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8 results on '"MESH : Muscle, Smooth, Vascular"'

1. Selective Involvement of Serum Response Factor in Pressure-Induced Myogenic Tone in Resistance Arteries

Author

Retailleau, Kevin, Toutain, Bertrand, Galmiche, Guillaume, Fassot, Celine, Sharif-Naeini, Reza, Kauffenstein, Gilles, Mericskay, Mathias, Duprat, Fabrice, Li, Linda, Mérot, Jean, Lardeux, Aurelie, Pizard, Anne, Baudrie, Véronique, Jeunemaitre, Xavier, Feil, Robert, Göthert, Joachim, Lacolley, Patrick, Henrion, Daniel, Li, Zhenlin, Loufrani, Laurent, Grimaud, L., Biologie Neurovasculaire et Mitochondriale Intégrée (BNMI), Université d'Angers (UA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Physiologie de la Nutrition et du Comportement Alimentaire (PNCA), AgroParisTech-Institut National de la Recherche Agronomique (INRA), Université Pierre et Marie Curie - Paris 6 (UPMC), Institut de pharmacologie moléculaire et cellulaire (IPMC), Université Nice Sophia Antipolis (... - 2019) (UNS), Université Côte d'Azur (UCA)-Université Côte d'Azur (UCA)-Centre National de la Recherche Scientifique (CNRS), UMR915, Nutrition et athérome, Institut National de la Santé et de la Recherche Médicale (INSERM), Défaillance Cardiovasculaire Aiguë et Chronique (DCAC), Centre Hospitalier Régional Universitaire de Nancy (CHRU Nancy)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lorraine (UL), Paris-Centre de Recherche Cardiovasculaire (PARCC - UMR-S U970), Hôpital Européen Georges Pompidou [APHP] (HEGP), Assistance publique - Hôpitaux de Paris (AP-HP) (APHP)-Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO)-Assistance publique - Hôpitaux de Paris (AP-HP) (APHP)-Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM), Service de Génétique [AP-HP Hôpital Européen Georges Pompidou, Paris], Assistance publique - Hôpitaux de Paris (AP-HP) (APHP)-Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO)-Assistance publique - Hôpitaux de Paris (AP-HP) (APHP)-Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO), Université Paris Descartes - Faculté de Médecine (UPD5 Médecine), Université Paris Descartes - Paris 5 (UPD5), Institut de Génétique Moléculaire de Montpellier (IGMM), Centre National de la Recherche Scientifique (CNRS)-Université de Montpellier (UM), Eberhard Karls Universität Tübingen, Universitätsklinikum Essen [Universität Duisburg-Essen] (Uniklinik Essen), Centre Hospitalier Universitaire d'Angers (CHU Angers), PRES Université Nantes Angers Le Mans (UNAM), Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université d'Angers (UA), Institut National de la Recherche Agronomique (INRA)-AgroParisTech, Centre National de la Recherche Scientifique (CNRS)-Université Nice Sophia Antipolis (... - 2019) (UNS), COMUE Université Côte d'Azur (2015-2019) (COMUE UCA)-COMUE Université Côte d'Azur (2015-2019) (COMUE UCA)-Université Côte d'Azur (UCA), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM), Service de Génétique [CHU HEGP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO), Eberhard Karls Universität Tübingen = Eberhard Karls University of Tuebingen, Biologie Neurovasculaire et Mitochondriale Intégrée, Université d'Angers ( UA ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Centre National de la Recherche Scientifique ( CNRS ), Physiologie de la Nutrition et du Comportement Alimentaire ( PNCA ), Institut National de la Recherche Agronomique ( INRA ) -AgroParisTech, Université Pierre et Marie Curie - Paris 6 ( UPMC ), Institut de pharmacologie moléculaire et cellulaire ( IPMC ), Université Nice Sophia Antipolis ( UNS ), Université Côte d'Azur ( UCA ) -Université Côte d'Azur ( UCA ) -Centre National de la Recherche Scientifique ( CNRS ), Institut National de la Santé et de la Recherche Médicale ( INSERM ), Défaillance Cardiovasculaire Aiguë et Chronique ( DCAC ), Centre Hospitalier Régional Universitaire de Nancy ( CHRU Nancy ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Université de Lorraine ( UL ), Paris-Centre de Recherche Cardiovasculaire ( PARCC - U970 ), Université Paris Descartes - Paris 5 ( UPD5 ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Hôpital Européen Georges Pompidou [APHP] ( HEGP ), Hôpital Européen Georges Pompidou [APHP] ( HEGP ), Université Paris Descartes - Faculté de Médecine ( UPD5 Médecine ), Université Paris Descartes - Paris 5 ( UPD5 ), Institut de Génétique Moléculaire de Montpellier ( IGMM ), Université de Montpellier ( UM ) -Centre National de la Recherche Scientifique ( CNRS ), Universitätsklinikum Essen [Universität Duisburg-Essen] ( Uniklinik Essen ), Centre Hospitalier Universitaire d'Angers ( CHU Angers ), PRES Université Nantes Angers Le Mans ( UNAM ), Université Paris Descartes - Paris 5 (UPD5)-Hôpital Européen Georges Pompidou [APHP] (HEGP), Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO)-Assistance publique - Hôpitaux de Paris (AP-HP) (APHP)-Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO)-Assistance publique - Hôpitaux de Paris (AP-HP) (APHP)-Institut National de la Santé et de la Recherche Médicale (INSERM), Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO)-Assistance publique - Hôpitaux de Paris (AP-HP) (APHP)-Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO)-Assistance publique - Hôpitaux de Paris (AP-HP) (APHP), Université Nice Sophia Antipolis (1965 - 2019) (UNS), COMUE Université Côte d'Azur (2015-2019) (COMUE UCA)-COMUE Université Côte d'Azur (2015-2019) (COMUE UCA)-Centre National de la Recherche Scientifique (CNRS)-Université Côte d'Azur (UCA), and Université de Montpellier (UM)-Centre National de la Recherche Scientifique (CNRS)

