Efeito da terapia fotodinâmica no fenótipo tronco de linhagens celulares de carcinoma epidermóide de boca

Detalhes bibliográficos
Autor(a) principal: Pinto, Marlene Aparecida Ferreira
Data de Publicação: 2021
Tipo de documento: Tese
Idioma: por
Título da fonte: Biblioteca Digital de Teses e Dissertações da Uninove
Texto Completo: http://bibliotecatede.uninove.br/handle/tede/2664
Resumo: Among the various types of cancer that affect the oral cavity, squamous cell carcinoma of the mouth (OSCC) is the most prevalent malignant neoplasm. Despite advances in treatment, patients with OSCC often present metastasis at diagnosis and recurrence, both associated with a poor prognosis. Recent studies have shown that the presence of cancer stem cells (CTC) are responsible for tumor growth and resistance to conventional therapy. An alternative for the treatment of CEB is Photodynamic Therapy (PDT). However, the effect of PDT on CTC in the CEB is unknown. Thus, this study aimed to evaluate in vitro the effect of PDT mediated by the photosensitizer 5-aminoulevulinic acid (5-ALA) on cell viability, maintenance of the stem cells phenotype, cell migration and expression of the genes Bmi1, Clagranulin, IVL and E- cadherin in two cell lines deriveted of OSCC (Ca1 and Luc4). All in vitro assays were conducted with both cell lines, which were divided into the following groups: Control, 5-ALA, LED 6J/cm² and PDT (5-ALA+6J/cm2). Cell viability was assessed by MTT and Neutral Red assays and the stem phenotype was assessed by flow cytometry to analyze the expression of CD44 and ESA to identify the CD44high/ESAhigh, CD44high/ESAlow and CD44low populations, as well as by colony and tumor sphere formation assays. Additionally, the PpIX production in each CTC population was evaluated. Cell migration was analyzed by wound healing assay and gene expression was analyzed by RT-qPCR. Both cell lines showed a significant reduction in cell viability after PDT. A percentage reduction of CD44high/ESAhigh cells was observed after PDT when compared to the other groups, which was followed by an increase in the percentage of cells with the CD44low phenotype, indicating cell differentiation induction. Additionally, reduced colony formation capacity and tumoral sphere formation after PDT. A CD44high / ESAhigh population of both cell lines had greater production of PpIX, indicating greater sensitivity of them to PDT. Cells from the Ca1 and Luc4 cell lines of the 5-ALA and PDT groups had less capacity to migrate compared to the other groups. An analysis of gene expression after PDT, showed a decrease in Bmi1 expression and increased Calgranulin expression in the Ca1 cell line, confirming the loss of the stem potential and induction of cell differentiation. Thus, the results shown here, indicate that a 5-ALA-PDT is capable of decreasing the cell viability and cell lines migration derived from OSCC, as well as reducing the percentage of cells with stem phenotype, inducing their differentiation.
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spelling Rodrigues, Maria Fernanda Setúbal Destrohttp://lattes.cnpq.br/7153553682954852Pavanni, Christianehttp://lattes.cnpq.br/2148910222765744Rodrigues, Maria Fernanda Setúbal Destrohttp://lattes.cnpq.br/7153553682954852Pavani, Christianehttp://lattes.cnpq.br/2148910222765744Dellê, Humbertohttp://lattes.cnpq.br/7435662740477057Franco, Adriana Lino dos Santoshttp://lattes.cnpq.br/7295388304888384Nunes, Fabio Daumashttp://lattes.cnpq.br/4909755821591847http://lattes.cnpq.br/9592750937209234Pinto, Marlene Aparecida Ferreira2021-11-04T17:38:34Z2021-08-04Pinto, Marlene Aparecida Ferreira. Efeito da terapia fotodinâmica no fenótipo tronco de linhagens celulares de carcinoma epidermóide de boca. 2021. 