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1. Transient viral exposure drives functionally-coordinated humoral immune responses in HIV-1 post-treatment controllers

Author

Molinos-Albert, Luis, Lorin, Valerie, Monceaux, Valérie, Orr, Sylvie, Essat, Asma, Dufloo, Jérémy, Schwartz, Olivier, Rouzioux, Christine, Meyer, Laurence, Hocqueloux, Laurent, Sáez-Cirión, Asier, Mouquet, Hugo, Group, Anrs Visconti Study, Prazuck, Thierry, Dieuleveult, Barbara De, Bani-Sadr, Firouzé, Hentzien, Maxime, Berger, Jean-Luc, Kmiec, Isabelle, Pichancourt, Gilles, Nasri, Safa, Hittinger, Gilles, Lambry, Véronique, Beauey, Anne-Cécile, Pialoux, Gilles, Palacios, Christia, Siguier, Martin, Adda, Anne, Foucoin, Jane, Weiss, Laurence, Karmochkine, Marina, Meghadecha, Mohamed, Ptak, Magali, Salmon-Ceron, Dominique, Blanche, Philippe, Piétri, Marie-Pierre, Molina, Jean-Michel, Taulera, Olivier, Lascoux-Combe, Caroline, Ponscarme, Diane, Bertaut, Jeannine Delgado, Makhloufi, Djamila, Godinot, Matthieu, Artizzu, Valérie, Yazdanpanah, Yazdan, Matheron, Sophie, Godard, Cindy, Julia, Zélie, Bernard, Louis, Bastides, Frédéric, Bourgault, Olivier, Jacomet, Christine, Goncalves, Emilie, Meybeck, Agnès, Huleux, Thomas, Cornavin, Pauline, Debab, Yasmine, Théron, David, Miailhes, Patrick, Cotte, Laurent, Pailhes, Sophie, Ogoudjobi, Stanislas, Viard, Jean Paul, Dulucq, Marie-Josée, Bodard, Loïc, Churaqui, Francoise, Guimard, Thomas, Laine, Laetitia, Immunologie humorale - Humoral Immunology, Institut Pasteur [Paris] (IP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Paris Cité (UPCité), HIV, Inflammation et persistance - HIV, Inflammation and Persistence, Institut Pasteur [Paris] (IP)-Université Paris Cité (UPCité), Centre de recherche en épidémiologie et santé des populations (CESP), Université de Versailles Saint-Quentin-en-Yvelines (UVSQ)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpital Paul Brousse-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Paris-Saclay, Virus et Immunité - Virus and immunity (CNRS-UMR3569), Institut Pasteur [Paris] (IP)-Centre National de la Recherche Scientifique (CNRS)-Université Paris Cité (UPCité), Service de Microbiologie Clinique [Paris], CHU Necker - Enfants Malades [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), La Source Hospital-CHR, Centre Hospitalier Universitaire de Reims (CHU Reims), Centre Hospitalier Henri Duffaut (Avignon), CHI de Toulon, CHU Tenon [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU), Hôpital Européen Georges Pompidou [APHP] (HEGP), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO), Hôpital Cochin [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), Hopital Saint-Louis [AP-HP] (AP-HP), Hôpital Edouard Herriot [CHU - HCL], Hospices Civils de Lyon (HCL), AP-HP - Hôpital Bichat - Claude Bernard [Paris], Hôpital Bretonneau, Centre Hospitalier Régional Universitaire de Tours (CHRU Tours), CHU Gabriel Montpied [Clermont-Ferrand], CHU Clermont-Ferrand, Centre Hospitalier Gustave Dron [Tourcoing], Hôpital Charles Nicolle [Rouen], CHU Rouen, Normandie Université (NU)-Normandie Université (NU), Hôpital de la Croix-Rousse [CHU - HCL], Hôpital Hôtel-Dieu [Paris], Institut mutualiste Monsouris (IMM), Centre Hospitalier Départemental - Hôpital de La Roche-sur-Yon (CHD Vendée), We thank the NIH AIDS Reagent Program (Division of AIDS, NIAID, NIH) for contributing to reagents and the Agence National de Recherche sur le SIDA et les hépatites virales (ANRS) for an equipment grant support (AO2013-2 #13553, H.M.). L.M. M-A. is the recipient of a postdoctoral fellowship from the Pasteur-Roux-Cantarini program (Institut Pasteur). H.M. received core grants from the Institut Pasteur, the INSERM and the Milieu Intérieur Program (ANR-10-LABX-69-01). This work was conducted in the context of the ANRS RHIVIERA consortium, and was supported by the ANRS and the NIH/NIAID (grant #: P01AI131365, H.M.)., ANRS VISCONTI Study Group Thierry Prazuck, Barbara De Dieuleveult, Firouzé Bani-Sadr, Maxime Hentzien, Jean-Luc Berger, Isabelle Kmiec, Gilles Pichancourt, Safa Nasri, Gilles Hittinger, Véronique Lambry, Anne-Cécile Beauey, Gilles Pialoux, Christia Palacios, Martin Siguier, Anne Adda, Jane Foucoin, Laurence Weiss, Marina Karmochkine, Mohamed Meghadecha, Magali Ptak, Dominique Salmon-Ceron, Philippe Blanche, Marie-Pierre Piétri, Jean-Michel Molina, Olivier Taulera, Caroline Lascoux-Combe, Diane Ponscarme, Jeannine Delgado Bertaut, Djamila Makhloufi, Matthieu Godinot, Valérie Artizzu, Yazdan Yazdanpanah, Sophie Matheron, Cindy Godard, Zélie Julia, Louis Bernard, Frédéric Bastides, Olivier Bourgault, Christine Jacomet, Emilie Goncalves, Agnès Meybeck, Thomas Huleux, Pauline Cornavin, Yasmine Debab, David Théron, Patrick Miailhes, Laurent Cotte, Sophie Pailhes, Stanislas Ogoudjobi, Jean Paul Viard, Marie-Josée Dulucq, Loïc Bodard, Francoise Churaqui, Thomas Guimard & Laetitia Laine, ANR-10-LABX-0069,MILIEU INTERIEUR,GENETIC & ENVIRONMENTAL CONTROL OF IMMUNE PHENOTYPE VARIANCE: ESTABLISHING A PATH TOWARDS PERSONALIZED MEDICINE(2010), Institut Pasteur [Paris]-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Paris Cité (UPC), Institut Pasteur [Paris]-Université Paris Cité (UPC), Virus et Immunité - Virus and immunity, and Institut Pasteur [Paris]-Centre National de la Recherche Scientifique (CNRS)-Université Paris Cité (UPC)

