Correlação entre os efeitos da radioterapia sobre a atividade osteogênica de células mesenquimais na instalação de implantes
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Data
2018-03-26
Autores
Godoi, Fernanda Herrera Costa
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Universidade Estadual Paulista (Unesp)
Resumo
A sobrevida de pacientes submetidos a tratamentos para câncer de cabeça e pescoço está aumentando e a busca por reabilitação é necessária para promover qualidade de vida. O objetivo deste estudo é avaliar o perfil da expressão gênica de transcritos relacionados na osteogênese e osteoclastogênese em uma cultura primária de células mesenquimais de fêmures de ratos submetidos a radioterapia e colocação de implantes de titânio. Setenta e dois ratos receberam implantes de titânio em ambos fêmures. Os animais foram divididos em quatro grupos: 1) Grupo controle (C): cirurgia de colocação dos implantes; 2) Implante + irradiação imediata (IrI): irurgia de implantes e seguido de irradiação imediata; 3) Implante + irradiação tardia (IrT): cirurgia de implantes e irradiação após 4 semanas; 4) Irradiação prévia + implantes (IrP): irradiação e após 4 semanas cirurgia de implantes. A dose de irradiação foi de 30 Gy fracionadas em duas sessões. As eutanásias nos períodos de 3, 14 e 49 dias após o tratamento. A células cultivadas sofreram diferenciação em osteoblastos. A expressão gênica dos genes Fosfatase alcalina (Alp), Colágeno 1 (Col-1), Integrina !1 (Itg ! ), Osteocalcina (Bglap), Osteopontina (Osp), Osteonectina (Osn), Sialoproteína Óssea (Bsp), Fator de crescimento transformador (Tgf- ! ), Osterix (Osx),Fator relacionado ao Runt (Runx2), Fator estimulador de colônias de macrófagos (M-csf), Interleucina -6 (Il-6) Apolipoproteína E (Apo-E), Prostaglandina E2 (PgE2), foram avaliados por qRT-PCR e os resultados validados por ELISA. A expressão mais alta de Alp foi encontrada no grupo IrP (p=0.0001) e foi subexpressa nos grupos IrI e IrT (p<0.0001 e p=0.0041 respectivamente). Resultados similares foram encontrados nos transcritos de Itg !, On, Bsp, Osx e Runx2. mRNA do Tgf- ! foi hiperexpresso em todos os grupos principalmente aos 49 dias. Depois de 49 dias, os níveis de proteína da Bglap e Il-6 foram correlacionados com a expressão do mRNA. A radioterapia imediata altera a atividade de diferenciaçãO das células mesenquimais dos fêmures de ratos.
Prognosis of patient submitted to head and neck cancer treatment has increased and the oral rehabilitation becomes necessary to improve their life quality. The aim of this study was to evaluate the gene expression profile of transcripts related to osteogenesis and osteoclastogenesis in primary culture of mesenchymal cells from rat femurs submitted to radiotherapy and installation of pure titanium implant. Seventy two rats received titanium implants in both femurs. The animals were divided in four groups: 1- Control (C) implant surgery; 2- Implant + immediate irradiation after 24 hours (IrI); 3- Implant + late irradiation after 4 weeks (IrL); 4- Implant + Previous irradiation 4 weeks before surgery (IrP) irradiation. The irradiation dose was 30Gy fractioned in two. The animals were euthanized in day 3, 14 and 49 after surgery. The mesenchymal cells from femurs were extracted and cultivated. The differentiation into osteoblastic cells was verified by calcification nodules formation. The gene expression of Alkaline Phosphatase (Alp), Collagen 1 (Col 1), Integrin β1 (ItgB1), Osteocalcin (Bglap), Osteopontin (Osp), Osteonectin (Osn), Bone Sialoprotein (Bsp), Transforming Growth Factor β-type (Tgf-β), Osterix (Osx), Runt-related transcription factor 2 (Runx2), Macrophage Colony- Stimulating Factor (M–csf), Interleukin-6 (Il-6), Apolipoprotein E (ApoE) and Prostaglandin E2 (PgE2) were evaluated by qRT-PCR and the results were validated by ELISA test. Higher mRNA of Alp expression was found in IrP group (p=0.0001) and it was downregulated in IrI and IrT groups (p<0.0001 and p=0.0041, respectively). Similar results were found for transcript levels of ItgB1, Osn, Bsp, Osx and Runx2. mRNA of Tgf-β was overexpressed in all groups mainly in 49 days. After 49 days, Osn and Bsp transcripts were downregulated in the 3 groups evaluated. The Bglap and IL-6 protein level was correlated to their mRNA expression. The radiotherapy alters immediately the differentiation and activity of mesenchymal cells from rat femurs; however these cells seem to recover becoming suitable for receiving implants.
Prognosis of patient submitted to head and neck cancer treatment has increased and the oral rehabilitation becomes necessary to improve their life quality. The aim of this study was to evaluate the gene expression profile of transcripts related to osteogenesis and osteoclastogenesis in primary culture of mesenchymal cells from rat femurs submitted to radiotherapy and installation of pure titanium implant. Seventy two rats received titanium implants in both femurs. The animals were divided in four groups: 1- Control (C) implant surgery; 2- Implant + immediate irradiation after 24 hours (IrI); 3- Implant + late irradiation after 4 weeks (IrL); 4- Implant + Previous irradiation 4 weeks before surgery (IrP) irradiation. The irradiation dose was 30Gy fractioned in two. The animals were euthanized in day 3, 14 and 49 after surgery. The mesenchymal cells from femurs were extracted and cultivated. The differentiation into osteoblastic cells was verified by calcification nodules formation. The gene expression of Alkaline Phosphatase (Alp), Collagen 1 (Col 1), Integrin β1 (ItgB1), Osteocalcin (Bglap), Osteopontin (Osp), Osteonectin (Osn), Bone Sialoprotein (Bsp), Transforming Growth Factor β-type (Tgf-β), Osterix (Osx), Runt-related transcription factor 2 (Runx2), Macrophage Colony- Stimulating Factor (M–csf), Interleukin-6 (Il-6), Apolipoprotein E (ApoE) and Prostaglandin E2 (PgE2) were evaluated by qRT-PCR and the results were validated by ELISA test. Higher mRNA of Alp expression was found in IrP group (p=0.0001) and it was downregulated in IrI and IrT groups (p<0.0001 and p=0.0041, respectively). Similar results were found for transcript levels of ItgB1, Osn, Bsp, Osx and Runx2. mRNA of Tgf-β was overexpressed in all groups mainly in 49 days. After 49 days, Osn and Bsp transcripts were downregulated in the 3 groups evaluated. The Bglap and IL-6 protein level was correlated to their mRNA expression. The radiotherapy alters immediately the differentiation and activity of mesenchymal cells from rat femurs; however these cells seem to recover becoming suitable for receiving implants.
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Palavras-chave
Implantes de titânio, Célula mesenquimal, Radioterapia, Osseointegração, Titanium implants, Mesenchymal cell, Radioterapy