Detección y aislamiento de células inmaduras neuronales a partir de líneas celulares de tumores del sistema nervioso / Detection and isolation of immature neural cells from tumoral cell lines of the nervous system

Introducción: Hoy se sugiere que la teoria de carcinogénesis establecida en las últimas décadas, según la cual el cáncer se produce por el acúmulo de mutaciones en proto-oncogenes y genes supresores de tumores podría cuestionarse, a favor de una teoría más completa que incluya la participación de las denominadas células stem tumorales. Estas células poseerían la capacidad de iniciar y mantener el tumor, además de su propia capacidad de diferenciación y autorrenovación. Objetivo: Determinar el fenotipo de marcadores de diferenciación y la posible existencia de células inmaduras (tal vez células stem) en las líneas celulares de tumores del sistema nervioso. Material y métodos: Se estudió el nivel de expressión de los genes CD133, nestina, Musashi-1, FAS, NCAM1 y GFAP en 27 líneas celulares de tumores del sistema nervioso mediante RT-PCR semicuantitativa y citometría. La línea de neuroblastoma IMR-32 fue sometida a separación celular mediante SdFFF. Resultados: Hemos podido separar diferentes subclones o células en diferentes estadios de diferenciación en la línea de neuroblastoma IMR-32. La correcta tipificación de estas líneas celulares podría ser relevante para establecer tratamientos quimioterápicos o de terapia génica, específicamente dirigidos contra los subclones celulares más inmaduros, que podrían corresponder (o no) a células stem tumorales (AU)

Introduction: Nowadays it is suggested that the theory of carcinogenesis established along the last decades, according to which cancer is produced by the accumulation of mutations in proto-oncogenes and tumor suppressor genes might be questioned in favor of a more complete theory that includes the participation of the so called tumor stem cells. These cells would represent those in charge of initiating and maintaining the tumor; with their own capacity of differentiation and autorenewal. Objective: To determine the phenotype of differentiation markers and the possible existence of immature cells (maybe stem cells) in cell lines of tumors of the nervous system. Material and methods: The level of expression of CD133, nestine, Musashi-1, FAS, NCAM1 and GFAP genes was studied in 27 cell lines of tumors of the nervous system by semiquantitative RT-PCR and cytometry. The neuroblastoma cell line IMR-32 was subjected to cell separation by SdFFF. Results:We could separate different subclones or cells in different stages of differentiation in the neuroblastoma cell line IMR-32. The correct description of these cells might be relevant to set up chemoterapeutic or gene therapy treatments specifically targeted against the most immature subclones, that might correspond (or not) to tumor stem cells (AU)

Study of the phenotypic relationship in the IMR-32 human neuroblastoma cell line by sedimentation field flow fractionation.

Neuroblastoma (NB) is the most common childhood solid tumor. Although spontaneous regression can occur in patients <1-year old, 70% of patients over the age of 1 year have a high-risk and difficult-to-treat NB. Cell type heterogeneity is observed either in the morphological appearance of NB tumors or in cell lines isolated from tumor specimens. NB consists of two principal neoplastic cell types: i) neuroblastic or N-type (undifferentiated cells); and ii) stromal or S-type (differentiated cells). As NB cells seem to have the capacity to differentiate spontaneously in vivo and in vitro, their heterogeneity could affect treatment outcome, in particular the response to apoptosis induced by chemotherapy. Therefore, it is important to understand the underlying process governing changes in differentiation in order to improve treatment response and NB patient outcome and the neoplastic population in IMR-32 represented a good model for such a study. Results showed that this cell line was extremely heterogeneous and highly variable in its stage of differentiation and we demonstrated that sedimentation field flow fractionation (SdFFF) permitted the isolation of 2 N-phenotypes and contributed to the understanding of the IMR-32 cell population dynamics. The first N-phenotype forms a pool of quiescent undifferentiated cells while the second one was able to proliferate (incorporation of BrdU) and also give rise to adherent S-type cells (PSA-N-CAM+ and N-CAM+). The results could also suggest a close interaction between these different cellular phenotypes to create the IMR-32 cell lineage.