Subjects
Multidisciplinary, MESH: Humans, General Physics and Astronomy, HIV Infections, General Chemistry, MESH: HIV Infections, Antibodies, Neutralizing, General Biochemistry, Genetics and Molecular Biology, Immunity, Humoral, MESH: Antibodies, Neutralizing, MESH: HIV-1, [SDV.MP.VIR]Life Sciences [q-bio]/Microbiology and Parasitology/Virology, HIV-1, Humans, Viremia, MESH: Immunity, Humoral, MESH: Viremia
Abstract

HIV-1 post-treatment controllers are rare individuals controlling HIV-1 infection for years after antiretroviral therapy interruption. Identification of immune correlates of control in post-treatment controllers could aid in designing effective HIV-1 vaccine and remission strategies. Here, we perform comprehensive immunoprofiling of the humoral response to HIV-1 in long-term post-treatment controllers. Global multivariate analyses combining clinico-virological and humoral immune data reveal distinct profiles in post-treatment controllers experiencing transient viremic episodes off therapy compared to those stably aviremic. Virally-exposed post-treatment controllers display stronger HIV-1 humoral responses, and develop more frequently Env-specific memory B cells and cross-neutralizing antibodies. Both are linked to short viremic exposures, which are also accompanied by an increase in blood atypical memory B cells and activated subsets of circulating follicular helper T cells. Still, most humoral immune variables only correlate with Th2-like circulating follicular helper T cells. Thus, post-treatment controllers form a heterogeneous group with two distinct viral behaviours and associated immune signatures. Post-treatment controllers stably aviremic present “silent” humoral profiles, while those virally-exposed develop functionally robust HIV-specific B-cell and antibody responses, which may participate in controlling infection.

