ABSTRACT
BACKGROUND: Tumour-initiating cells (TICs) or cancer stem cells can exist as a small population in malignant tissues. The signalling pathways activated in TICs that contribute to tumourigenesis are not fully understood. METHODS: Several breast cancer cell lines were sorted with CD24 and CD44, known markers for enrichment of breast cancer TICs. Tumourigenesis was analysed using sorted cells and total RNA was subjected to gene expression profiling and gene set enrichment analysis (GSEA). RESULTS: We showed that several breast cancer cell lines have a small population of CD24(-/low)/CD44(+) cells in which TICs may be enriched, and confirmed the properties of TICs in a xenograft model. GSEA revealed that CD24(-/low)/CD44(+) cell populations are enriched for genes involved in transforming growth factor-beta, tumour necrosis factor, and interferon response pathways. Moreover, we found the presence of nuclear factor-kappaB (NF-kappaB) activity in CD24(-/low)/CD44(+) cells, which was previously unrecognised. In addition, NF-kappaB inhibitor dehydroxymethylepoxyquinomicin (DHMEQ) prevented tumourigenesis of CD24(-/low)/CD44(+) cells in vivo. CONCLUSION: Our findings suggest that signalling pathways identified using GSEA help to identify molecular targets and biomarkers for TIC-like cells.
Subject(s)
Breast Neoplasms/pathology , CD24 Antigen/analysis , Cell Separation/methods , Gene Expression Profiling , Hyaluronan Receptors/analysis , Neoplasm Proteins/analysis , Neoplastic Stem Cells/physiology , Animals , Benzamides/pharmacology , Benzamides/therapeutic use , Biomarkers , Breast Neoplasms/genetics , Cyclohexanones/pharmacology , Cyclohexanones/therapeutic use , Female , Genetic Vectors/pharmacology , Humans , Mice , Mice, Inbred NOD , Mice, SCID , NF-kappa B/antagonists & inhibitors , NF-kappa B/physiology , Neoplasm Proteins/antagonists & inhibitors , Neoplasm Proteins/physiology , Neoplastic Stem Cells/chemistry , Neoplastic Stem Cells/transplantation , Oligonucleotide Array Sequence Analysis , Signal Transduction/physiology , Transduction, Genetic , Xenograft Model Antitumor AssaysABSTRACT
Mammalian corticogenesis occurs through a complex process that includes neurogenesis, in which neural progenitor cells proliferate, differentiate, and migrate. It has been reported recently that neurogenesis occurs in the subventricular zone (SVZ), a region previously thought to be the primary site of gliogenesis. It has been recognized that in the SVZ, intermediate progenitor cells, derived from radial glial cells that are multipotent neural stem cells, produce only neurons. However, the molecular mechanisms underlying the regulation of neural stem cells and intermediate progenitor cells as well as their contribution to overall corticogenesis remain unknown. The docking protein FRS2alpha is a major mediator of signaling by means of FGFs and neurotrophins. FRS2alpha mediates many of its pleiotropic cellular responses by recruiting the adaptor protein Grb2 and the protein tyrosine phosphatase Shp2 upon ligand stimulation. Here, we report that targeted disruption of Shp2-binding sites in FRS2alpha leads to severe impairment in cerebral cortex development in mutant mice. The defect in corticogenesis appears to be due at least in part to abnormalities in intermediate progenitor cells. Genetic evidence is provided that FRS2alpha plays critical roles in the maintenance of intermediate progenitor cells and in neurogenesis in the cerebral cortex. Moreover, FGF2-responsive neurospheres, which are cell aggregates derived from neural stem/progenitor cells (NSPCs), from FRS2alpha mutant mice were smaller than those of WT mice. However, mutant NSPCs were able to self-renew, demonstrating that Shp2-binding sites on FRS2alpha play an important role in NSPC proliferation but are dispensable for NSPC self-renewing capacity after FGF2 stimulation.