Subjects
Male, Serum Response Factor, MESH : RNA, Messenger, Vasodilator Agents, MESH: Vascular Resistance, MESH : Actins, MESH : Blotting, Western, Filamin, Mechanotransduction, Cellular, Muscle, Smooth, Vascular, MESH : Mechanotransduction, Cellular, Mice, Contractile Proteins, 0302 clinical medicine, MESH: Protein Kinase Inhibitors, MESH: Animals, MESH: Myosin-Light-Chain Kinase, MESH : Patch-Clamp Techniques, MESH : p38 Mitogen-Activated Protein Kinases, 0303 health sciences, Microscopy, Confocal, MESH : Vasoconstrictor Agents, MESH : Gene Expression Regulation, Electrical impedance myography, MESH : Reverse Transcriptase Polymerase Chain Reaction, Arteries, MESH : Vascular Resistance, MESH : Arterial Pressure, MESH: Serum Response Factor, MESH: Vasoconstriction, MESH : Tail, MESH : Vasoconstriction, Cardiology and Cardiovascular Medicine, Tail, medicine.medical_specialty, Blotting, Western, MESH: Myography, MESH: Actins, MESH : Vasodilator Agents, 03 medical and health sciences, [SDV.MHEP.CSC]Life Sciences [q-bio]/Human health and pathology/Cardiology and cardiovascular system, Serum response factor, MESH: Membrane Potentials, MESH: Blotting, Western, MESH: Contractile Proteins, RNA, Messenger, MESH : Microscopy, Confocal, Myosin-Light-Chain Kinase, MESH: Arteries, Phenylephrine, Dose-Response Relationship, Drug, MESH: Mechanotransduction, Cellular, MESH : Muscle, Smooth, Vascular, MESH : Protein Kinase Inhibitors, MESH: Vasodilator Agents, Endocrinology, Vasoconstriction, Vascular Resistance, Patch-Clamp Techniques, Time Factors, Contraction (grammar), Medizin, MESH : Dose-Response Relationship, Drug, 030204 cardiovascular system & hematology, MESH : Filamins, p38 Mitogen-Activated Protein Kinases, MESH: Mice, Knockout, Membrane Potentials, MESH: Dose-Response Relationship, Drug, MESH : Vasodilation, MESH : Calcium Signaling, MESH: Reverse Transcriptase Polymerase Chain Reaction, Vasoconstrictor Agents, MESH : Membrane Potentials, MESH: Microscopy, Confocal, MESH : Contractile Proteins, MESH: Filamins, Mice, Knockout, MESH : Myography, Reverse Transcriptase Polymerase Chain Reaction, MESH: Real-Time Polymerase Chain Reaction, Microfilament Proteins, MESH: Tail, MESH: Muscle, Smooth, Vascular, [ SDV.MHEP.CSC ] Life Sciences [q-bio]/Human health and pathology/Cardiology and cardiovascular system, MESH: Gene Expression Regulation, MESH: Myosin Light Chains, Vasodilation, MESH : Time Factors, medicine.drug, Myosin Light Chains, Myosin light-chain kinase, MESH : Microfilament Proteins, Filamins, MESH : Male, MESH: Arterial Pressure, MESH : Myosin Light Chains, MESH : Real-Time Polymerase Chain Reaction, Biology, Real-Time Polymerase Chain Reaction, MESH : Myosin-Light-Chain Kinase, MESH: Calcium Signaling, MESH: Vasodilation, MESH: Microfilament Proteins, MESH: Vasoconstrictor Agents, Internal medicine, MESH : Mice, MESH: Patch-Clamp Techniques, medicine, Animals, Arterial Pressure, Calcium Signaling, Patch clamp, Protein Kinase Inhibitors, MESH: Mice, Actin, MESH: RNA, Messenger, 030304 developmental biology, MESH: Time Factors, Myography, Actins, MESH : Arteries, MESH: Male, MESH: p38 Mitogen-Activated Protein Kinases, Gene Expression Regulation, MESH : Mice, Knockout, MESH : Serum Response Factor, MESH : Animals
Abstract

Objective— In resistance arteries, diameter adjustment in response to pressure changes depends on the vascular cytoskeleton integrity. Serum response factor (SRF) is a dispensable transcription factor for cellular growth, but its role remains unknown in resistance arteries. We hypothesized that SRF is required for appropriate microvascular contraction. Methods and Results— We used mice in which SRF was specifically deleted in smooth muscle or endothelial cells, and their control. Myogenic tone and pharmacological contraction was determined in resistance arteries. mRNA and protein expression were assessed by quantitative real-time PCR (qRT-PCR) and Western blot. Actin polymerization was determined by confocal microscopy. Stress-activated channel activity was measured by patch clamp. Myogenic tone developing in response to pressure was dramatically decreased by SRF deletion (5.9±2.3%) compared with control (16.3±3.2%). This defect was accompanied by decreases in actin polymerization, filamin A, myosin light chain kinase and myosin light chain expression level, and stress-activated channel activity and sensitivity in response to pressure. Contractions induced by phenylephrine or U46619 were not modified, despite a higher sensitivity to p38 blockade; this highlights a compensatory pathway, allowing normal receptor-dependent contraction. Conclusion— This study shows for the first time that SRF has a major part to play in the control of local blood flow via its central role in pressure-induced myogenic tone in resistance arteries.