Published
2022

2. Novel role of UHRF1 in the epigenetic repression of the latent HIV-1

Author

Roxane Verdikt, Maryam Bendoumou, Sophie Bouchat, Lorena Nestola, Alexander O. Pasternak, Gilles Darcis, Véronique Avettand-Fenoel, Caroline Vanhulle, Amina Aït-Ammar, Marion Santangelo, Estelle Plant, Valentin Le Douce, Nadège Delacourt, Aurelija Cicilionytė, Coca Necsoi, Francis Corazza, Caroline Pereira Bittencourt Passaes, Christian Schwartz, Martin Bizet, François Fuks, Asier Sáez-Cirión, Christine Rouzioux, Stéphane De Wit, Ben Berkhout, Virginie Gautier, Olivier Rohr, Carine Van Lint, Université libre de Bruxelles (ULB), University of Amsterdam [Amsterdam] (UvA), Université de Liège, CHU Necker - Enfants Malades [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), Institut Cochin (IC UM3 (UMR 8104 / U1016)), Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Université Paris Cité (UPCité), University College Dublin [Dublin] (UCD), Institut Pasteur [Paris] (IP), HIV, Inflammation et persistance - HIV, Inflammation and Persistence, Institut Pasteur [Paris] (IP)-Université Paris Cité (UPCité), Université de Strasbourg (UNISTRA), CVL acknowledges funding from the Belgian National Fund for Scientific Research (F.R.S.-FNRS, Belgium), the « Fondation Roi Baudouin », the NEAT (European AIDS Treatment Network) program, the Internationale Brachet Stiftung, ViiV Healthcare, the Walloon Region (« Fonds de Maturation »), « Les Amis des Instituts Pasteur à Bruxelles, asbl », and the University of Brussels (Action de Recherche Concertée ULB grant) related to her work on HIV latency. The laboratory of CVL is part of the ULB-Cancer Research Center (U-CRC). RV was funded by an 'Aspirant' fellowship (F.R.S-FNRS), a fellowship from 'Les Amis des Instituts Pasteur à Bruxelles, asbl' and is a Belgian American Educational Foundation (BAEF) fellow and a scientific collaborator of the ULB. LN is supported by a 'PDR' grant from the F.R.S-FNRS. GD is a postdoctoral clinical master specialist for the F.R.S-FNRS. A A-A is a fellow of the 'Wallonie-Bruxelles International' Program and the Marie Skłodowska Curie COFUND action. EP is a fellow of the 'Télévie Program' (F.R.S-FNRS). MBD and MS are funded by FRIA fellowships (F.R.S.-FNRS). CVL is 'Directeur de Recherches' at the F.R.S-FNRS. Work in OR's laboratory was supported by grants from the French Agency for Research on AIDS and Viral Hepatitis (ANRS), the Sidaction and the 'Alsace contre le Cancer' Foundation. This work was supported by the European Union's Horizon 2020 research and innovation program under grant agreement No 691119-EU4HIVCURE-H2020-MSCA-RISE-2015. This work is supported by 1UM1AI164562-01, co-funded by National Heart, Lung and Blood Institute, National Institute of Diabetes and Digestive and Kidney Diseases, National Institute of Neurological Disorders and Stroke, National Institute on Drug Abuse and the National Institute of Allergy and Infectious, We thank the members of the ANRS (French National Agency for Research on AIDS and Viral Hepatitis) RHIVIERA (Remission of HIV Era) Consortium for helpful discussions. We thank the HIV-1+ individuals for their willingness to participate in this study. We thank the nursing team of CHU Saint-Pierre Hospital (Elodie Goudeseune, Joëlle Cailleau and Annick Caestecker) who cared for the HIV+ individuals. We thank Jacqueline Pineau from the transfusion centre of Charleroi (Belgium) for providing blood from healthy donors. We thank Hilde Vereertbrugghen from Francis Corazza's laboratory for excellent technical assistance. We thank Motoko Unoki for her precious advice regarding UHRF1 RNA interference. We thank Mitia Duerinckx and Benoît Van Driessche for their support in the probabilistic and statistical analyses., European Project: 691119,H2020,H2020-MSCA-RISE-2015,EU4HIVCURE(2016), Saez-Cirion, Asier, Accelerating HIV Cure in Europe - EU4HIVCURE - - H20202016-02-01 - 2020-01-31 - 691119 - VALID, Medical Microbiology and Infection Prevention, AII - Infectious diseases, Service of Molecular Virology, Department of Molecular Biology (DBM), Université Libre de Bruxelles (ULB), Gosselies 6041, Belgium, Department of Medical Microbiology, Amsterdam UMC, University of Amsterdam, Laboratory of Experimental Virology, Amsterdam 1105 AZ, the Netherland, Infectious Diseases Department, Liège University Hospital, Liège 4000, Belgium, Centre for Research in Infectious Diseases, University College Dublin, Dublin 4, Ireland, Service des Maladies Infectieuses, CHU St-Pierre, Université Libre de Bruxelles (ULB), Brussels 1000, Belgium, Laboratory of Immunology, IRISLab, CHU Brugmann, Université Libre de Bruxelles (ULB), Brussels 1020, Belgium, Dynamique des interactions hôte pathogène (DIHP), Laboratory of Cancer Epigenetics, Faculty of Medicine, ULB-Cancer Research Center (U-CRC), Université Libre de Bruxelles (ULB), Brussels 1070, Belgium, and AP-HP, Hôpital Necker-Enfants-Malades, Service de Microbiologie clinique, Paris 75015, France