77 f. Tese( Programa de Pós-Graduação em Biofotônica Aplicada às Ciências da Saúde) - Universidade Nove de Julho, São Paulo.http://bibliotecatede.uninove.br/handle/tede/2664Among the various types of cancer that affect the oral cavity, squamous cell carcinoma of the mouth (OSCC) is the most prevalent malignant neoplasm. Despite advances in treatment, patients with OSCC often present metastasis at diagnosis and recurrence, both associated with a poor prognosis. Recent studies have shown that the presence of cancer stem cells (CTC) are responsible for tumor growth and resistance to conventional therapy. An alternative for the treatment of CEB is Photodynamic Therapy (PDT). However, the effect of PDT on CTC in the CEB is unknown. Thus, this study aimed to evaluate in vitro the effect of PDT mediated by the photosensitizer 5-aminoulevulinic acid (5-ALA) on cell viability, maintenance of the stem cells phenotype, cell migration and expression of the genes Bmi1, Clagranulin, IVL and E- cadherin in two cell lines deriveted of OSCC (Ca1 and Luc4). All in vitro assays were conducted with both cell lines, which were divided into the following groups: Control, 5-ALA, LED 6J/cm² and PDT (5-ALA+6J/cm2). Cell viability was assessed by MTT and Neutral Red assays and the stem phenotype was assessed by flow cytometry to analyze the expression of CD44 and ESA to identify the CD44high/ESAhigh, CD44high/ESAlow and CD44low populations, as well as by colony and tumor sphere formation assays. Additionally, the PpIX production in each CTC population was evaluated. Cell migration was analyzed by wound healing assay and gene expression was analyzed by RT-qPCR. Both cell lines showed a significant reduction in cell viability after PDT. A percentage reduction of CD44high/ESAhigh cells was observed after PDT when compared to the other groups, which was followed by an increase in the percentage of cells with the CD44low phenotype, indicating cell differentiation induction. Additionally, reduced colony formation capacity and tumoral sphere formation after PDT. A CD44high / ESAhigh population of both cell lines had greater production of PpIX, indicating greater sensitivity of them to PDT. Cells from the Ca1 and Luc4 cell lines of the 5-ALA and PDT groups had less capacity to migrate compared to the other groups. An analysis of gene expression after PDT, showed a decrease in Bmi1 expression and increased Calgranulin expression in the Ca1 cell line, confirming the loss of the stem potential and induction of cell differentiation. Thus, the results shown here, indicate that a 5-ALA-PDT is capable of decreasing the cell viability and cell lines migration derived from OSCC, as well as reducing the percentage of cells with stem phenotype, inducing their differentiation.Dentre os vários tipos de câncer que afetam a cavidade bucal, o carcinoma epidermóide de boca (CEB) é a neoplasia maligna de maior prevalência. Apesar dos avanços no tratamento, os pacientes com CEB frequentemente apresentam metástases no momento do diagnóstico e recorrência, ambos associados com um prognóstico reservado. Estudos recentes têm demonstrado que a presença de células-tronco de câncer (CTC) são responsáveis pelo crescimento tumoral e resistência à terapia convencional. Uma alternativa para o tratamento do CEB é a Terapia Fotodinâmica (TFD). Porém, o efeito da TFD em CTC no CEB é pouco conhecido. Desta maneira, este estudo teve como objetivo avaliar in vitro o efeito da TFD mediada pelo fotossensibilizador ácido 5-aminulevulínico (5-ALA) na viabilidade celular, manutenção do fenótipo tronco, migração celular e expressão dos genes Bmi1, Clagranulina, IVL e E-caderina em duas linhagens celulares derivadas de CEB (Ca1 e Luc4). Todos os ensaios in vitro foram conduzidos com ambas linhagens celulares, as quais foram divididas nos seguintes grupos: Controle, 5-ALA, LED 6J/cm2 e TFD (5-ALA+6J/cm2). A viabilidade celular foi avaliada por meio dos ensaios de MTT e Vermelho neutro e o fenótipo tronco foi avaliado por meio de citometria de fluxo para análise da expressão de CD44 e ESA para identificação das populações CD44high/ESAhigh, CD44high/ESAlow e CD44low, bem como pelos ensaios de formação de colônias e esferas tumorais. Adicionalmente, foi avaliada a produção de PpIX em cada população de CTC. A migração celular foi avaliada por meio do ensaio de ferida e a expressão gênica foi avaliada por meio de RT-qPCR. Ambas as linhagens mostraram redução significante da viabilidade celular após a TFD. Foi observada redução na porcentagem de células CD44high/ESAhigh após a TFD quando comparada aos demais grupos, o que foi acompanhado por um aumento da porcentagem de células com fenótipo CD44low, indicando indução da diferenciação celular. Adicionalmente, houve redução da capacidade de formação