ABSTRACT
Basal-like breast cancer (BBC) and glioblastoma multiforme (GBM) are aggressive cancers associated with poor prognosis. BBC and GBM have stem cell-like gene expression signatures, which are in part driven by forkhead box O (FOXO) transcription factors. To gain further insight into the impact of FOXO1 in BBC, we treated BT549 cells with AS1842856 and performed RNA sequencing. AS1842856 binds to unphosphorylated FOXO1 and inhibits its ability to directly bind to DNA. Gene Set Enrichment Analysis indicated that a set of WNT pathway target genes, including lymphoid enhancer-binding factor 1 (LEF1) and transcription factor 7 (TCF7), were robustly induced after AS1842856 treatment. These same genes were also induced in GBM cell lines U87MG, LN18, LN229, A172, and DBTRG upon AS1842856 treatment. By contrast, follow-up RNA interference (RNAi) targeting of FOXO1 led to reduced LEF1 and TCF7 gene expression in BT549 and U87MG cells. In agreement with RNAi experiments, CRISPR Cas9-mediated FOXO1 disruption reduced the expression of canonical WNT genes LEF1 and TCF7 in U87MG cells. The loss of TCF7 gene expression in FOXO1 disruption mutants was restored by exogenous expression of the DNA-binding-deficient FOXO1-H215R. Therefore, FOXO1 induces TCF7 in a DNA-binding-independent manner, similar to other published FOXO1-activated genes such as TCF4 and hes family bHLH transcription factor 1. Our work demonstrates that FOXO1 promotes canonical WNT gene expression in examined BBC and GBM cells, similar to results found in Drosophila melanogaster, T-cell development, and murine acute myeloid leukemia models.
Subject(s)
Drosophila melanogaster , Glioblastoma , Animals , Mice , Cell Differentiation , DNA , Glioblastoma/genetics , Stem Cells , HumansABSTRACT
Blood extraction by means of a vacuum system is a technique which has been employed in our field for some time, although this method does not receive all the popularity it should. This method offers something which the classical syringe can not: greater security. Proper blood extraction leads to a decrease in problems/errors which are susceptible to appear during the pre-analysis period. The use of a system which combine the characteristics of a vacuum with those of classical syringes, such as the Holdex de Vacuette (Bio Greiner, Madrid, España-H) can help to change this negative tendency. However, the special characteristics of this system (bendable to adapt to the site of a phlebotomy with the tube used to extract a blood sample) could cause an increase in the hemolysis in samples due to theoretically greater difficulty blood flow has in this circuit in comparison with other usual procedures in our filed (syringe and vacuum system. To test this assumption, 512 patients who had been asked to undergo a routine biochemical analysis were randomly chosen. These patients were divided into 3 groups, one for each extraction method used; 171 used Holdex de Vacuette(H), 168 used Vacutainer (V), and 173 used a syringe(J). The presence of hemolysis in these tubes was tested and the number of cases was compared to those detected among all the samples carried out during the study period. No statistically significant differences were discovered either among the 3 extraction systems used (H 2.1 or 16%, V 2.1 or 15%, J 4.2 or 23%) nor with the all samples analyzed (39, 2.02% total; 8, 1.56% study group, p < 0.001). These results let us conclude that the number of cases of hemolysis which occur when using Holdex is not greater than the number which occur when using other extraction systems and this method can be a good alternative to the conventional vacuum system.
Subject(s)
Blood Specimen Collection/instrumentation , Hemolysis , Adolescent , Adult , Aged , Aged, 80 and over , Equipment Design , Female , Humans , Male , Middle Aged , Needlestick Injuries/prevention & control , VacuumABSTRACT
La extracción sanguínea con sistemas de vacío (SV) es una técnica implantada en nuestro centro desde hace tiempo, aunque no goza de toda la popularidad que debería. Ofrece algo que la toma "clásica" con jeringuilla no puede: mayor seguridad. Una correcta extracción sanguínea favorece el descenso de problemas-errores del periodo preanalítico. Utilizar un sistema que combine las características del vacío con las de las clásicas jeringuillas, como el Holdex de Vacuette (Bio Greiner, Madrid, España-H-) puede ayudar a cambiar esta tendencia negativa. No obstante, las especiales características del sistema (acodado para adaptar el lugar de flebotomía con el tubo para muestra sanguínea) podría originar un aumento de la hemólisis debido a la teórica mayor dificultad del flujo de sangre en este circuito, en comparación con otros procedimientos usuales en nuestro medio (jeringuilla y SV). Para comprobar este supuesto se seleccionaron aleatoriamente 512 pacientes divididos en tres grupos atendiendo al sistema de extracción empleado (H: 171, Sistema BD Vacutainer® -V-: 168, jeringuilla -J-: 173). Se comprobó la presencia de hemólisis en estos tubos comparada con el total de muestras procesadas en el periodo de estudio. No se hallaron diferencias estadísticamente significativas ni entre los tres sistemas de extracción (H: 2,1, 16 por ciento; V: 2,1,15 por ciento; J: 4, 2,23 por ciento) ni con el total de muestras analizadas (39 ,2,02 por ciento total; 8, 1,56 por ciento grupo estudiado; p < 0,001). Estos resultados nos permiten concluir que la incidencia de hemólisis con el Holdex no es mayor que con otros sistemas de extracción y puede ser una buena alternativa al SV convencional (AU)
