A cell viability study on an experimental model of human gingival fibroblasts cultured without foetal bovine serum

El objetivo del estudio es aportar más informaciones metodológicas y conceptuales a cerca del efecto del SBF 10% en ensayos de viabilidad celular sobre fibroblastos gingivales humanos, con la finalidad de optimizar las condiciones experimentales para la obtención de resultados más predictivos. Se utilizarán el ensayo colorimétrico WST-1 para la evaluación de proliferación celular, el ensayo de LDH para evaluación de la integridad de membrana celular, el ensayo de cuantificación de ADN mediante espectrometría y la microscopía óptica de contraste de fases para observar las alteraciones morfológicas. Entre 6 y 24 h, las células sufren poco con la ausencia del SBF 10% y no resulta imprescindible su utilización para evaluar proliferación celular y la integridad de membranas plasmática y nuclear. Sin embargo, la morfología de las células se altera, de forma intensa, a partir de 6 h de ausencia de SBF 10%, por lo que es conveniente utilizarlo para dichos estudios morfológicos.The aim of this study is to improve the methodological and conceptual knowledges about the effect of 10% foetal bovine serum in cell viability assays on human gingival fibroblasts, in order to optimize the experimental conditions to obtain more predictable results. In this study, different methods were used to determine cell proliferation, cell and nuclear membrane integrity and morphological study by using WST-1 assay, LDH assay, DNA quantification and phase contrast microscopy. Our results demonstrated that 10% foetal bovine serum deprivation for 6h up to 24h did not show changes in terms of cell proliferation, cell and nuclear membrane integrity. However, important morphological changes were observed under foetal bovine serum deprivation conditions after 6h. We conclude that, 10% foetal bovine serum does not result essential its utilization to evaluate cell proliferation and their deprivation do not compromise cell survival range and could be suppressed in cell proliferation, cell and nuclear membrane assays without affecting the assays results. In contrast, the utilization of 10% foetal bovine serum becomes crucial in morphological studies and we strongly recommend 10% foetal bovine serum supplement in morphological studies

Practical formation in histology. A study of the motivational priorizartion model between spanish and argentinian students

La incidencia de los factores motivacionales en la formación práctica universitaria ha sido escasamente investigada. En el presente trabajo, se investiga un conjunto de factores de carácter motivacional con el objeto de determinar el modelo de prioridad que establecen, en relación con la formación práctica de histología, los alumnos que cursan dicha materia en la Facultad de Odontología. El estudio se realiza en dos universidades: la de Granada en España y la Nacional de Córdoba en la República Argentina. Los resultados demuestran que, en la cesta motivacional investigada, los valores más elevados en ambos grupos de estudiantes -aunque significativamente más altos en el grupo argentino-, son los que hacen referencia a los deseos motivacionales relacionados con los intereses vinculados al futuro éxito profesional en el ejercicio de la odontología -utilidad diagnóstica y terapéutica y adquisición de habilidades-, observándose cifras más bajas en los factores relacionados con los deseos motivacionales vinculados a la autosatisfacción como la curiosidad o el placer del conocimiento. Se discuten los resultados en relación con la variable geográfica y la psicopedagogía de la motivación.The incidence of motivational factors on the practical formation of University students has not been studied in deep. In this work, we studied a group of motivational factors in order to determine the priority model that the Dentistry students establish in relation to their practical formation in Histology. This study was carried out in two universities: the University of Granada, Spain, and the National University of Cordoba, Argentina. The results showed that the highest scores of the motivational chart analyzed here, corresponded to those interests that were associated to the future professional success in the field of Dentistry -diagnostic and therapeutic usefulness and acquisition of abilities-, especially in students from Argentina. The lowest scores were found for factors related to motivational desires associated to self-satisfaction, such as curiosity and pleasure of knowledge. The results are discussed in relationship with the geographical origin and motivational psicopedagogy

Effects of Four Formulations of Prostaglandin Analogs on Eye Surface Cells. A Comparative Study