Subject(s)
Cerebral Cortex/growth & development , Intracellular Signaling Peptides and Proteins/metabolism , Membrane Proteins/metabolism , Neurons/cytology , Protein Tyrosine Phosphatases/metabolism , Receptor, Fibroblast Growth Factor, Type 2/physiology , Stem Cells/cytology , Animals , Binding Sites , Cell Proliferation , Cells, Cultured , Cerebral Cortex/cytology , Embryo, Mammalian , Immunohistochemistry , Intracellular Signaling Peptides and Proteins/physiology , Membrane Proteins/physiology , Mice , Mice, Mutant Strains , Protein Tyrosine Phosphatase, Non-Receptor Type 11 , Protein Tyrosine Phosphatases/physiologyABSTRACT
The docking protein FRS2alpha is a major mediator of fibroblast growth factor (FGF) signaling. However, the physiological role of FRS2alpha in vivo remains unknown. In this report, we show that Frs2alpha-null mouse embryos have a defect in anterior-posterior (A-P) axis formation and are developmentally retarded, resulting in embryonic lethality by embryonic day 8. We demonstrate that FRS2alpha is essential for the maintenance of self-renewing trophoblast stem (TS) cells in response to FGF4 in the extraembryonic ectoderm (ExE) that gives rise to tissues of the placenta. By analyzing chimeric embryos, we found that FRS2alpha also plays a role in cell movement through the primitive streak during gastrulation. In addition, experiments are presented demonstrating that Bmp4 expression in TS cells is controlled by mitogen-activated protein kinase-dependent FGF4 stimulation. Moreover, both the expression of Bmp4 in ExE and activation of Smad1/5 in epiblasts are reduced in Frs2alpha-null embryos. These experiments underscore the critical role of FRS2alpha in mediating multiple processes during embryonic development and reveal a potential new link between FGF and Bmp4 signaling pathways in early embryogenesis.
Subject(s)
Embryo, Mammalian/drug effects , Fibroblast Growth Factors/pharmacology , Membrane Proteins/metabolism , Proto-Oncogene Proteins/pharmacology , Signal Transduction/drug effects , Animals , Body Patterning , Bone Morphogenetic Protein 4 , Bone Morphogenetic Proteins/deficiency , Bone Morphogenetic Proteins/genetics , Cell Movement , Cell Survival , Chimera/abnormalities , Chimera/embryology , Chimera/genetics , DNA-Binding Proteins/metabolism , Embryo, Mammalian/cytology , Embryo, Mammalian/embryology , Embryo, Mammalian/metabolism , Enzyme Activation , Extracellular Signal-Regulated MAP Kinases/metabolism , Fibroblast Growth Factor 4 , Gastrula/drug effects , Gastrula/pathology , Gene Deletion , Gene Expression Regulation, Developmental/drug effects , Membrane Proteins/deficiency , Membrane Proteins/genetics , Mice , Mice, Knockout , Nodal Protein , Promoter Regions, Genetic/genetics , Smad Proteins , Smad1 Protein , Trans-Activators/metabolism , Transforming Growth Factor beta/deficiency , Transforming Growth Factor beta/geneticsABSTRACT
In previous studies (Masuda and Nakayama, 1983), diethyldithiocarbamate (DTC) and carbon disulfide (CS2) have been found to be protective against acute nephrotoxicity induced by CHCl3 and 1,1-dichloroethylene in normal and CCl4-poisoned mice, and it has been suggested that the protective action of DTC and CS2 might be mediated through inhibition of bioactivation of these nephrotoxicants in the kidney. As an extension of these studies, similar experiments were undertaken with furan, bromobenzene and cephaloridine, other nephrotoxic agents that are also thought to require metabolic activation. DTC or CS2 prevented mice from suffering renal injury induced by furan and bromobenzene, as evidenced by suppression of elevations in plasma urea nitrogen concentration and kidney calcium content and of morphologic alterations. Cephaloridine nephrotoxicity, however, was not prevented. In CCl4-poisoned mice, furan nephrotoxicity was augmented, whereas bromobenzene and cephaloridine nephrotoxicity was suppressed. The augumented furan nephrotoxicity was also prevented by DTC or CS2. These observations suggest that furan, like CHCl3 and 1,1-dichloroethylene, may exert nephrotoxicity through active metabolites formed in the kidney. For bromobenzene and cephaloridine nephrotoxicity, a renal bioactivation mechanism is suspected. DTC and CS2 also protected against hepatotoxicity induced by furan and thiophene.
