Formação de invadopódios na migração de células neoplásicas malignas expostas à fibronectina
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Data
2018-02-28
Autores
Cucielo, Maira Smaniotto [UNESP]
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Universidade Estadual Paulista (Unesp)
Resumo
Durante processos patológicos como o câncer, a atividade migratória de células neoplásicas aumenta, sendo um passo crítico para a disseminação dessas células a órgãos distantes. O processo migratório dessas células depende da formação de protrusões citoplasmáticas chamadas invadopódios, através da nucleação e polimerização de actina e degradação da matriz extracelular (MEC). A MEC é uma rede complexa de proteínas que fornece sustentação e moléculas sinalizadoras responsáveis pela homeostase dos tecidos. Dentre essas proteínas, a fibronectina (FN) é uma glicoproteína multiadesiva fundamental no comportamento celular normal e apresenta-se aumentada no tecido tumoral. Esse trabalho teve como objetivo avaliar a influência da FN, isolada ou em associação com componentes da membrana basal, no processo migratório de células tumorais prostáticas, com ênfase no mecanismo de formação dos invadopódios. Para isso, as linhagens celulares LNCaP e PC-3 foram divididas nos seguintes grupos experimentais: Controle, FN solúvel (10µg/mL), FN coating (10µg/mL), FN+Geltrex e Geltrex com exposição aos componentes por 4 dias. Nas células LNCaP, foi possível observar alterações morfológicas dos grupos expostos a FN (solúvel, coating e em associação com Geltrex) com diminuição na área celular em comparação ao grupo controle. A proliferação celular aumentou nos grupos com recobrimento da superfície (FN coating, FN+Geltrex e Geltrex). Também foi possível observar a diminuição na taxa de motilidade dessas células em todos os grupos expostos. A expressão das proteínas chaves para formação dos invadopódios (Cortactina, Dinamina-2 e Actina-F) também mostraram-se diminuídas na presença dos componentes da MEC e ainda, as proteínas pFAK e ERK, mostraram-se aumentadas. Já na linhagem celular PC-3, podemos observar redução na taxa de proliferação celular nos grupos com recobrimento da superfície (FN coating, FN+Geltrex e Geltrex) e aumento na taxa de motilidade nos grupos com recobrimento de Geltrex (FN+Geltrex e Geltrex). A Cortactina mostrou menor expressão no grupos FN+Geltrex quando comparada ao controle, e os demais grupos não apresentaram diferenças. Para a proteína Dinamina-2, observamos aumento na expressão no grupo FN solúvel quando comparada ao controle, e diminuição nos grupos FN coating e FN+Geltrex. Nas células PC-3, podemos observar a inconsistência nos dados obtidos, já que essa linhagem possui características avançadas na biologia dos tumores, como independência androgênica e a fatores de crescimento. Na linhagem celular LNCaP, foi possível observar maior suscetibilidade das respostas celulares ao microambiente oferecido, sendo assim, a FN e os componentes da MB, inibiram a formação dos invadopódios e a capacidade móvel dessas linhagem. Fica claro que a FN tem papel supressor na motilidade celular para essas células tumorais, porém o processo de formação dos invadopódios é complexo e dependente de várias vias de sinalização. Os nossos resultados são importantes, em sua maioria inéditos, pois revelam a importância e a influência da FN na inibição do comportamento migratório e formação das projeções citoplasmáticas de células tumorais e encorajam estudos adicionais sobre essa relevante temática de migração celular no câncer.
