Andrée Houbion

Publications (12)37.82 Total impact

• Source

  Available from: narilis.be

  Article: Hypersensitivity of mtDNA-depleted cells to staurosporine-induced apoptosis: roles of Bcl-2 downregulation and cathepsin B.

  Guillaume Rommelaere, Sébastien Michel, Ludovic Mercy, Antoine Fattaccioli, Catherine Demazy, Noelle Ninane, Andrée Houbion, Patricia Renard, Thierry Arnould
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  ABSTRACT: We show that mitochondrial DNA (mtDNA)-depleted 143B cells are hypersensitive to staurosporine-induced cell death as evidenced by a more pronounced DNA fragmentation, a stronger activation of caspase-3, an enhanced poly(ADP-ribose) polymerase-1 (PARP-1) cleavage, and a more dramatic cytosolic release of cytochrome c. We also show that B-cell CLL/lymphoma-2 (Bcl-2), B-cell lymphoma extra large (Bcl-X(L)), and myeloid cell leukemia-1 (Mcl-1) are constitutively less abundant in mtDNA-depleted cells, that the inhibition of Bcl-2 and Bcl-X(L) can sensitize the parental cell line to staurosporine-induced apoptosis, and that overexpression of Bcl-2 or Bcl-X(L) can prevent the activation of caspase-3 in ρ(0)143B cells treated with staurosporine. Moreover, the inactivation of cathepsin B with CA074-Me significantly reduced cytochrome c release, caspase-3 activation, PARP-1 cleavage, and DNA fragmentation in mtDNA-depleted cells, whereas the pan-caspase inhibitor failed to completely prevent PARP-1 cleavage and DNA fragmentation in these cells, suggesting that caspase-independent mechanisms are responsible for cell death even if caspases are activated. Finally, we show that cathepsin B is released in the cytosol of ρ(0) cells in response to staurosporine, suggesting that the absence of mitochondrial activity leads to a facilitated permeabilization of lysosomal membranes in response to staurosporine.
  AJP Cell Physiology 11/2010; 300(5):C1090-106. · 3.54 Impact Factor
• Article: Mild mitochondrial uncoupling induces 3T3-L1 adipocyte de-differentiation by a PPARgamma-independent mechanism, whereas TNFalpha-induced de-differentiation is PPARgamma dependent.

  Silvia Tejerina, Aurélia De Pauw, Sébastien Vankoningsloo, Andrée Houbion, Patricia Renard, Françoise De Longueville, Martine Raes, Thierry Arnould
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  ABSTRACT: Impairment of mitochondrial activity affects lipid-metabolizing tissues and mild mitochondrial uncoupling has been proposed as a possible strategy to fight obesity and associated diseases. In this report, we characterized the 3T3-L1-adipocyte ;de-differentiation' induced by carbonyl cyanide (p-trifluoromethoxy)-phenylhydrazone (FCCP), a mitochondrial uncoupler. We found a decrease in triglyceride (TG) content in adipocytes incubated with this molecule. We next analyzed the expression of genes encoding adipogenic markers and effectors and compared the differentially expressed genes in adipocytes treated with FCCP or TNFalpha (a cytokine known to induce adipocyte de-differentiation). Furthermore, a significant decrease in the transcriptional activity of PPARgamma and C/EBPalpha transcription factors was found in adipocytes with impaired mitochondrial activity. However, although these modifications were also found in TNFalpha-treated adipocytes, rosiglitazone and 9-cis retinoic acid (PPARgamma and RXR ligands) were unable to prevent triglyceride loss in FCCP-treated cells. Metabolic assays also revealed that TG reduction could be mediated by a downregulation of lipid synthesis rather than an upregulation of fatty acid oxidation. Finally, lipolysis stimulated by the uncoupler also seems to contribute to the TG reduction, a process associated with perilipin A downregulation. These results highlight some new mechanisms that might potentially be involved in adipocyte de-differentiation initiated by a mitochondrial uncoupling.
  Journal of Cell Science 01/2009; 122(Pt 1):145-55. · 6.11 Impact Factor
• Article: Gene expression silencing with 'specific' small interfering RNA goes beyond specificity - a study of key parameters to take into account in the onset of small interfering RNA off-target effects.

