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Master's Dissertation
DOI
https://doi.org/10.11606/D.23.2021.tde-27082021-154856
Document
Author
Full name
Raphael da Silva Gomes
E-mail
Institute/School/College
Knowledge Area
Date of Defense
Published
São Paulo, 2021
Supervisor
Committee
Sipert, Carla Renata (President)
Andrade, Flaviana Bombarda de
Caldeira, Celso Luiz
Nardino, Marcela Claudino da Silva
Title in Portuguese
Hidróxido de Cálcio na modulação de células da papila apical e ligamento periodontal in vitro
Keywords in Portuguese
Células-tronco
Hidróxido de Cálcio
Ligamento Periodontal
OPG
Papila Apical
RANKL
Abstract in Portuguese
A interação entre Receptor para a Ativação do Fator de Transcrição Nuclear kappaB (RANK), Ligante do ativador do Receptor do Fator Nuclear kappaB (RANKL), a Osteoprotegerina (OPG) e Estimulador de Colônias de Macrófagos (M-CSF), tem sido relatadas como as principais citocinas responsáveis para remodulação óssea. Considera-se de extrema relevância entender a relação entre eles nas células de papila apical (CPA) e ligamento periodontal (CLP) considerando que eles estão envolvidos no processo de revascularização pulpar e reabsorção óssea periapical em casos de periodontite apical. Sendo assim, o presente trabalho in vitro teve como objetivo esclarecer o papel do lipopolissacarídeo (LPS) nas CPA e CLP, detectar a expressão de RANKL e OPG e compreender a modulação dessas células. Para elucidar esses pontos, CPA e CLP foram estabelecidas em meio de cultura e estimuladas por LPS. Foi realizada análise de citotoxicidade por um ensaio de Brometo de 3-(4,5-dimetiliazol-2-il)-2,5-difenitetrazólio (MTT). As células então foram estimuladas com a concentração definida após o experimento anterior com 0,01 µg/mL e depois de 1 hora com concentrações de 1000 e 250 µg/mL de hidróxido de cálcio (Ca(OH)2) para avaliar e quantificar a expressão de RANK-L, M-CSF e OPG por um ensaio Enzyme Linked Immunosorbent Assay (ELISA). Foi realizado novamente ensaio de MTT para avaliação da citotoxicidade do LPS e do Ca(OH)2 . Os resultados demostraram aumento de OPG induzido por LPS em CPA, mas redução quando Ca(OH)2 foi incubado na sequência. A produção de OPG nas CLP não demonstrou diferenças estatisticamente significativas. RANKL e M-CSF não foram detectados durante o experimento. Como conclusão, este estudo demonstrou em células de papila apical que o LPS pode aumentar os níveis de OPG enquanto, o Ca(OH)2 pode diminuí-los. Esta modulação não foi detectada em células de ligamento periodontal. Sendo assim, LPS e Ca(OH)2 podem influenciar a modulação dos eventos moleculares de reabsorção óssea por células de papila apical.
Title in English
Calcium Hydroxide in the modulation of cells of the apical papilla and periodontal ligament in vitro
Keywords in English
Apical papilla
Calcium hydroxide
OPG
Periodontal ligament
RANKL
Stem cells
Abstract in English
The interaction between kappaB Nuclear Transcription Factor Activation Receptor (RANK), kappaB Nuclear Factor Receptor Activator Ligand (RANKL), Osteoprotegerin (OPG) and Macrophage Colony Stimulator (M-CSF), has been reported as the main cytokines responsible for bone remodeling. It is considered extremely important to understand the relationship between them in the cells of the apical papilla (CPA) and periodontal ligament (CLP) considering that they are involved in the process of pulp revascularization and periapical bone resorption in cases of apical periodontitis. Thus, the present in vitro study aimed to clarify the role of lipopolysaccharide (LPS) in CPA and CLP, detect RANKL and OPG expression and understand the modulation of these cells. To clarify these points, CPA and CLP were established in culture medium and stimulated by LPS. Cytotoxicity analysis was carried out by a 3- (4,5-dimethylazol-2- yl) -2,5-diphenitetrazolium (MTT) Bromide assay. The Cells were then stimulated to the defined concentration after the previous experiment with 0.01 µg / mL and after 1 hour with concentrations of 1000 and 250 µg / mL of calcium hydroxide (Ca (OH) 2) to evaluate and quantify the expression of RANK-L, M-CSF and OPG by an Enzyme Linked Immunosorbent Assay (ELISA) assay. CPA and CLP again by MTT assay to evaluate the cytotoxicity of LPS and Ca (OH) 2. The results showed an increase in OPG induced by LPS in CPA, but a reduction when Ca (OH) 2 was incubated in the sequence. The production of OPG in the PLCs did not show statistically significant differences. RANKL and M-CSF were not detected during the experiment. In conclusion, this study demonstrated in apical papilla cells that LPS can increase OPG levels while Ca (OH) 2 can decrease them. This modulation was not detected in cells of periodontal ligament. Thus, LPS and Ca (OH) 2 can influence the modulation of bone resorption molecular events by apical papilla cells.
 
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Publishing Date
2022-01-19
 
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