We evaluated the cytotoxic effects of four prostaglandin analogs (PGAs) used to treat glaucoma. First we established primary cultures of conjunctival stromal cells from healthy donors. Then cell cultures were incubated with different concentrations (0, 0.1, 1, 5, 25, 50 and 100%) of commercial formulations of bimatoprost, tafluprost, travoprost and latanoprost for increasing periods (5 and 30 min, 1 h, 6 h and 24 h) and cell survival was assessed with three different methods: WST-1, MTT and calcein/AM-ethidium homodimer-1 assays. Our results showed that all PGAs were associated with a certain level of cell damage, which correlated significantly with the concentration of PGA used, and to a lesser extent with culture time. Tafluprost tended to be less toxic than bimatoprost, travoprost and latanoprost after all culture periods. The results for WST-1, MTT and calcein/AM-ethidium homodimer-1 correlated closely. When the average lethal dose 50 was calculated, we found that the most cytotoxic drug was latanoprost, whereas tafluprost was the most sparing of the ocular surface in vitro. These results indicate the need to design novel PGAs with high effectiveness but free from the cytotoxic effects that we found, or at least to obtain drugs that are functional at low dosages. The fact that the commercial formulation of tafluprost used in this work was preservative-free may support the current tendency to eliminate preservatives from eye drops for clinical use.This study was supported by the Spanish Plan Nacional de Investigación Científica, Desarrollo e Innovación Tecnológica (I+D+I) from the Spanish Ministry of Economy and Competitiveness (Instituto de Salud Carlos III), grants FIS PI11/1582 and FIS PI14/0955 (co-financed by FEDER funds, European Union) and by Grant P10-CTS-6060 from the Consejería de Economía, Innovación, Ciencia y Empleo, Junta de Andalucia, Spain (Proyectos de Excelencia)

Quality control of dental adhesive system in an “in vitro” experimental model of human gingival fibroblasts

En este trabajo se evaluó la viabilidad de fibroblastos gingivales humanos en contacto con distintos sistemas adhesivos dentales mediante la utilización del método de cuantificación de LDH libre y de un análisis morfológico. Se utilizaron fibroblastos gingivales humanos cultivados en placas de 24 pocillos a una concentración de 20000 células/500 ìl de medio de cultivo DMEM con 10% FBS y antibióticos. Los materiales evaluados fueron: Adper SE Plus (ADSE), Adper Single Bond (SB) y One Coat (OC). A las 24 horas se realizó la cuantificación de la liberación de la enzima lactato deshidrogenasa (LDH) en un lector de placas ELX-800. Para el análisis morfológico las células adheridas en los pocillos fueron observadas con un microscopio óptico de luz invertida Nikon Japan Optifhot-2. Las células controles no recibieron tratamiento. Al evaluar LDH los valores fueron: ADSE 42,1%, SB 46,5%, OC 39,6%. A nivel morfológico las células en contacto con los distintos materiales usados en la experiencia mostraron alteraciones caracterizadas por células que toman formas esféricas y que presentan ruptura de sus membranas plasmáticas. En conclusión, los distintos sistemas adhesivos dentales utilizados generarían alteraciones irreversibles en la viabilidad celular de los fibroblastos gingivales humanos por la ruptura de sus membranas plasmáticas.The objetive of this study was to evaluate human gingival fibroblast viability in contact with different dental adhesive systems by means of both free LDH quantification method and morphological analyses. Human gingival fibroblast grow to in 24 wells plates to 20.000cells/500ul concentration DMEM culture medium whith 10% and antibiotics were used. Materials assessed were: Adper SE Plus (ADSE), Adper Single Bond (SB), One Coat (OC). After 24 hours deshidrogenasa lactate enzyme releasing quantification was performed. The analysis was carried out through ELX- 800 (Biotek) plates reader. For morphologic analysis cells adhered in the wells were observed by light microscopy Nikon Japan Optifhot-2. Control cells were not treated. When evaluating LDH, results were: ADSE 42,1%, SB 46,5%, OC 39,6%. At morphogical level cells in contact to different experimental materials showed some alterations characterized by cells which become spherical and present plasmatic membrane disruption. These results allow us to conclude that different dental adhesive systems used do creat or posibility cellular viability irreversible alterations in human gingival fibroblast due to plasmatic membrane disruption

