The purpose of this research has been deciphering the Warburg paradox, the biochemical enigma unsolved since 1923. We solved it by demonstrating that its specific character, i.e.
View Article and Find Full Text PDFWe previously showed that cellular RedOx state governs the G-S transition of AH130 hepatoma, a tumor spontaneously reprogrammed to the embryonic stem cell stage. This transition is impaired when the mithocondrial electron transport system is blocked by specific inhibitors (antimycin A) or the respiratory chain is saturated by adding to the cells high concentrations of pyruvate. The antimycin A or pyruvate block is removed by the addition of adequate concentrations of folate (F).
View Article and Find Full Text PDFWe defined the stem cell profile of K562 line, demonstrating the expression of the Embryonic Transcription Factors Oct3/4, Sox2, Klf4 and Nanog. This profile was associated with a high vulnerability to the physiological oxidizable substrate pyruvate. remarkably, this substrate was shown to be innocuous, even at the highest doses, to normal differentiated cells.
View Article and Find Full Text PDFWe have previously shown that peculiar metabolic features of cell adaptation and survival in hypoxia imply growth restriction points that are typical of embryonic stem cells and disappear with differentiation. Here we provide evidence that such restrictions can be exploited as specific antiblastic targets by physiological factors such as pyruvate, tetrahydrofolate, and glutamine. These metabolites act as powerful cytotoxic agents on cancer stem cells (CSCs) when supplied at doses that perturb the biochemical network, sustaining the resumption of aerobic growth after the hypoxic dormant state.
View Article and Find Full Text PDFOne undisputed milestone of traditional oncology is neoplastic progression, which consists of a progressive selection of dedifferentiated cells driven by a chance sequence of genetic mutations. Recently it has been demonstrated that the overexpression of well-defined transcription factors reprograms somatic cells to the pluripotent stem status. The demonstration raises crucial questions as to whether and to what extent this reprogramming contributes to tumorigenesis, and whether the epigenetic changes involved in it are reversible.
View Article and Find Full Text PDFA novel tethered bilayer lipid micromembrane (tBLmicroM) was prepared and characterized. It consists of a mercury cap electrodeposited on a platinum microelectrode, about 20 microm in diameter. The micromembrane was prepared by tethering to the mercury cap a thiolipid monolayer and by then self-assembling a lipid monolayer on top of it.
View Article and Find Full Text PDFTumors are ecosystems which develop from stem cells endowed with unlimited self-renewal and genetic instability, under the effects of mutagenesis and natural selection imposed by environmental changes. While changes and variations made possible by genetic instability are practically unlimited, the microenvironment progressively reduces those possibilities in the struggle for life imposed by hypoxia and nutrient shortage typical of tumor environments. This entails the tendency to evolve a convergent phenotype resistant to microenvironmental restrictions (first of all hypoxia), which progressively dominates the clonal selection.
View Article and Find Full Text PDFThe HERG potassium channel was incorporated in a mercury-supported tethered bilayer lipid membrane (tBLM) obtained by anchoring a thiolipid monolayer to the mercury surface and by self-assembling a lipid monolayer on top of it from a lipid film spread on the surface of an electrolyte solution. HERG was then incorporated in this tBLM from its micellar solution in Triton X-100, thus avoiding the use of vesicles in the preparation of the tBLM and of proteoliposomes in channel incorporation. The HERG "inward" current following a repolarization step was obtained by subtracting the current recorded upon addition of the specific inhibitor WAY from that recorded prior to this addition.
View Article and Find Full Text PDFWe worked out an experimental protocol able to purge the stem cell compartment of the SH-SY5Y neuroblastoma clone. This protocol was based on the prolonged treatment of the wild-type cell population with either hypoxia or the antiblastic etoposide. Cell fate was monitored by immunocytochemical and electrophysiologic (patch-clamp) techniques.
View Article and Find Full Text PDFWe showed that resistance to severe hypoxia defines hierarchical levels within normal hematopoietic populations and that hypoxia modulates the balance between generation of progenitors and maintenance of hematopoietic stem cells (HSC) in favor of the latter. This study deals with the effects of hypoxia (0.1% oxygen) in vitro on Friend's murine erythroleukemia (MEL) cells, addressing the question of whether a clonal leukemia cell population comprise functionally different cell subsets characterized by different hypoxia resistance.
View Article and Find Full Text PDFThe macrophage colony-stimulating factor (M-CSF, CSF-1) regulates survival, proliferation and differentiation of mononuclear phagocytes, as well as macrophage motility and morphology. The latter features are usually regulated by ECM-mediated activation of integrins and subsequent tyrosine phosphorylation of cellular proteins, including focal adhesion kinase (FAK). FAK is phosphorylated by downstream receptor tyrosine kinases as well.