de colônias e formação de esferas tumorais após a FTD. A população CD44high/ESAhigh de ambas as linhagens mostraram maior produção de PpIX, indicando maior sensibilidade das mesmas à TFD. As células das linhagens Ca1 e Luc4 dos grupos 5-ALA e TFD mostraram menor capacidade de migração em relação aos demais grupos. A análise da expressão gênico mostrou que após TFD, houve diminuição da expressão de Bmi1 e aumento da expressão de Calgranulina na linhagem Ca1, confirmando a perda do potencial tronco e indução da diferenciação celular. Desta maneira, os resultados aqui demonstrados indicam que a TFD-5-ALA é capaz de diminuir a viabilidade e migração celular de linhagens celulares derivadas de CEB, bem como reduz a porcentagem de células com fenótipo tronco, induzindo sua diferenciação.Submitted by Nadir Basilio (nadirsb@uninove.br) on 2021-11-04T17:38:34Z No. of bitstreams: 1 Marlene Aparecida Ferreira Pinto.pdf: 2951835 bytes, checksum: 0ab6be1786cd507a64fa7777783e01ad (MD5)Made available in DSpace on 2021-11-04T17:38:34Z (GMT). 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dc.title.por.fl_str_mv Efeito da terapia fotodinâmica no fenótipo tronco de linhagens celulares de carcinoma epidermóide de boca
dc.title.alternative.eng.fl_str_mv Effect of photodynamic therapy on the stem phenotipe of oral squamous cell lines carcinom
title Efeito da terapia fotodinâmica no fenótipo tronco de linhagens celulares de carcinoma epidermóide de boca
spellingShingle Efeito da terapia fotodinâmica no fenótipo tronco de linhagens celulares de carcinoma epidermóide de boca
Pinto, Marlene Aparecida Ferreira
carcinoma epidermóide de boca
terapia fotodinâmica
5-ALA
células-tronco de câncer
oral squamous cell carcinoma
photodynamic therapy
5-ALA
cancer stem cell
CIENCIAS DA SAUDE
title_short Efeito da terapia fotodinâmica no fenótipo tronco de linhagens celulares de carcinoma epidermóide de boca
title_full Efeito da terapia fotodinâmica no fenótipo tronco de linhagens celulares de carcinoma epidermóide de boca
title_fullStr Efeito da terapia fotodinâmica no fenótipo tronco de linhagens celulares de carcinoma epidermóide de boca
title_full_unstemmed Efeito da terapia fotodinâmica no fenótipo tronco de linhagens celulares de carcinoma epidermóide de boca
title_sort Efeito da terapia fotodinâmica no fenótipo tronco de linhagens celulares de carcinoma epidermóide de boca
author Pinto, Marlene Aparecida Ferreira
author_facet Pinto, Marlene Aparecida Ferreira
author_role author
dc.contributor.advisor1.fl_str_mv Rodrigues, Maria Fernanda Setúbal Destro
dc.contributor.advisor1Lattes.fl_str_mv http://lattes.cnpq.br/7153553682954852
dc.contributor.advisor-co1.fl_str_mv Pavanni, Christiane
dc.contributor.advisor-co1Lattes.fl_str_mv http://lattes.cnpq.br/2148910222765744
dc.contributor.referee1.fl_str_mv Rodrigues, Maria Fernanda Setúbal Destro
dc.contributor.referee1Lattes.fl_str_mv http://lattes.cnpq.br/7153553682954852
dc.contributor.referee2.fl_str_mv Pavani, Christiane
dc.contributor.referee2Lattes.fl_str_mv http://lattes.cnpq.br/2148910222765744
dc.contributor.referee3.fl_str_mv Dellê, Humberto
dc.contributor.referee3Lattes.fl_str_mv http://lattes.cnpq.br/7435662740477057
dc.contributor.referee4.fl_str_mv Franco, Adriana Lino dos Santos
dc.contributor.referee4Lattes.fl_str_mv http://lattes.cnpq.br/7295388304888384
dc.contributor.referee5.fl_str_mv Nunes, Fabio Daumas
dc.contributor.referee5Lattes.fl_str_mv http://lattes.cnpq.br/4909755821591847
dc.contributor.authorLattes.fl_str_mv http://lattes.cnpq.br/9592750937209234
dc.contributor.author.fl_str_mv Pinto, Marlene Aparecida Ferreira
contributor_str_mv Rodrigues, Maria Fernanda Setúbal Destro
Pavanni, Christiane
Rodrigues, Maria Fernanda Setúbal Destro
Pavani, Christiane
Dellê, Humberto
Franco, Adriana Lino dos Santos
Nunes, Fabio Daumas
dc.subject.por.fl_str_mv carcinoma epidermóide de boca
terapia fotodinâmica
5-ALA
células-tronco de câncer
topic carcinoma epidermóide de boca
terapia fotodinâmica
5-ALA
células-tronco de câncer
oral squamous cell carcinoma
photodynamic therapy
5-ALA
cancer stem cell
CIENCIAS DA SAUDE
dc.subject.eng.fl_str_mv oral squamous cell carcinoma
photodynamic therapy
5-ALA
cancer stem cell
dc.subject.cnpq.fl_str_mv CIENCIAS DA SAUDE