Evaluation of the students' perceptions about the virtual self-learning process in a postgraduate professional program

Objetivos: Evaluar la percepción de los estudiantes de postgrado sobre la enseñanza virtual en tres cuestiones básicas: acceso al conocimiento, grado de satisfacción y preferencia temporal en enseñanza virtual y presencial. Metodología: El estudio se realizó a través de un cuestionario con tres grupos de ítems vinculados a los objetivos arriba indicados. Se utilizó la prueba t de Student de doble cola. Resultados y Discusión: Para los alumnos, la plataforma virtual permite acceder al profesor sin llegar a sustituirlo (p=0,0000), e igual ocurre con la ampliación de conocimientos, la cual no sustituye a los libros ni a otras fuentes (p=0,0003) (p=0,0158). La libertad de horario constituye la percepción más valorada, superando significativamente a la dificultad de uso de la plataforma (p=0,0000), la modificación del horario extralaboral (p=0,0652), el estrés académico (p=0,0020) o la conciliación de la vida familiar y laboral (p=0,0016). Los estudiantes, a pesar de la accesibilidad y la satisfacción mostrada sobre la enseñanza virtual, asignan más porcentaje a la enseñanza presencial que a la virtual, tanto en teoría como en práctica (p=0,0000) y (p=0,0000).Objectives: To evaluate the perception of postgraduate students on virtual learning in three topics: access to knowledge, satisfaction level, and time-distribution preferences in virtual and non-virtual education. Methodology: This study was carried out by using a questionnaire with three groups of items associated with the 3 objectives. The Student t test was used to analyze the results. Results and Discussion: The virtual platform allowed the students to access the instructors, but without replacing them (p=0.0000). Also the student's perception is that they can increase their knowledge without replacing books or other resources (p=0.0003) (p=0.0158). The freedom of work time was significantly more valued that the need to overcome the difficulty of using the platform (p=0.0000), changing the working hours (p=0.0652), academic stress (p=0.0020) and balancing work and family life (p=0.0016). Despite the accessibility and satisfaction with the platform, students gave more percentage of time to non-virtual learning than to virtual learning in both theory and practice (p=0.0000) and (p=0.0000)

Identification of Histological Patterns in Clinically Affected and Unaffected Palm Regions in Dupuytren's Disease

Dupuytren's disease is a fibro-proliferative disease characterized by a disorder of the extracellular matrix (ECM) and high myofibroblast proliferation. However, studies failed to determine if the whole palm fascia is affected by the disease. The objective of this study was to analyze several components of the extracellular matrix of three types of tissues—Dupuytren's diseased contracture cords (DDC), palmar fascia clinically unaffected by Dupuytren's disease contracture (NPF), and normal forehand fascia (NFF). Histological analysis, quantification of cells recultured from each type of tissue, mRNA microarrays and immunohistochemistry for smooth muscle actin (SMA), fibrillar ECM components and non-fibrillar ECM components were carried out. The results showed that DDC samples had abundant fibrosis with reticular fibers and few elastic fibers, high cell proliferation and myofibroblasts, laminin and glycoproteins, whereas NFF did not show any of these findings. Interestingly, NPF tissues had more cells showing myofibroblasts differentiation and more collagen and reticular fibers, laminin and glycoproteins than NFF, although at lower level than DDC, with similar elastic fibers than DDC. Immunohistochemical expression of decorin was high in DDC, whereas versican was highly expressed NFF, with no differences for aggrecan. Cluster analysis revealed that the global expression profile of NPF was very similar to DDC, and reculturing methods showed that cells corresponding to DDC tissues proliferated more actively than NPF, and NPF more actively than NFF. All these results suggest that NPF tissues may be affected, and that a modification of the therapeutic approach used for the treatment of Dupuytren's disease should be considered.This work was supported by CTS-115 (Tissue Engineering Group), University of Granada/Spain