View Article and Find Full Text PDFWe explored the stem cell compartment of the SH-SY5Y neuroblastoma (NB) clone and its development by a novel approach, integrating clonal and immunocytochemical investigations with patch-clamp measurements of ion currents simultaneously expressed on single cells. The currents selected were the triad IHERG, IKDR, INa, normally expressed at varying mutual ratios during development of neural crest stem cells, from which NB derives upon neoplastic transformation. These ratios could be used as electrophysiological clusters of differentiation (ECDs), identifying otherwise indistinguishable stages in maturation.
View Article and Find Full Text PDFAdhesive receptors of the integrin family are primarily involved in cell-extracellular matrix adhesion. Additionally, integrins trigger multiple signaling pathways that are involved in cell migration, proliferation, survival, and differentiation. We previously demonstrated that the activation of integrins containing the beta(1) subunit leads to a selective increase in potassium currents carried by the human ether-a-go-go-related gene (hERG) channels in neuroblastoma and leukemia cells; this current activation modulates adhesion-dependent differentiation in these cells.
View Article and Find Full Text PDFThe acquisition of the capacity to invade surrounding tissues confers a more malignant phenotype to tumor cells and is necessary for the establishment of metastases. The understanding of the molecular mechanisms underlying cell invasion in human solid tumors such as colorectal cancers could provide not only more sensitive prognostic analyses but also novel molecular targets for cancer therapy. We report in this article that K(+) ion channels belonging to the HERG family are important determinants for the acquisition of an invasive phenotype in colorectal cancers.
View Article and Find Full Text PDFDeciphering the expression pattern of K+ channel encoding genes during development can help in the understanding of the establishment of cellular excitability and unravel the molecular mechanisms of neuromuscular diseases. We focused our attention on genes belonging to the erg family, which is deeply involved in the control of neuromuscular excitability in Drosophila flies and possibly other organisms. Both in situ hybridisation and RNase Protection Assay experiments were used to study the expression pattern of mouse (m)erg1, m-erg2 and m-erg3 genes during mouse embryo development, to allow the pattern to be compared with their expression in the adult.
View Article and Find Full Text PDFBiochem Biophys Res Commun
February 2003
From the adrenergic SH-SY5Y human neuroblastoma clone, we isolated a subclone (21S) endowed with a glial-oriented phenotype. At difference from the parental clone, 21S cells responded to depolarizing stimuli with overshooting action potentials, whose repolarization phase was composed of an initial rapid episode, followed by a long-lasting plateau and a slow return to the resting potential (V(REST)). The action potential depolarization phase was sustained by a TTX-sensitive Na(+) current, while the first repolarizing episode was produced by the scanty delayed rectifier potassium current (I(KDR)) expressed in 21S cells.
View Article and Find Full Text PDFThe role of K(+) channel activity during cell cycle progression has become a research topic of considerable interest. Blocking of K(+) channels inhibits the proliferation of many cell types, although the mechanism of this inhibition is unclear. There is speculation that K(+) channels differentially regulate the electrical potential of the plasma membrane (V(m)) during proliferation.
View Article and Find Full Text PDFWe had previously shown that murine macrophages expressing v-Fes, the oncogenically activated counterpart of the c-Fes cytoplasmic tyrosine kinase, proliferate independently of Macrophage Colony-Stimulating Factor (MCSF) and that the Extracellular signal-Regulated Kinase (ERK) pathway mediates the mitogenic effect of v-Fes. In this study, the response of c-fes- and v-fes-overexpressing cells to MCSF was investigated. A critical modulation of the activation of Mitogen-activated ERK Kinase (MEK) and ERK based on the MCSF dose was characterized.
View Article and Find Full Text PDFNative rat lactotrophs express thyrotrophin-releasing hormone-dependent K+ currents consisting of fast and slow deactivating components that are both sensitive to the class III anti-arrhythmic drugs that block the eag-related gene (ERG) K+ current (I(ERG)). Here we describe in MMQ prolactin-releasing pituitary cells the isolation of the slowly deactivating long-lasting component (I(ERGS)), which, unlike the fast component (I(ERGF)), is insensitive to verapamil 2 microm but sensitive to a novel scorpion toxin (ErgTx-2) that hardly affects I(ERGF). The time constants of I(ERGS) activation, deactivation, and recovery from inactivation are more than one order of magnitude greater than in I(ERGF), and the voltage-dependent inactivation is left-shifted by approximately 25 mV.
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