description Among the various types of cancer that affect the oral cavity, squamous cell carcinoma of the mouth (OSCC) is the most prevalent malignant neoplasm. Despite advances in treatment, patients with OSCC often present metastasis at diagnosis and recurrence, both associated with a poor prognosis. Recent studies have shown that the presence of cancer stem cells (CTC) are responsible for tumor growth and resistance to conventional therapy. An alternative for the treatment of CEB is Photodynamic Therapy (PDT). However, the effect of PDT on CTC in the CEB is unknown. Thus, this study aimed to evaluate in vitro the effect of PDT mediated by the photosensitizer 5-aminoulevulinic acid (5-ALA) on cell viability, maintenance of the stem cells phenotype, cell migration and expression of the genes Bmi1, Clagranulin, IVL and E- cadherin in two cell lines deriveted of OSCC (Ca1 and Luc4). All in vitro assays were conducted with both cell lines, which were divided into the following groups: Control, 5-ALA, LED 6J/cm² and PDT (5-ALA+6J/cm2). Cell viability was assessed by MTT and Neutral Red assays and the stem phenotype was assessed by flow cytometry to analyze the expression of CD44 and ESA to identify the CD44high/ESAhigh, CD44high/ESAlow and CD44low populations, as well as by colony and tumor sphere formation assays. Additionally, the PpIX production in each CTC population was evaluated. Cell migration was analyzed by wound healing assay and gene expression was analyzed by RT-qPCR. Both cell lines showed a significant reduction in cell viability after PDT. A percentage reduction of CD44high/ESAhigh cells was observed after PDT when compared to the other groups, which was followed by an increase in the percentage of cells with the CD44low phenotype, indicating cell differentiation induction. Additionally, reduced colony formation capacity and tumoral sphere formation after PDT. A CD44high / ESAhigh population of both cell lines had greater production of PpIX, indicating greater sensitivity of them to PDT. Cells from the Ca1 and Luc4 cell lines of the 5-ALA and PDT groups had less capacity to migrate compared to the other groups. An analysis of gene expression after PDT, showed a decrease in Bmi1 expression and increased Calgranulin expression in the Ca1 cell line, confirming the loss of the stem potential and induction of cell differentiation. Thus, the results shown here, indicate that a 5-ALA-PDT is capable of decreasing the cell viability and cell lines migration derived from OSCC, as well as reducing the percentage of cells with stem phenotype, inducing their differentiation.
publishDate 2021
dc.date.accessioned.fl_str_mv 2021-11-04T17:38:34Z
dc.date.issued.fl_str_mv 2021-08-04
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dc.identifier.citation.fl_str_mv Pinto, Marlene Aparecida Ferreira. Efeito da terapia fotodinâmica no fenótipo tronco de linhagens celulares de carcinoma epidermóide de boca. 2021. 77 f. Tese( Programa de Pós-Graduação em Biofotônica Aplicada às Ciências da Saúde) - Universidade Nove de Julho, São Paulo.
dc.identifier.uri.fl_str_mv http://bibliotecatede.uninove.br/handle/tede/2664
identifier_str_mv Pinto, Marlene Aparecida Ferreira. Efeito da terapia fotodinâmica no fenótipo tronco de linhagens celulares de carcinoma epidermóide de boca. 2021. 77 f. Tese( Programa de Pós-Graduação em Biofotônica Aplicada às Ciências da Saúde) - Universidade Nove de Julho, São Paulo.
url http://bibliotecatede.uninove.br/handle/tede/2664
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language por
dc.relation.cnpq.fl_str_mv 8765449414823306929
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dc.publisher.none.fl_str_mv Universidade Nove de Julho
dc.publisher.program.fl_str_mv Programa de Pós-Graduação em Biofotônica Aplicada às Ciências da Saúde
dc.publisher.initials.fl_str_mv UNINOVE
dc.publisher.country.fl_str_mv Brasil
dc.publisher.department.fl_str_mv Saúde
publisher.none.fl_str_mv Universidade Nove de Julho
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repository.name.fl_str_mv Biblioteca Digital de Teses e Dissertações da Uninove - Universidade Nove de Julho (UNINOVE)
repository.mail.fl_str_mv bibliotecatede@uninove.br||bibliotecatede@uninove.br
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