Estudio de la expresión génica en la enfermedad periodontal y obtención de un modelo de matriz biológica descelularizada para la regeneración tisular guiada

Tesis Univ. Granada. Departamento de HistologíaEste trabajo ha sido financiado por los siguientes proyectos de investigación: FIS PI11/2680 del Fondo de Investigación Sanitaria del Instituto de Salud Carlos III (Ministerio de Economía y Competitividad) y FPS PI/0512/2011 de la Fundación Progreso y Salud (Junta de Andalucía)

Dossiê Temático: Engenharia de Tecidos em Odontologia

Included in this dossier are three articles that explore three interesting topics: the governmental policies on tissue engineering in Latin America, the approaches and methods used to elaborate a bio-tooth, and the applications of platelet-rich fibrin in regenerative dentistry.Se incluyen en este dossier tres artículos que exploran tres temas interesantes: las políticas gubernamentales en materia de ingeniería tisular en Latinoamérica, los enfoques y métodos empleados para elaborar un biodiente, y las aplicaciones de la fibrina rica en plaquetas en odontología regenerativa.Incluídos neste dossiê estão três artigos que exploram três tópicos interessantes: as políticas governamentais sobre engenharia de tecidos na América Latina, as abordagens e métodos usados para elaborar um bio-dente e as aplicações da fibrina rica em plaquetas na odontologia regenerativa

Generation and characterization of novel magnetic field-responsive biomaterials

We report the preparation of novel magnetic field-responsive tissue substitutes based on biocompatible multi-domain magnetic particles dispersed in a fibrin–agarose biopolymer scaffold. We characterized our biomaterials with several experimental techniques. First we analyzed their microstructure and found that it was strongly affected by the presence of magnetic particles, especially when a magnetic field was applied at the start of polymer gelation. In these samples we observed parallel stripes consisting of closely packed fibers, separated by more isotropic net-like spaces. We then studied the viability of oral mucosa fibroblasts in the magnetic scaffolds and found no significant differences compared to positive control samples. Finally, we analyzed the magnetic and mechanical properties of the tissue substitutes. Differences in microstructural patterns of the tissue substitutes correlated with their macroscopic mechanical properties. We also found that the mechanical properties of our magnetic tissue substitutes could be reversibly tuned by noncontact magnetic forces. This unique advantage with respect to other biomaterials could be used to match the mechanical properties of the tissue substitutes to those of potential target tissues in tissue engineering applicationPeer Reviewe

Sequential keratinocytic differentiation and maturation in a three-dimensional model of human artificial oral mucosa

Dupuytren's disease is a fibro-proliferative disease characterized by a disorder of the extracellular matrix (ECM) and high myofibroblast proliferation. However, studies failed to determine if the whole palm fascia is affected by the disease. The objective of this study was to analyze several components of the extracellular matrix of three types of tissues—Dupuytren's diseased contracture cords (DDC), palmar fascia clinically unaffected by Dupuytren's disease contracture (NPF), and normal forehand fascia (NFF). Histological analysis, quantification of cells recultured from each type of tissue, mRNA microarrays and immunohistochemistry for smooth muscle actin (SMA), fibrillar ECM components and non-fibrillar ECM components were carried out. The results showed that DDC samples had abundant fibrosis with reticular fibers and few elastic fibers, high cell proliferation and myofibroblasts, laminin and glycoproteins, whereas NFF did not show any of these findings. Interestingly, NPF tissues had more cells showing myofibroblasts differentiation and more collagen and reticular fibers, laminin and glycoproteins than NFF, although at lower level than DDC, with similar elastic fibers than DDC. Immunohistochemical expression of decorin was high in DDC, whereas versican was highly expressed NFF, with no differences for aggrecan. Cluster analysis revealed that the global expression profile of NPF was very similar to DDC, and reculturing methods showed that cells corresponding to DDC tissues proliferated more actively than NPF, and NPF more actively than NFF. All these results suggest that NPF tissues may be affected, and that a modification of the therapeutic approach used for the treatment of Dupuytren's disease should be considered.Peer Reviewe