Published
2013

2. IgG from patients with pulmonary arterial hypertension and/or systemic sclerosis binds to vascular smooth muscle cells and induces cell contraction

Author

Cynthia Calzas, Khaldoun Ghazal, Y Sahbatou, Nada Kherbeck, Loïc Guillevin, Babette B. Weksler, Franck Verrecchia, Mathieu C. Tamby, Hanadi Dib, Cédric Broussard, Marc Humbert, Gérald Simonneau, Azzedine Yaici, Véronique Witko-Sarsat, Guillaume Bussone, Claire Sanson, Luc Mouthon, Institut Cochin (IC UM3 (UMR 8104 / U1016)), Centre National de la Recherche Scientifique (CNRS)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM), Physiopathologie des Adaptations Nutritionnelles (PhAN), Université de Nantes - UFR de Médecine et des Techniques Médicales (UFR MEDECINE), Université de Nantes (UN)-Université de Nantes (UN)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE), Service de pneumologie [Béclère], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-AP-HP - Hôpital Antoine Béclère [Clamart], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), Hypertension arterielle pulmonaire physiopathologie et innovation thérapeutique, Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre chirurgical Marie Lannelongue, Service de médecine interne et centre de référence des maladies rares [CHU Cochin], Hôpital Cochin [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Physiopathologie de la Résorption Osseuse et Thérapie des Tumeurs Osseuses Primitives, Université de Nantes (UN)-Institut National de la Santé et de la Recherche Médicale (INSERM), Université Paris-Sud - Paris 11 (UP11)-Assistance publique - Hôpitaux de Paris (AP-HP) (APHP)-Hôpital Antoine Béclère, Centre chirurgical Marie Lannelongue-Institut National de la Santé et de la Recherche Médicale (INSERM), Assistance publique - Hôpitaux de Paris (AP-HP) (APHP)-CHU Cochin [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpital Cochin [AP-HP], Institut Cochin ( UM3 (UMR 8104 / U1016) ), Université Paris Descartes - Paris 5 ( UPD5 ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Centre National de la Recherche Scientifique ( CNRS ), Université de Nantes ( UN ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ), Université Paris-Sud - Paris 11 ( UP11 ) -Assistance publique - Hôpitaux de Paris (AP-HP)-Hôpital Antoine Béclère, Centre chirurgical Marie Lannelongue-Institut National de la Santé et de la Recherche Médicale ( INSERM ), and Assistance publique - Hôpitaux de Paris (AP-HP)-CHU Cochin [AP-HP]

Subjects
Male, Pathology, MESH : Hypertension, Pulmonary, Vascular smooth muscle, MESH: Muscle Contraction, MESH : Aged, 030204 cardiovascular system & hematology, MESH: Heat-Shock Proteins, MESH : Fluorescent Antibody Technique, Indirect, Autoantigens, Muscle, Smooth, Vascular, MESH: Scleroderma, Systemic, 0302 clinical medicine, MESH: Collagen, [ SDV.IMM ] Life Sciences [q-bio]/Immunology, Immunology and Allergy, MESH : Female, Fluorescent Antibody Technique, Indirect, Cells, Cultured, Heat-Shock Proteins, MESH: Aged, MESH: Immunoglobulin G, 0303 health sciences, MESH: Middle Aged, medicine.diagnostic_test, MESH: Muscle, Smooth, Vascular, MESH : Adult, Middle Aged, Connective tissue disease, 3. Good health, medicine.anatomical_structure, MESH: Autoantigens, MESH: Young Adult, [SDV.IMM]Life Sciences [q-bio]/Immunology, Female, Collagen, Blood vessel, Artery, Muscle Contraction, MESH: Cells, Cultured, Adult, medicine.medical_specialty, MESH : Immunoglobulin G, MESH : Male, Hypertension, Pulmonary, Immunology, MESH : Young Adult, MESH : Scleroderma, Systemic, Immunofluorescence, General Biochemistry, Genetics and Molecular Biology, 03 medical and health sciences, Young Adult, Rheumatology, MESH : Cells, Cultured, medicine, Humans, MESH: Fluorescent Antibody Technique, Indirect, MESH : Middle Aged, 030304 developmental biology, Aged, Autoimmune disease, Scleroderma, Systemic, MESH: Humans, MESH: Hypertension, Pulmonary, business.industry, MESH : Humans, MESH : Muscle, Smooth, Vascular, MESH: Adult, MESH : Heat-Shock Proteins, medicine.disease, Pulmonary hypertension, MESH: Male, MESH : Autoantigens, Phosphoprotein, Immunoglobulin G, MESH : Collagen, MESH : Muscle Contraction, business, MESH: Female
Abstract

International audience; OBJECTIVES: Pulmonary arterial hypertension (PAH) is characterised by remodelling of pulmonary arteries with enhanced vascular smooth muscle cell (VSMC) contraction, migration and proliferation. The authors investigated the presence of antibodies to human VSMCs in the serum of patients with systemic sclerosis with or without PAH and idiopathic PAH (iPAH). METHODS AND RESULTS: Antibodies to VSMCs were detected by immunofluorescence in sera from healthy controls and patients with scleroderma without PAH, scleroderma-associated PAH and iPAH. Serum IgG from these patients induced contraction of VSMCs in a collagen matrix in contrast with IgG from healthy controls. Several protein spots of interest and target antigens were identified by two-dimensional immunoblotting and MS, including stress-induced phosphoprotein 1 and α-enolase. Finally, antibodies to stress-induced phosphoprotein 1 were detected by ELISA in sera from 84%, 76% and 24% of patients with scleroderma without PAH, scleroderma-associated PAH and iPAH, respectively, compared with only 3% of healthy controls. CONCLUSION: The authors have identified IgG that binds to VSMCs in the serum of patients with scleroderma and iPAH. These antibodies may be pathogenic by modulating vascular contraction. The target antigens of these antibodies are stress-induced phosphoprotein 1 and α-enolase.