  Sébastien Vankoningsloo, Françoise de Longueville, Stéphanie Evrard, Pierre Rahier, Andrée Houbion, Antoine Fattaccioli, Mélanie Gastellier, José Remacle, Martine Raes, Patricia Renard, Thierry Arnould
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  ABSTRACT: RNA-mediated gene silencing (RNA interference) is a powerful way to knock down gene expression and has revolutionized the fields of cellular and molecular biology. Indeed, the transfection of cultured cells with small interfering RNAs (siRNAs) is currently considered to be the best and easiest approach to loss-of-function experiments. However, several recent studies underscore the off-target and potential cytotoxic effects of siRNAs, which can lead to the silencing of unintended mRNAs. In this study, we used a low-density microarray to assess gene expression modifications in response to five different siRNAs in various cell types and transfection conditions. We found major differences in off-target signature according to: (a) siRNA sequence; (b) cell type; (c) duration of transfection; and (d) post-transfection time before analysis. These results contribute to a better understanding of important parameters that could impact on siRNA side effects in knockdown experiments.
  FEBS Journal 07/2008; 275(11):2738-53. · 3.79 Impact Factor
• Article: CREB activation induced by mitochondrial dysfunction triggers triglyceride accumulation in 3T3-L1 preadipocytes.

  Sébastien Vankoningsloo, Aurélia De Pauw, Andrée Houbion, Silvia Tejerina, Catherine Demazy, Françoise de Longueville, Vincent Bertholet, Patricia Renard, José Remacle, Paul Holvoet, Martine Raes, Thierry Arnould
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  ABSTRACT: Several mitochondrial pathologies are characterized by lipid redistribution and microvesicular cell phenotypes resulting from triglyceride accumulation in lipid-metabolizing tissues. However, the molecular mechanisms underlying abnormal fat distribution induced by mitochondrial dysfunction remain poorly understood. In this study, we show that inhibition of respiratory complex III by antimycin A as well as inhibition of mitochondrial protein synthesis trigger the accumulation of triglyceride vesicles in 3T3-L1 fibroblasts. We also show that treatment with antimycin A triggers CREB activation in these cells. To better delineate how mitochondrial dysfunction induces triglyceride accumulation in preadipocytes, we developed a low-density DNA microarray containing 89 probes, which allows gene expression analysis for major effectors and/or markers of adipogenesis. We thus determined gene expression profiles in 3T3-L1 cells incubated with antimycin A and compared the patterns obtained with differentially expressed genes during the course of in vitro adipogenesis induced by a standard pro-adipogenic cocktail. After an 8-day treatment, a set of 39 genes was found to be differentially expressed in cells treated with antimycin A, among them CCAAT/enhancer-binding protein alpha (C/EBPalpha), C/EBP homologous protein-10 (CHOP-10), mitochondrial glycerol-3-phosphate dehydrogenase (GPDmit), and stearoyl-CoA desaturase 1 (SCD1). We also demonstrate that overexpression of two dominant negative mutants of the cAMP-response element-binding protein CREB (K-CREB and M1-CREB) and siRNA transfection, which disrupt the factor activity and expression, respectively, inhibit antimycin-A-induced triglyceride accumulation. Furthermore, CREB knockdown with siRNA also downregulates the expression of several genes that contain cAMP-response element (CRE) sites in their promoter, among them one that is potentially involved in synthesis of triglycerides such as SCD1. These results highlight a new role for CREB in the control of triglyceride metabolism during the adaptative response of preadipocytes to mitochondrial dysfunction.
  Journal of Cell Science 05/2006; 119(Pt 7):1266-82. · 6.11 Impact Factor
• Article: Growth and cytotoxic activity by Mycobacterium ulcerans in protein‐free media

  Armand Mve-Obiang, José Remacle, Juan Carlos Palomino, Andrée Houbion, Françoise Portaels
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  ABSTRACT: The pathogenic slow-growing Mycobacterium ulcerans has, until now, been cultured in liquid media containing albumin. Here, we report the first description of use of Sauton medium and modified Reid medium, two protein-free media, in which M. ulcerans was able to grow and produce its toxin, a major virulence factor of this environmental organism which causes a skin disease commonly called Buruli ulcer. These results suggest that Sauton and modified Reid may be useful for certain fields of M. ulcerans research requiring protein-free growth conditions.
  FEMS Microbiology Letters 01/2006; 181(1):153 - 157. · 2.04 Impact Factor
• Article: Mitochondrial biogenesis in mtDNA-depleted cells involves a Ca2+-dependent pathway and a reduced mitochondrial protein import.