During pathological processes like cancer, the migratory activity of neoplasic cells increases, being a critical step for the dissemination of these cells to distant organs. The migratory process depends on the formation of cytoplasmic protrusions called invadopodia, through actin polymerization and extracellular matrix degradation (ECM). ECM is a complex protein network that provides support and molecules responsible for tissue homeostasis. Among these proteins, fibronectin (FN) is an essential adhesive glycoprotein in normal conditions however, is unregulated in tumor tissue. The objective of this study was to evaluate the influence of FN, alone or in association with basement membrane (BM) components, within migratory process of prostatic tumor cells, with emphasis on the mechanism of invadopodia. For this, the LNCaP and PC-3 cell lines were divided into the following experimental groups: Control, FN soluble (10μg / mL), FN coating (10μg / mL), FN + Geltrex and Geltrex. Exposure time was established into 4 days. We could observe morphological alterations of the groups exposed to FN in LNCaP cells, with a decrease in the cellular area in comparison to the control group. Cell proliferation was increased in the groups with surface coating (FN coating, FN + Geltrex and Geltrex). It was also possible to observe a decrease in the motility rate of these cells in all the exposed groups. Key proteins expression important to invadopodia formation (Cortactin, Dynamin-2 and Actin-F) was also shown to be decreased in the presence of the ECM components, and the pFAK and ERK proteins were also increased. PC-3 cell line, was observed a reduction in cell proliferation rate in the groups with surface coating (FN coating, FN + Geltrex and Geltrex). Besides, an increase in the motility rate in the Geltrex coated groups (FN + Geltrex and Geltrex). Cortactin showed lower expression in the FN + Geltrex groups when compared to control, and the other groups did not present significant differences. FN soluble group showed an increase in the expression of Dynamin-2 when compared to the control, and a decrease in FN coating and FN + Geltrex groups. We note an inconsistency in data to PC-3 cells, since this lineage has advanced characteristics in the tumor biology, as androgen and growth factors independence. It was possible to observe a higher susceptibility of the cellular responses to LNCaP microenvironment, thus, the FN and BM components inhibited invadopodia formation and mobile capacity of this lineage. It is clear that FN plays a suppressive role in cell motility for these type of tumor, but the process of invadopodia formation is complex and depends on several signaling pathways. Our results showed important data, as they reveal the importance and influence of FN in inhibiting migratory behavior and formation of cytoplasmic projections of tumor cells and encourage further studies on this relevant issue of cell migration concerning cancer.
During pathological processes like cancer, the migratory activity of neoplasic cells increases, being a critical step for the dissemination of these cells to distant organs. The migratory process depends on the formation of cytoplasmic protrusions called invadopodia, through actin polymerization and extracellular matrix degradation (ECM). ECM is a complex protein network that provides support and molecules responsible for tissue homeostasis. Among these proteins, fibronectin (FN) is an essential adhesive glycoprotein in normal conditions however, is unregulated in tumor tissue. The objective of this study was to evaluate the influence of FN, alone or in association with basement membrane (BM) components, within migratory process of prostatic tumor cells, with emphasis on the mechanism of invadopodia. For this, the LNCaP and PC-3 cell lines were divided into the following experimental groups: Control, FN soluble (10μg / mL), FN coating (10μg / mL), FN + Geltrex and Geltrex. Exposure time was established into 4 days. We could observe morphological alterations of the groups exposed to FN in LNCaP cells, with a decrease in the cellular area in comparison to the control group. Cell proliferation was increased in the groups with surface coating (FN coating, FN + Geltrex and Geltrex). It was also possible to observe a decrease in the motility rate of these cells in all the exposed groups. Key proteins expression important to invadopodia formation (Cortactin, Dynamin-2 and Actin-F) was also shown to be decreased in the presence of the ECM components, and the pFAK and ERK proteins were also increased. PC-3 cell line, was observed a reduction in cell proliferation rate in the groups with surface coating (FN coating, FN + Geltrex and Geltrex). Besides, an increase in the motility rate in the Geltrex coated groups (FN + Geltrex and Geltrex). Cortactin showed lower expression in the FN + Geltrex groups when compared to control, and the other groups did not present significant differences. FN soluble group showed an increase in the expression of Dynamin-2 when compared to the control, and a decrease in FN coating and FN + Geltrex groups. We note an inconsistency in data to PC-3 cells, since this lineage has advanced characteristics in the tumor biology, as androgen and growth factors independence. It was possible to observe a higher susceptibility of the cellular responses to LNCaP microenvironment, thus, the FN and BM components inhibited invadopodia formation and mobile capacity of this lineage. It is clear that FN plays a suppressive role in cell motility for these type of tumor, but the process of invadopodia formation is complex and depends on several signaling pathways. Our results showed important data, as they reveal the importance and influence of FN in inhibiting migratory behavior and formation of cytoplasmic projections of tumor cells and encourage further studies on this relevant issue of cell migration concerning cancer.
Descrição
Palavras-chave
Fibronectina, Invadopódio, Membrana basal, Migração, Câncer de próstata