Published
2012

3. A label-free biosensor assay for botulinum neurotoxin B in food and human serum

Author

Christelle Mazuet, Christian Lévêque, Emmanuel Jover, Géraldine Ferracci, Michael Seagar, Marie-Pierre Blanchard, Séverine Marconi, Michel R. Popoff, Centre de recherche en neurobiologie - neurophysiologie de Marseille ( CRN2M ), Aix Marseille Université ( AMU ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Centre National de la Recherche Scientifique ( CNRS ), Carcinogenèse Hépatique et Virologie Moléculaire, Institut National de la Santé et de la Recherche Médicale ( INSERM ), Centre National de Référence des Bactéries Anaérobies et du Botulisme - Bactéries Anaérobies et Toxines ( CNR ), Institut Pasteur [Paris], Institut des Neurosciences Cellulaires et Intégratives ( INCI ), Université Louis Pasteur - Strasbourg I-Centre National de la Recherche Scientifique ( CNRS ), Unité de Neurobiologie des canaux Ioniques et de la Synapse ( UNIS - Inserm U1072 ), Centre de recherche en neurobiologie - neurophysiologie de Marseille (CRN2M), Aix Marseille Université (AMU)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Institut National de la Santé et de la Recherche Médicale (INSERM), Centre National de Référence des Bactéries Anaérobies et Botulisme - National Reference Center Anaerobic Bacteria and Botulism (CNR), Institut Pasteur [Paris] (IP), Institut des Neurosciences Cellulaires et Intégratives (INCI), Université Louis Pasteur - Strasbourg I-Centre National de la Recherche Scientifique (CNRS), Unité de Neurobiologie des canaux Ioniques et de la Synapse (UNIS - Inserm U1072), and Centre National de Référence des Bactéries Anaérobies et du Botulisme - Bactéries Anaérobies et Toxines (CNR)

Subjects
Serum, Botulinum Toxins, MESH: Annexin A5, Vesicle-Associated Membrane Protein 2, medicine.medical_treatment, MESH : Corpus Striatum, 01 natural sciences, Biochemistry, Substrate Specificity, MESH: Botulinum Toxins, MESH: Helium, Mice, MESH : Glutamic Acid, Clostridium botulinum, MESH : Serum, MESH: Animals, MESH: Receptors, GABA-A, MESH : Adaptation, Physiological, MESH: Vesicle-Associated Membrane Protein 2, 0303 health sciences, VAMP2, MESH : Substrate Specificity, Vesicle, In vitro toxicology, MESH: Reference Values, MESH: Serum, MESH: Glutamic Acid, MESH : Protein Array Analysis, MESH : Pulmonary Artery, 3. Good health, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biomolecules [q-bio.BM], MESH: Surface Plasmon Resonance, MESH : Thrombosis, MESH: N-Methylaspartate, MESH : Inert Gas Narcosis, MESH : Botulism, MESH : Synaptic Vesicles, Biological Assay, MESH: Membrane Proteins, MESH : Food, MESH : Surface Plasmon Resonance, MESH : Dopamine, MESH: Pulmonary Artery, Biophysics, MESH: Anesthetics, MESH: Botulism, MESH: Biological Assay, MESH: Tetralogy of Fallot, MESH : Atmospheric Pressure, Synaptic vesicle, MESH : Neurons, MESH : Food Analysis, 03 medical and health sciences, In vivo, Humans, MESH: Thrombosis, Molecular Biology, MESH : Crystallography, MESH: Receptors, N-Methyl-D-Aspartate, Protease, MESH: Humans, MESH : Tetralogy of Fallot, 010401 analytical chemistry, MESH: Child, Preschool, MESH : Humans, MESH: Atmospheric Pressure, MESH : Receptors, GABA-A, MESH : Muscle, Smooth, Vascular, Surface Plasmon Resonance, MESH : Clostridium botulinum, medicine.disease, 0104 chemical sciences, MESH: Peritoneovenous Shunt, Food, MESH : Membrane Proteins, MESH : Lipid Bilayers, MESH : Biological Assay, MESH : Receptors, N-Methyl-D-Aspartate, MESH : Annexin A5, Food Analysis, MESH: Neurons, Biosensing Techniques, MESH : Child, Preschool, MESH : Nitrogen, MESH: Corpus Striatum, [ SDV.BBM.BC ] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biomolecules [q-bio.BM], Botulism, Botulinum Toxins, Type A, MESH: Nitrogen, Chemistry, MESH: Protein Array Analysis, MESH : Rats, MESH: Lipid Bilayers, MESH: Crystallography, MESH : Hydrogen, MESH: Hydrogen, MESH: Muscle, Smooth, Vascular, MESH : Anesthetics, MESH : Botulinum Toxins, MESH: Fibrosis, MESH : Fibrosis, MESH: Inert Gas Narcosis, MESH : Urate Oxidase, Synaptic Vesicles, MESH : Peritoneovenous Shunt, MESH: Urate Oxidase, MESH : Vesicle-Associated Membrane Protein 2, MESH: Biosensing Techniques, MESH: Food, Carrot juice, MESH: Clostridium botulinum, MESH: Rats, MESH: Food Analysis, Protein Array Analysis, MESH: Substantia Nigra, MESH: Dopamine, In Vitro Techniques, MESH : Reference Values, MESH : Helium, MESH : Mice, medicine, Animals, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM], MESH: Mice, 030304 developmental biology, MESH : Substantia Nigra, MESH : Biosensing Techniques, Cell Biology, MESH: Synaptic Vesicles, Molecular biology, MESH: Adaptation, Physiological, Rats, MESH: Substrate Specificity, MESH : Animals, MESH : N-Methylaspartate
Abstract

International audience; Botulinum neurotoxins (BoNTs) are among the most toxic substances known. Surveillance and diagnostics require methods for rapid detection of BoNTs in complex media such as foodstuffs and human serum. We have developed in vitro assays to specifically detect the protease activity of botulinum neurotoxin B (BoNT/B) on a time scale of minutes. Cleavage of the BoNT/B substrate VAMP2, a membrane SNARE protein associated with synaptic vesicles, was monitored using real-time surface plasmon resonance to measure vesicle capture by specific antibodies coupled to microchips. The assay is functional in low-ionic-strength buffers and stable over a wide range of pH values (5.5-9.0). Endoproteolytic cleavage of VAMP2 was detected in 10 min with 2 pM native BoNT/B holotoxin. Contamination of liquid food products such as carrot juice, apple juice, and milk with low picomolar amounts of BoNT/B was revealed within 3h. BoNT/B activity was detected in sera from patients with type B botulism but not in healthy controls or patients with other neurological diseases. This robust, sensitive, and rapid protein chip assay is appropriate for monitoring BoNT/B in food products and diagnostic tests for type B botulism and could replace the current in vivo mouse bioassay.