  Ludovic Mercy, Aurélia de Pauw, Laetitia Payen, Silvia Tejerina, Andrée Houbion, Catherine Demazy, Martine Raes, Patricia Renard, Thierry Arnould
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  ABSTRACT: Alterations in mitochondrial activity resulting from defects in mitochondrial DNA (mtDNA) can modulate the biogenesis of mitochondria by mechanisms that are still poorly understood. In order to study mitochondrial biogenesis in cells with impaired mitochondrial activity, we used rho-L929 and rho(0)143 B cells (partially and totally depleted of mtDNA, respectively), that maintain and even up-regulate mitochondrial population, to characterize the activity of major transcriptional regulators (Sp1, YY1, MEF2, PPARgamma, NRF-1, NRF-2, CREB and PGC-1alpha) known to control the expression of numerous nuclear genes encoding mitochondrial proteins. Among these regulators, cyclic AMP-responsive element binding protein (CREB) activity was the only one to be increased in mtDNA-depleted cells. CREB activation mediated by a calcium-dependent pathway in these cells also regulates the expression of cytochrome c and the abundance of mitochondrial population as both are decreased in mtDNA-depleted cells that over-express CREB dominant negative mutants. Mitochondrial biogenesis in mtDNA-depleted cells is also dependent on intracellular calcium as its chelation reduces mitochondrial mass. Despite a slight increase in mitochondrial mass in mtDNA-depleted cells, the mitochondrial protein import activity was reduced as shown by a decrease in the import of radiolabeled matrix-targeted recombinant proteins into isolated mitochondria and by the reduced mitochondrial localization of ectopically expressed HA-apoaequorin targeted to the mitochondria. Decrease in ATP content, in mitochondrial membrane potential as well as reduction in mitochondrial Tim44 abundance could explain the lower mitochondrial protein import in mtDNA-depleted cells. Taken together, these results suggest that mitochondrial biogenesis is stimulated in mtDNA-depleted cells and involves a calcium-CREB signalling pathway but is associated with a reduced mitochondrial import for matrix proteins.
  FEBS Journal 11/2005; 272(19):5031-55. · 3.79 Impact Factor
• Article: Mitochondrial dysfunction induces triglyceride accumulation in 3T3-L1 cells: role of fatty acid beta-oxidation and glucose.

  Sébastien Vankoningsloo, Marie Piens, Christophe Lecocq, Audrey Gilson, Aurélia De Pauw, Patricia Renard, Catherine Demazy, Andrée Houbion, Martine Raes, Thierry Arnould
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  ABSTRACT: Mitochondrial cytopathy has been associated with modifications of lipid metabolism in various situations, such as the acquisition of an abnormal adipocyte phenotype observed in multiple symmetrical lipomatosis or triglyceride (TG) accumulation in muscles associated with the myoclonic epilepsy with ragged red fibers syndrome. However, the molecular signaling leading to fat metabolism dysregulation in cells with impaired mitochondrial activity is still poorly understood. Here, we found that preadipocytes incubated with inhibitors of mitochondrial respiration such as antimycin A (AA) accumulate TG vesicles but do not acquire specific markers of adipocytes. Although the uptake of TG precursors is not stimulated in 3T3-L1 cells with impaired mitochondrial activity, we found a strong stimulation of glucose uptake in AA-treated cells mediated by calcium and phosphatidylinositol 3-kinase/Akt1/glycogen synthase kinase 3beta, a pathway known to trigger the translocation of glucose transporter 4 to the plasma membrane in response to insulin. TG accumulation in AA-treated cells is mediated by a reduced peroxisome proliferator-activated receptor gamma activity that downregulates muscle carnitine palmitoyl transferase-1 expression and fatty acid beta-oxidation, and by a direct conversion of glucose into TGs accompanied by the activation of carbohydrate-responsive element binding protein, a lipogenic transcription factor. Taken together, these results could explain how mitochondrial impairment leads to the multivesicular phenotype found in some mitochondria-originating diseases associated with a dysfunction in fat metabolism.
  The Journal of Lipid Research 07/2005; 46(6):1133-49. · 5.56 Impact Factor
• Article: Aging as a multi-step process characterized by a lowering of entropy production leading the cell to a sequence of defined stages. II. Testing some predictions on aging human fibroblasts in culture

  Olivier Toussaint, Andrée Houbion, José Remacle
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  ABSTRACT: The concepts of irreversible thermodynamics have been used in order to develop a theory of aging considered as a multi-step process leading the cell through a sequence of defined stages characterized by a lower level of entropy production and finally to a critical level of errors involving cell death (Toussaint et al., 1991). One of the predictions of this model is that external stresses which can be considered as fluctuations would accelerate the evolution of the cell from one state to the other according to the intensity of the stress. Seven morphotypes have been observed in the serially cultivated human fibroblasts, cells passing progressively from one morphotype to the other. In this paper, we experimentally tested the effect of two different molecules, tert-butylhydroperoxide and ethanol, in order to determine their influence on the shift from one morphotype to the other. When applied for a single period of time on cultivated cells, both molecules effectively showed a modification in the pattern of the different morphotyes which was dependant on the stress intensity: a decreased proportion of the early morphotypes and an increased proportion of the late and post-mitotic morphotypes were observed within three days after the stresses. Similar results were obtained when successive stresses were performed at every subculture. The results also indicated that all stages are not equally stable with morphotypes III and IV being the most stable. The positive effect on the increased shift of these cells from one morphotype to the other by two different stresses firms one of the prediction of the ethermodynamic model which states that cellular aging can be considered as a multi-step process which can be speeded up by various external modifications.
  Mechanisms of Ageing and Development 09/1992; · 3.44 Impact Factor
• Article: Alteration of enzymes in ageing human fibroblasts in culture. III. Modification of superoxide dismutase as an environmental and reversible process

