Collection and culture of human connective tissue cells from gingival explant technique for oral tissue bioengineering / Colección de células de tejido conectivo humano a partir de la técnica de explante gingival para bioingeniería de tejidos orales

SUMMARY: Cell culture is an important tool in medical, odontological and biological research laboratories, supporting cell therapies and tissue bioengineering strategies. Gingival fibroblasts present structural function, being able to modulate their metabolic capacity, which is reflected in the tissue morphology. The possibility of culturing fibroblasts in vitro, in monolayer or on three-dimensional scaffolds, for subsequent transplants in vivo opens important perspectives for the periodontal surgical clinic. The objective of the present article is to present a method of obtaining and cultivating viable human gingival fibroblasts for in vitro research. Explants derived from periodontal surgical discards were used, grown in 25 cm2 bottles to obtain a primary cell culture. After observing the proliferation and growth of the fibroblasts that interconnected and formed a monolayer network, involving the periphery of the explants, it was possible to remove the explants, to make the passage and the new subcultures were obtained in a ratio of 1:1. After 7 days, the amount of viable cells was analyzed in triplicate, using the Neubauer chamber technique, in cell culture bottles of 25 mm2 (T25) and 75 mm2 (T75). Fibroblasts were described and subclassified morphologically. The results showed a growth pattern in both bottles, but with a larger number in bottles of 75 cm2. Cells with fibroblastic morphology were subclassified into reticular and fusiform, being predominant those with fusiform morphology. In conclusion, culture of explant of human gingival connective tissue is a viable method for obtaining gingival connective tissue cells suitable for laboratory tests in cell culture, aiming at obtaining constructs for gingival tissue engineering.

RESUMEN: El cultivo celular es una herramienta importante en los laboratorios de investigación médica, odontológica y biológica, que apoyan las terapias celulares y las estrategias de bioingeniería de tejidos. Los fibroblastos gingivales presentan una función estructural, pudiendo modular su capacidad metabólica, que se refleja en la morfología tisular. La posibilidad de cultivar fibroblastos in vitro, en monocapa o en andamios tridimensionales, para trasplantes posteriores in vivo abre perspectivas importantes para la clínica de cirugía periodontal. El objetivo del presente artículo es presentar un método para obtener y cultivar fibroblastos gingivales humanos viables para investigación in vitro. Se utilizaron explantes derivados de los descartes quirúrgicos periodontales, crecidos en frascos de 25 cm2 para obtener un cultivo de células primarias. Después de observar la proliferación y el crecimiento de los fibroblastos que se interconectaron y formaron una red de monocapa, que involucraba la periferia de los explantes, fue posible eliminar los explantes, hacer el pasaje y los nuevos subcultivos se obtuvieron en una proporción de 1:1. Después de 7 días, la cantidad de células viables se analizó por triplicado, utilizando la técnica de cámara de Neubauer, en botellas de cultivo celular de 25 mm2 (T25) y 75 mm2 (T75). Los fibroblastos fueron descritos y sub-clasificados morfológicamente. Los resultados mostraron un patrón de crecimiento en ambas botellas, pero con un número mayor en botellas de 75 cm2. Las células con morfología fibroblástica se subclasificaron en reticulares y fusiformes, predominando aquellas con morfología fusiforme. En conclusión, el cultivo de explante de tejido conectivo gingival humano es un método viable para obtener células de tejido conectivo gingival adecuadas para pruebas de laboratorio en cultivos celulares, con el objetivo de obtener construcciones para la ingeniería del tejido gingival.

Histometry Analysis of Osteoclasts, Osteoblasts and Fibroblasts in Immediate Placement Implants with Addition of Aloe Vera Extract

Objective: To analyze the effect of immediate placement of implants with extract from the new bone formation histometically. Material and Methods: In this true-experimental design with randomized post test control group, 9 mongrel dogs weighing 10 to 12 kg were used, which were divided into 3 groups, based on observation time of 14 days, 28 days and 56 days. On the installation of implants (∅3.5x10 mm) sequentially, the former socket extraction of the lower jaw's right second premolar tooth in the study sample injected 10% Aloe vera gel extract and left second left premolar tooth without injection of 10% Aloe vera extract. To compare independent groups use the Mann-Whitney test. All analysis were carried out using SPSS version 20. Results: There was an increase in the number of osteoblast cells in both treatment and control groups, but the value of the treatment group was greater. There were significant differences in the number of osteoblast cells between the treatment and control groups 14 days (p=0.019), 28 days: (p=0.018), and 56 days (p=0.009). There were no significant differences in the number of fibroblast cells between the treatment and control groups (p>0.05). But at observations 28 and 56 days, it was showed a significant difference in the number of fibroblast cells between the treatment and control groups (p=0.353 and p=0.024, respectively). Conclusion: Immediate placement of implants with 10% Aloe vera extract gel on extracted socket increases the number of osteoblasts and suppresses the number of osteoclasts and fibroblasts.

Biocompatibility of four root canal sealers: a histopathological evaluation in rat subcutaneous connective tissue

The aim of this study was to evaluate the subcutaneous biocompatibility of: Epiphany, AH Plus, Pulp Canal Sealer and Sealapex root canal sealers. Sixty rats were randomly assigned to 4 groups, according to the sealer. Polyethylene tubes containing the tested materials were inserted into the connective tissue. The implants were removed after 7, 15 and 30 days, and the tissue samples were processed, stained and examined by light microscopy. The descriptive analysis considered: thickness of the fibrous capsule, severity of the inflammatory reaction, and presence of giant cells. After 7 days, all sealers induced moderate to severe inflammatory reaction. After 15 days, Epiphany and AH Plus sealers showed a moderate inflammatory reaction, while Pulp Canal Sealer and Sealapex induced severe and mild inflammatory reactions, respectively. After 30 days, mild inflammatory reactions were observed for Epiphany, Sealapex and AH Plus. Sealapex induced the lowest inflammatory response at all evaluation periods, and only Pulp Canal Sealer did not show a decreased in the inflammatory reaction over time.

O objetivo deste estudo foi avaliar a biocompatibilidade subcutânea de cimentos endodônticos radiculares: Epiphany, AH Plus, Pulp Canal Sealer e Sealapex. Sessenta ratos foram divididos aleatoriamente em 4 grupos, de acordo com o cimento. Tubos de polietileno contendo os materiais testados foram inseridos no tecido conjuntivo. Os implantes foram removidos após 7, 15 e 30 dias, e as amostras de tecido foram processadas, coradas e examinadas por microscopia de luz. A análise descritiva considerou: espessura da cápsula fibrosa, severidade da reação inflamatória e presença de células gigantes. Após 7 dias, todos os cimentos induziram moderada e severa reação inflamatória. Após 15 dias, os cimentos Epiphany e AH Plus apresentaram uma reação inflamatória moderada, enquanto Pulp Canal Sealer e Sealapex induziram severa e leve reações inflamatórias, respectivamente. Após 30 dias, leve reação inflamatória foi observada para o Epiphany, Sealapex e AH Plus. Sealapex induziu a menor resposta inflamatória em todos os períodos de avaliação, e somente Pulp Canal Sealer não apresentou uma diminuição da reação inflamatória ao longo do tempo.