Published
2011

4. TRPC1 is regulated by caveolin-1 and is involved in oxidized LDL-induced apoptosis of vascular smooth muscle cells

Author

Bertrand Marcheix, Cécile Vindis, Cécile Ingueneau, Anne Nègre-Salvayre, Anne Athias, Robert Salvayre, Philippe Gambert, Uyen Huynh-Do, Institut de médecine moléculaire de Rangueil ( I2MR ), Université Paul Sabatier - Toulouse 3 ( UPS ) -IFR31-IFR150-Institut National de la Santé et de la Recherche Médicale ( INSERM ), Departement of Nephrology and Hypertension, and Department of Clinical research, University of Bern, Plateforme Lipidomique [Dijon] ( LAP ), Lipides - Nutrition - Cancer (U866) ( LNC ), Université de Bourgogne ( UB ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ) -AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement-Ecole Nationale Supérieure de Biologie Appliquée à la Nutrition et à l'Alimentation de Dijon ( ENSBANA ) -Université de Bourgogne ( UB ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ) -AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement-Ecole Nationale Supérieure de Biologie Appliquée à la Nutrition et à l'Alimentation de Dijon ( ENSBANA ) -IFR100 - Structure fédérative de recherche Santé-STIC-Centre Hospitalier Universitaire de Dijon - Hôpital François Mitterrand ( CHU Dijon ), Institut de médecine moléculaire de Rangueil (I2MR), Université Toulouse III - Paul Sabatier (UT3), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées- Institut Fédératif de Recherche Bio-médicale Institution (IFR150)-Institut National de la Santé et de la Recherche Médicale (INSERM), Plateforme Lipidomique [Dijon] (LAP), Lipides - Nutrition - Cancer (U866) (LNC), Université de Bourgogne (UB)-Institut National de la Santé et de la Recherche Médicale (INSERM)-AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement-Ecole Nationale Supérieure de Biologie Appliquée à la Nutrition et à l'Alimentation de Dijon (ENSBANA)-Université de Bourgogne (UB)-Institut National de la Santé et de la Recherche Médicale (INSERM)-AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement-Ecole Nationale Supérieure de Biologie Appliquée à la Nutrition et à l'Alimentation de Dijon (ENSBANA)-IFR100 - Structure fédérative de recherche Santé-STIC-Centre Hospitalier Universitaire de Dijon - Hôpital François Mitterrand (CHU Dijon), and Simon, Marie Francoise

Subjects
MESH: Lipoproteins, LDL, Vascular smooth muscle, Oxysterol, Caveolin 1, Apoptosis, Biology, MESH: Base Sequence, MESH : RNA, Small Interfering, Muscle, Smooth, Vascular, TRPC1, Transient receptor potential channel, MESH: RNA, Small Interfering, MESH : Cells, Cultured, Humans, MESH: Caveolin 1, RNA, Small Interfering, MESH: TRPC Cation Channels, Cells, Cultured, TRPC Cation Channels, MESH: Humans, Base Sequence, MESH : Gene Expression Regulation, MESH: Apoptosis, MESH : Humans, MESH : TRPC Cation Channels, MESH : Muscle, Smooth, Vascular, Articles, Cell Biology, MESH: Muscle, Smooth, Vascular, Actin cytoskeleton, MESH: Gene Expression Regulation, Cell biology, Lipoproteins, LDL, Gene Expression Regulation, MESH : Caveolin 1, Molecular Medicine, lipids (amino acids, peptides, and proteins), MESH : Base Sequence, MESH : Lipoproteins, LDL, Homeostasis, MESH : Apoptosis, MESH: Cells, Cultured
Abstract

International audience; Oxidized low-density lipoprotein (oxLDL) induced-apoptosis of vascular cells may participate in plaque instability and rupture. We have previously shown that vascular smooth muscle cells (VSMC) stably expressing caveolin-1 were more susceptible to oxLDL-induced apoptosis than VSMC expressing lower level of caveolin-1, and this was correlated with enhanced Ca(2+) entry and pro-apoptotic events. In this study, we aimed to identify the molecular events involved in oxLDL-induced Ca(2+) influx and their regulation by the structural protein caveolin-1. In VSMC, transient receptor potential canonical-1 (TRPC1) silencing by ARN interference prevents the Ca(2+) influx and reduces the toxicity induced by oxLDL. Moreover, caveolin-1 silencing induces concomitant decrease of TRPC1 expression and reduces oxLDL-induced apoptosis of VSMC. OxLDL enhanced the cell surface expression of TRPC1, as shown by biotinylation of cell surface proteins, and induced TRPC1 translocation into caveolar compartment, as assessed by subcellular fractionation. OxLDL-induced TRPC1 translocation was dependent on actin cytoskeleton and associated with a dramatic rise of 7-ketocholesterol (a major oxysterol in oxLDL) into caveolar membranes, whereas the caveolar content of cholesterol was unchanged. Altogether, the reported results show that TRPC1 channels play a role in Ca(2+) influx and Ca(2+) homeostasis deregulation that mediate apoptosis induced by oxLDL. These data also shed new light on the role of caveolin-1 and caveolar compartment as important regulators of TRPC1 trafficking to the plasma membrane and apoptotic processes that play a major role in atherosclerosis.