  Michel Somville, Anne Houben, Martine Raes, Andrée Houbion, Vincent Henin, José Remacle
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  ABSTRACT: The alteration of superoxide dismutase (SOD) defined as the change occurring in its thermostability and observed in ageing cells, concerned the cytoplasmic but not the mitochondrial enzymes. Altered SOD disappeared if it was incubated in a supernatant from the young cells whereas supernatants from the old cells induced the alteration. The alteration induced on purified SOD was found to be associated with the appearance of thermolabile tetramers. Cytoplasmic SOD tetramers were also observed in the supernatants of the old cells. The addition of NADPH into the incubation medium could reverse this alteration; also when cultivated in the presence of vincamine the alteration, normally present in the old cells, disappears. The alteration of SOD is therefore associated with the formation of tetramers; it is a reversible process influenced by the cytoplasmic composition of the old cells.
  Mechanisms of Ageing and Development 02/1985; · 3.44 Impact Factor
• Article: Sensitive assay for nicotinamide adenine dinucleotide phosphate and its reduced form based on the bioluminescence method

  José Ramacle, Andrée Houbion, Martine Raes
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  ABSTRACT: An assay of reduced nicotinamide adenine dinucleotide phosphate (NADPH) by bioluminescence was investigated and applied for NADP+. The NADP+ is first reduced by glucose-6-phosphate dehydrogenase and then assayed in a mixture containing a NADPH/flavin mononucleotide oxidoreductase which in turn activates luciferase. Many interferences were observed and the method was modified accordingly. NADP+ and NADPH can be assayed separately or simultaneously within the range 1–100 pmol, which is sufficiently sensitive to be applied to biological materials. Many details and precautions must be taken into consideration.
  Analytica Chimica Acta. 170:109-116.
• Article: Development of a sensitive multi-well colorimetric assay for active NF{{kappa}}B

  Patricia Renard, Isabelle Ernest, Andrée Houbion, Muriel Art, Hervé Le Calvez, Martine Raes, José Remacle
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  ABSTRACT: The transcription factor nuclear factor &kgr;B (NF&kgr;B) is a key factor in the immune response triggered by a wide variety of molecules such as inflammatory cytokines, or some bacterial and viral products. This transcription factor represents a new target for the development of anti-inflammatory molecules, but this type of research is currently hampered by the lack of a convenient and rapid screening assay for NF&kgr;B activation. Indeed, NF&kgr;B DNA-binding capacity is traditionally estimated by radioactive gel shift assay. Here we propose a new DNA-binding assay based on the use of multi-well plates coated with a cold oligonucleotide containing the consensus binding site for NF&kgr;B. The presence of the DNA-bound transcription factor is then detected by anti-NF&kgr;B antibodies and revealed by colorimetry. This assay is easy to use, non-radioactive, highly reproducible, specific for NF&kgr;B, more sensitive than regular radioactive gel shift and very convenient for high throughput screening.
• Article: Alteration of enzymes in ageing human fibroblasts in culture IV. Effect of glutathione on the alteration of glucose-6-phosphate dehydrogenase

  D.L. Mbemba Funfu, Andrée Houbion, José Remacle
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  ABSTRACT: Alteration and inactivation of glucose-6-phosphate dehydrogenase (G6PD) can be induced in human fibroblasts by incubation of a cell supernatant at 4°C and pH 7.4. When added in such conditions, glutathione (GSH) had a stabilizing effect on the enzyme. On the other hand, substances which are known to deplete the cells of their GSH content, dramatically increase the inactivation rate. When analysed by gel filtraton after 24 h of incubation at 4°C, the inactive G6PD appears as a dimeric protein when GSH is present, while as a monomer in the control experiment. Reactivation of the monomers was stimulated with GSH. The heat inactivation of the dimeric fraction first started with a sharp activity increase of 20%. This increase vanished when the enzyme was first reactivated before the thermolability experiment. We propose that what is called altered G6PD is the expression of a quick reactivation of an inactive, labile dimer. Finally, a schematic view of the G6PD alteration is proposed.
  Mechanisms of Ageing and Development.