaluate the influence of transforming growth factor-beta 1 (TGF-â1) in the osteogenic differentiation of human dermal fibroblasts cultured in a collagen sponge. Methods: The collagen sponges were cut in 16x2-mm sections and distributed into four groups according to the culture medium: CONTROL (DMEM culture medium); TGF-â1 (DMEM culture medium with 10 ng/ml of TGF-â1); OSTEOG (DMEM culture medium with 0.5 ìg/ml of ascorbic acid, 10 mmol/l of â-glycerophosphate and 10 nmol/L of dexamethasone); and OSTEOG.TGF-â1 (osteogenic medium with 10 ng/ml of TGF-â1). Measurements of the alkaline phosphatase (ALP) activity and the amount of osteocalcin (OC) in the supernatant, as well as measurement of the penetration of cells in the sponge by histology and measurement of calcium phosphate deposits by the Von Kossa test, were performed on days 2, 7, 14, 21, 28, and 56 . Results: ALP activity and OC level: There were no differences between the CONTROL and TGF-â1 groups in any of the measurements between any of the measurement points. However, the measured values were significantly lower than the OSTEOG and OSTEOG.TGF-â1 groups. The OSTEOG.TGF-â1 group achieved significantly higher. Interaction Cell/Sponge: in all groups the cells were at the top of sponge in the beginning and there were a penetration into the sponge up to the end of the experiments. Deposits of Calcium Phosphate: There were no presence of deposits in the CONTROL and TGF-â1 groups. There were evidence of the deposits in the OSTEOG. and OSTEG. TGF-â1 groups from the 14° day up to 56° day. Conclusion: TGF-â1 in osteogenic medium increased the ALP activity and the OC levels but did not influence the interaction cell/sponge and the presence of mineralized deposits of calcium phosphate into the collagen sponge.Introducao: A busca por uma fonte alternativa de celulas para a Engenharia Tecidual ossea, se deve a morbidade da area doadora e maior dificuldade de expansao in vitro quando da utilizacao de celulas de linhagem osteoblastica, como as celulas da medula ossea. Portanto, a busca de outras fontes celulares para contribuir para esta terapia, se torna fundamental. Objetivo: Avaliar a influencia do fator de crescimento transformante beta1(TGF-ƒÀ1) na diferenciacao celular de fibroblastos dermicos humanos cultivados em esponja de colageno. Metodos: As esponjas de colageno foram cortadas em pecas de 16mm de diametro e 2 mm de espessura sendo distribuidas em quatro grupos denominados de acordo com o meio de cultura: CONTROLE (Padrao); TGF-ƒÀ1 (Padrao acrescido de 10ng/mL de TGF-ƒÀ1); OSTEOG. Padrao acrescido de 0,5ƒÊg/mL de acido ascorbico, 10 mMol/L de ƒÀ-glicerofosfato e 10nMol/L de dexametasona) e OSTEOG.TGF- ƒÀ1(meio osteogenico acrescido de 10ng/mL de TGF-ƒÀ1). As avaliacoes da atividade da fosfatase alcalina (FAL) e quantidade de osteocalcina (OC) foram realizadas no sobrenadante. A interacao das celulas com a esponja de colageno foi avaliada pela exame histologico e a presenca de depositos de calcio fosfato realizada pelo teste de Von Kossa, todas as avaliacoes foram realizadas no segundo, setimo, 14 ‹, 21 ‹, 28 ‹ e 56 ‹dias. Resultados: Atividade da FAL e Quantidade de OC: nao houve diferenca entre os grupos CONTROLE e TGF- ƒÀ1 em nenhuma das avaliacoes e em nenhum dos pontos de avaliacao, no entanto na comparacao com os outros dois grupos, os valores foram significativamente menores, o grupo OSTEOG.TGF- ƒÀ1 obteve valores significativamente. Interacao Celula/Esponja: em todos os grupos as celulas inicialmente estavam na superficie com posterior penetracao para o interior da esponja. Depositos Mineralizados: nao existiu a presenca nos grupos CONTROLE e TGF- ƒÀ1 e nos grupos OSTEOG. e OSTEOG. TGF- ƒÀ1 existiu a presenca de depositos a partir do 14 ‹ dia ate o 56 ‹ dia. Conclusao: O TGF-ƒÀ1 no meio osteogenico, na cultura de fibroblastos dermicos humanos em esponja de colageno, aumentou a atividade da FAL e a quantidade de OC, nao alterando a interacao celula/esponja e a presenca de depositos mineralizados de calcio fosfato na estrutura da esponja de colagenoTEDEBV UNIFESP: Teses e dissertaçõe