Published
2009

5. Plasminogen activation: a mediator of vascular smooth muscle cell apoptosis in atherosclerotic plaques

Author

Rossignol, Patrick, Luttun, Aernout, Martin-Ventura, Jose Luis, Lupu, Florea, Carmeliet, Peter, Collen, Désiré, Anglès-Cano, Eduardo, Lijnen, Henri Roger, Center for Molecular and Vascular Biology, Catholic University of Leuven ( KU Leuven ), Center for Transgene Technology and Gene Therapy, Flanders Interuniversity Institute for Biotechnology, Hémostase, bio-ingénierie et remodelage cardiovasculaires ( LBPC ), Université Paris Diderot - Paris 7 ( UPD7 ) -Université Paris 13 ( UP13 ) -Université Sorbonne Paris Cité ( USPC ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Institut Galilée, Cardiovascular Biology Research Program, Oklahoma Medical Research Foundation, Sérine protéases et physiopathologie de l'unité neurovasculaire, Université de Caen Normandie ( UNICAEN ), Normandie Université ( NU ) -Normandie Université ( NU ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ), Inserm, Leducq Foundation, Catholic University of Leuven - Katholieke Universiteit Leuven (KU Leuven), Hémostase, bio-ingénierie et remodelage cardiovasculaires (LBPC), Université Paris Diderot - Paris 7 (UPD7)-Université Paris 13 (UP13)-Université Sorbonne Paris Cité (USPC)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut Galilée, Oklahoma Medical Research Foundation (OMRF), Université de Caen Normandie (UNICAEN), and Normandie Université (NU)-Normandie Université (NU)-Institut National de la Santé et de la Recherche Médicale (INSERM)

Subjects
MESH : Antiplasmin, MESH : Plasminogen Activator Inhibitor 1, MESH : Atherosclerosis, [SDV.BC]Life Sciences [q-bio]/Cellular Biology, MESH : Mice, Inbred C57BL, MESH: Mice, Knockout, MESH: Plasminogen Activator Inhibitor 1, MESH : Tunica Media, MESH: Atherosclerosis, MESH: Mice, Inbred C57BL, MESH: Antiplasmin, MESH: Plasminogen, MESH : Plasmin, MESH : Apolipoproteins E, MESH : Mice, MESH: Animals, genetically altered mice, MESH: Mice, MESH: Plasmin, [ SDV.BC ] Life Sciences [q-bio]/Cellular Biology, MESH: Apoptosis, apoptosis, MESH : Aorta, MESH: Aorta, MESH : Muscle, Smooth, Vascular, MESH: Muscle, Smooth, Vascular, MESH: Apolipoproteins E, MESH : Disease Models, Animal, MESH: Tunica Media, MESH : Plasminogen, cardiovascular system, plasminogen, MESH : Mice, Knockout, MESH : Animals, MESH: Disease Models, Animal, atherosclerosis, vascular smooth muscle cell, MESH : Apoptosis
Abstract

21 pages; International audience; BACKGROUND: Apoptosis of vascular cells is considered to be a major determinant of atherosclerotic plaque vulnerability and potential rupture. Plasmin can be generated in atherosclerotic plaques and recent in vitro data suggest that plasminogen activation may trigger vascular smooth muscle cell (VSMC) apoptosis. AIM: To determine whether plasminogen activation may induce aortic VSMC apoptosis ex vivo and in vivo. METHODS AND RESULTS: Mice with single or combined deficiencies of apolipoprotein E (ApoE) and plasminogen activator inhibitor-1 (PAI-1) were used. Ex vivo incubation with plasminogen of isolated aortic tunica media from PAI-1-deficient mice induced plasminogen activation and VSMC apoptosis, which was inhibited by alpha2-antiplasmin. In vivo, levels of plasmin, active caspase 3 and VSMC apoptotic index were significantly higher in atherosclerotic aortas from mice with combined ApoE and PAI-1 deficiencies than in those from littermates with single ApoE deficiency. A parallel decrease in VSMC density was observed. CONCLUSIONS: These data strongly suggest that plasminogen activation may contribute to VSMC apoptosis in atherosclerotic plaques.

Published
2006

6. Serotonin transporter inhibition prevents and reverses monocrotaline-induced pulmonary hypertension in rats

Author

Serge Adnot, Bernadette Raffestin, Saadia Eddahibi, William Raoul, Patricia Zadigue, Dominique Rideau, Rima Benferhat, Christophe Guignabert, Michel Hamon, Physiopathologie et Thérapeutiques Respiratoires, Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpital Henri Mondor-Institut National de la Santé et de la Recherche Médicale (INSERM)-IFR10, Service de chirurgie thoracique et d'anatomopathologie, Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpital Hôtel Dieu, Département de physiologie, Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpital Ambroise Paré [AP-HP]-Université Paris Descartes - Paris 5 (UPD5), Neuropsychopharmacologie moléculaire, cellulaire et fonctionnelle, Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut National de la Santé et de la Recherche Médicale (INSERM), This research was supported by grants from INSERM, the Ministère de la Recherche, the Délégation à la Recherche Clinique de l'AP-HP, and the Fondation de France. We are grateful to the pharmaceutical companies Janssen, Lilly, Lundbeck, Roche, and SmithKline Beecham for generously donating the drugs used in this study., Assistance publique - Hôpitaux de Paris (AP-HP)-Hôpital Henri Mondor-Institut National de la Santé et de la Recherche Médicale ( INSERM ) -IFR10, Assistance publique - Hôpitaux de Paris (AP-HP)-Hôpital Hôtel Dieu, Assistance publique - Hôpitaux de Paris (AP-HP)-Hôpital Ambroise Paré-Université Paris Descartes - Paris 5 ( UPD5 ), and Université Pierre et Marie Curie - Paris 6 ( UPMC ) -Institut National de la Santé et de la Recherche Médicale ( INSERM )

Subjects
MESH : Hypertension, Pulmonary, MESH : Monocrotaline, muscle, MESH: Receptor, Serotonin, 5-HT1B, MESH: Receptor, Serotonin, 5-HT1D, [CHIM.THER]Chemical Sciences/Medicinal Chemistry, 030204 cardiovascular system & hematology, [ SDV.BA ] Life Sciences [q-bio]/Animal biology, [SDV.MHEP.PSR]Life Sciences [q-bio]/Human health and pathology/Pulmonology and respiratory tract, MESH : Serotonin Plasma Membrane Transport Proteins, Muscle, Smooth, Vascular, MESH: Monocrotaline, chemistry.chemical_compound, 0302 clinical medicine, MESH : Receptor, Serotonin, 5-HT2A, MESH : Cell Proliferation, MESH: Up-Regulation, Medicine, MESH: Animals, MESH: Serotonin Plasma Membrane Transport Proteins, Neurotransmitter, Receptor, MESH : Up-Regulation, Lung, Serotonin transporter, remodeling, Serotonin Plasma Membrane Transport Proteins, Monocrotaline, biology, MESH : Rats, [ SDV.MHEP.PHY ] Life Sciences [q-bio]/Human health and pathology/Tissues and Organs [q-bio.TO], [SDV.BA]Life Sciences [q-bio]/Animal biology, [ CHIM.THER ] Chemical Sciences/Medicinal Chemistry, MESH: Muscle, Smooth, Vascular, [ SDV.MHEP.CSC ] Life Sciences [q-bio]/Human health and pathology/Cardiology and cardiovascular system, 3. Good health, Up-Regulation, MESH: Serotonin Uptake Inhibitors, medicine.anatomical_structure, Serotonin 5-HT2 Receptor Antagonists, Cardiology and Cardiovascular Medicine, Selective Serotonin Reuptake Inhibitors, smooth, medicine.medical_specialty, MESH: Rats, MESH : Lung, Hypertension, Pulmonary, [SDV.BC]Life Sciences [q-bio]/Cellular Biology, Serotonin 5-HT1 Receptor Antagonists, 03 medical and health sciences, Downregulation and upregulation, [SDV.MHEP.CSC]Life Sciences [q-bio]/Human health and pathology/Cardiology and cardiovascular system, Physiology (medical), Internal medicine, [ SDV.MHEP ] Life Sciences [q-bio]/Human health and pathology, MESH: Cell Proliferation, Fluoxetine, pulmonary heart disease, [SDV.MHEP.PHY]Life Sciences [q-bio]/Human health and pathology/Tissues and Organs [q-bio.TO], MESH: Fluoxetine, Animals, MESH: Lung, Cell Proliferation, MESH: Hypertension, Pulmonary, business.industry, MESH : Serotonin Uptake Inhibitors, [ SDV.BC ] Life Sciences [q-bio]/Cellular Biology, Monocyte, MESH: Receptor, Serotonin, 5-HT2A, MESH : Fluoxetine, MESH : Muscle, Smooth, Vascular, medicine.disease, Pulmonary hypertension, Rats, MESH : Receptor, Serotonin, 5-HT1B, Endocrinology, 030228 respiratory system, chemistry, biology.protein, MESH : Receptor, Serotonin, 5-HT1D, Serotonin, MESH : Animals, business, [ SDV.MHEP.PSR ] Life Sciences [q-bio]/Human health and pathology/Pulmonology and respiratory tract, [SDV.MHEP]Life Sciences [q-bio]/Human health and pathology
Abstract

Background—Progression of pulmonary hypertension (PH) is associated with increased lung expression of the serotonin transporter (5-HTT), which leads to hyperplasia of the pulmonary artery smooth muscle cells (PA-SMCs). Given the postulated causal relation between 5-HTT overexpression and PH, we herein investigated whether the highly selective 5-HTT inhibitor fluoxetine prevented and/or reversed PH induced by monocrotaline (MCT) in rats. Selective 5-HT1B/1D, 5-HT2A, and 5-HT2Breceptor antagonists were used for comparative testing.Methods and Results—MCT injection (60 mg/kg SC) was followed by an early peak in lung 5-HTT expression on day 1, which preceded the onset of PH. Established PH on day 15 was associated with a sustained 5-HTT increase. Continued fluoxetine treatment completely prevented PA-SMC proliferation and PH development and also suppressed the late 5-HTT increase, without affecting the early peak. The 5-HT receptor antagonists did not affect PH. Fluoxetine (10 mg · kg−1· d−1PO) started 3 weeks after MCT injection completely reversed established PH, normalizing PA pressure and structure. MCT-induced PH was also associated with increased expression of various cytokines, but only interleukin-1β and monocyte chemotactic protein-1 increased at the early phase and stimulated 5-HTT expression by cultured PA-SMCs.Conclusion—Upregulation of lung 5-HTT induced by MCT appears necessary to initiate the development of pulmonary vascular remodeling, whereas a sustained increase in 5-HTT expression may underlie both the progression and the maintenance of MCT-induced PH. Complete reversal of established PH by fluoxetine provides a rationale for new therapeutic strategies in human PH.

Published
2005

7. Resveratrol: preventing properties against vascular alterations and ageing

Author

Dominique Delmas, Norbert Latruffe, Brigitte Jannin, Laboratoire de Biochimie Moléculaire et Cellulaire ( LBMC ), Université de Bourgogne ( UB ), Lipides - Nutrition - Cancer (U866) ( LNC ), and Université de Bourgogne ( UB ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ) -AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement-Ecole Nationale Supérieure de Biologie Appliquée à la Nutrition et à l'Alimentation de Dijon ( ENSBANA )

Subjects
MESH : Oxidative Stress, Aging, Antioxidant, Platelet Aggregation, Arteriosclerosis, medicine.medical_treatment, Resveratrol, Pharmacology, medicine.disease_cause, Muscle, Smooth, Vascular, chemistry.chemical_compound, MESH : Vasodilation, MESH : Foam Cells, MESH : Platelet Aggregation, Stilbenes, MESH : Cardiovascular Diseases, MESH : Macrophages, chemistry.chemical_classification, Neovascularization, Pathologic, Phytoalexin, food and beverages, Vasodilation, Biochemistry, Cardiovascular Diseases, medicine.symptom, Biotechnology, Lipoproteins, Inflammation, Health Promotion, MESH : Lipoproteins, Biology, MESH : Stilbenes, MESH : Arteriosclerosis, medicine, Humans, [ SDV.BBM ] Life Sciences [q-bio]/Biochemistry, Molecular Biology, Reactive oxygen species, Mechanism (biology), Macrophages, MESH : Humans, MESH : Neovascularization, Pathologic, MESH : Muscle, Smooth, Vascular, MESH : Aging, Oxidative Stress, chemistry, Ageing, MESH : Health Promotion, Oxidative stress, Food Science, Foam Cells
Abstract

International audience; Cardiovascular diseases are the leading cause of death in developed countries where the common pathological substrate underlying this process is atherosclerosis. Several new concepts have emerged in relation to mechanisms that contribute to the regulation of the vascular diseases and associated inflammatory effects. Recently, potential antioxidants (vitamin E, polyphenols) have received much attention as potential anti-atherosclerotic agents. Among the polyphenols with health benefic properties, resveratrol, a phytoalexin of grape, seem to be a good candidate protecting the vascular walls from oxidation, inflammation, platelet aggregation, and thrombus formation. In this review, we focus on the mechanism of resveratrol cardiovascular benefic effects. We analyze, in relation with the different steps of atherosclerotic process, the resveratrol properties at multiple levels, such as cellular signaling, enzymatic pathways, apoptosis, and gene expression. We show and discuss the relationship with reactive oxygen species, regulation of pro-inflammatory genes including cycloxygenases and cytokines in molecular inflammatory and aging processes, and how the regulation of these activites by resveratrol can lead to a prevention of vascular diseases.

Published
2005

8. Chronology of cellular alterations during 7-ketocholesterol-induced cell death on A7R5 rat smooth muscle cells: analysis by time lapse-video microscopy and conventional fluorescence microscopy

Author

Zahm, Jean-Marie, Baconnais, Sonia, Monier, Serge, Bonnet, Noël, Bessède, Ginette, Gambert, Philippe, Puchelle, Edith, Lizard, Gérard, Dynamique cellulaire et moléculaire de la muqueuse respiratoire, Université de Reims Champagne-Ardenne (URCA)-IFR53-Institut National de la Santé et de la Recherche Médicale (INSERM), Laboratoire des Lipoprotéines humaines et interactions vasculaires, Université de Bourgogne (UB)-Institut National de la Santé et de la Recherche Médicale (INSERM), Université de Reims Champagne-Ardenne ( URCA ) -IFR53-Institut National de la Santé et de la Recherche Médicale ( INSERM ), Université de Bourgogne ( UB ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ), and Birembaut, Philippe

Subjects
MESH: Cell Death, Time Factors, MESH : Microscopy, Fluorescence, MESH : Actins, Apoptosis, Cell Count, MESH: Microscopy, Fluorescence, MESH : Cell Count, [SDV.MHEP.PSR]Life Sciences [q-bio]/Human health and pathology/Pulmonology and respiratory tract, Muscle, Smooth, Vascular, Membrane Potentials, MESH : Image Cytometry, MESH : Rat, MESH : Membrane Potentials, MESH: Animals, MESH : DNA Fragmentation, Enzyme Inhibitors, Ketocholesterols, Image Cytometry, Microscopy, Video, Cell Death, MESH : Rats, MESH : Microscopy, Video, MESH: Ketocholesterols, MESH: Muscle, Smooth, Vascular, Mitochondria, MESH: Enzyme Inhibitors, MESH : Mitochondria, MESH: Rats, MESH: Mitochondria, DNA Fragmentation, MESH: Actins, MESH: Cell Adhesion, MESH: Rat, Cell Adhesion, Animals, MESH: Membrane Potentials, MESH: DNA Fragmentation, MESH : Ketocholesterols, MESH : Enzyme Inhibitors, MESH: Cell Count, MESH: Apoptosis, Rats, Inbred Strains, MESH : Muscle, Smooth, Vascular, Actins, Rats, MESH: Microscopy, Video, MESH : Cell Adhesion, Microscopy, Fluorescence, MESH : Cell Death, [SDV.MHEP.PSR] Life Sciences [q-bio]/Human health and pathology/Pulmonology and respiratory tract, MESH : Animals, MESH : Apoptosis, [ SDV.MHEP.PSR ] Life Sciences [q-bio]/Human health and pathology/Pulmonology and respiratory tract, MESH: Image Cytometry
Abstract

BACKGROUND: Time-lapse video microscopy was used to determine whether mitochondrial and nuclear changes (decrease in mitochondrial transmembrane potential, condensation, and/or fragmentation of the nuclei, morphologic features typical of apoptosis) occurring during 7-ketocholesterol-induced cell death on A7R5 rat smooth muscle cells took place before or after the loss of cell adhesion. In addition, changes in actin organization were followed by conventional fluorescence microscopy. METHODS: Morphologic, functional, and spatial changes at the mitochondrial level were investigated with 3,3'-dihexyloxacarbocyanine iodide and/or MitoTracker Red, and nuclear morphology was characterized by staining with Hoechst 33342. Actin fibers, which are major components of the filament network of the cytoskeleton, were visualized with phalloidin linked to fluorescein. The numbers of adherent and nonadherent cells were determined by cell counting. RESULTS: 7-Ketocholesterol-induced cell death was associated with a rapid alteration of actin fibers, a loss of intercellular junctions, and cell shape modifications. Analysis of mitochondrial transmembrane potential showed successively a hyperpolarization and a more or less pronounced progressive decrease followed by a dramatic drop associated with an increase in Hoechst 33342 staining, reflecting chromatin condensation and morphologic changes in the nuclei. CONCLUSIONS: During cell death induced by 7-ketocholesterol in A7R5 rat smooth muscle cells, the different methods of microscopy allowed us to establish that alterations of actin fibers and mitochondrial dysfunctions occurred before condensation and/or fragmentation of the nuclei, which preceded the loss of cell adhesion.

Published
2003

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