Subjects
Ubiquitin-Protein Ligases, Clinical Sciences, HIV Infections, Epigenetic Repression, Decitabine, General Biochemistry, Genetics and Molecular Biology, Genetics, Humans, 2.2 Factors relating to the physical environment, 2.1 Biological and endogenous factors, Aetiology, UHRF1, [SDV.MP.VIR] Life Sciences [q-bio]/Microbiology and Parasitology/Virology, Acquired Immunodeficiency Syndrome, [SDV.BBM.BM]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology, General Medicine, DNA Methylation, Reactivation, Virus Latency, Infectious Diseases, Good Health and Well Being, [SDV.MP.VIR]Life Sciences [q-bio]/Microbiology and Parasitology/Virology, HIV-1, CCAAT-Enhancer-Binding Proteins, Public Health and Health Services, HIV/AIDS, Epigenetics, HIV-1 latency, Infection, EGCG
Abstract

BackgroundThe multiplicity, heterogeneity, and dynamic nature of human immunodeficiency virus type-1 (HIV-1) latency mechanisms are reflected in the current lack of functional cure for HIV-1. Accordingly, all classes of latency-reversing agents (LRAs) have been reported to present variable ex vivo potencies. Here, we investigated the molecular mechanisms underlying the potency variability of one LRA: the DNA methylation inhibitor 5-aza-2'-deoxycytidine (5-AzadC).MethodsWe employed epigenetic interrogation methods (electrophoretic mobility shift assays, chromatin immunoprecipitation, Infinium array) in complementary HIV-1 infection models (latently-infected T-cell line models, primary CD4+ T-cell models and ex vivo cultures of PBMCs from HIV+ individuals). Extracellular staining of cell surface receptors and intracellular metabolic activity were measured in drug-treated cells. HIV-1 expression in reactivation studies was explored by combining the measures of capsid p24Gag protein, green fluorescence protein signal, intracellular and extracellular viral RNA and viral DNA.FindingsWe uncovered specific demethylation CpG signatures induced by 5-AzadC in the HIV-1 promoter. By analyzing the binding modalities to these CpG, we revealed the recruitment of the epigenetic integrator Ubiquitin-like with PHD and RING finger domain 1 (UHRF1) to the HIV-1 promoter. We showed that UHRF1 redundantly binds to the HIV-1 promoter with different binding modalities where DNA methylation was either non-essential, essential or enhancing UHRF1 binding. We further demonstrated the role of UHRF1 in the epigenetic repression of the latent viral promoter by a concerted control of DNA and histone methylations.InterpretationA better understanding of the molecular mechanisms of HIV-1 latency allows for the development of innovative antiviral strategies. As a proof-of-concept, we showed that pharmacological inhibition of UHRF1 in ex vivo HIV+ patient cell cultures resulted in potent viral reactivation from latency. Together, we identify UHRF1 as a novel actor in HIV-1 epigenetic silencing and highlight that it constitutes a new molecular target for HIV-1 cure strategies.FundingFunding was provided by the Belgian National Fund for Scientific Research (F.R.S.-FNRS, Belgium), the «Fondation Roi Baudouin», the NEAT (European AIDS Treatment Network) program, the Internationale Brachet Stiftung, ViiV Healthcare, the Télévie, the Walloon Region («Fonds de Maturation»), «Les Amis des Instituts Pasteur à Bruxelles, asbl», the University of Brussels (Action de Recherche Concertée ULB grant), the Marie Skodowska Curie COFUND action, the European Union's Horizon 2020 research and innovation program under grant agreement No 691119-EU4HIVCURE-H2020-MSCA-RISE-2015, the French Agency for Research on AIDS and Viral Hepatitis (ANRS), the Sidaction and the "Alsace contre le Cancer" Foundation. This work is supported by 1UM1AI164562-01, co-funded by National Heart, Lung and Blood Institute, National Institute of Diabetes and Digestive and Kidney Diseases, National Institute of Neurological Disorders and Stroke, National Institute on Drug Abuse and the National Institute of Allergy and Infectious Diseases.

Published
2022

3. Optimal maturation of the SIV-specific CD8 + T-cell response after primary infection is associated with natural control of SIV. ANRS SIC study

Author

Roger Le Grand, Jeremie Guedj, Nathalie Bosquet, Antoine Blancher, David Price, Annie David, Bruno Vaslin, Gianfranco Pancino, Emma Gostick, Naya Sylla, Olivier Lambotte, Caroline Passaes, Pierre Versmisse, Véronique Avettand-Fenoel, Vincent Madelain, Valérie Monceaux, Asier Sáez-Cirión, Christine Rouzioux, Antoine Millet, Michaela Müller-Trutwin, HIV, Inflammation et persistance - HIV, Inflammation and Persistence, Institut Pasteur [Paris] (IP), Infectious Diseases Models for Innovative Therapies (IDMIT), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Paris-Saclay, UFR Médecine [Santé] - Université Paris Cité (UFR Médecine UPCité), Université Paris Cité (UPCité), Infection, Anti-microbiens, Modélisation, Evolution (IAME (UMR_S_1137 / U1137)), Université Paris 13 (UP13)-Université Paris Diderot - Paris 7 (UPD7)-Université Sorbonne Paris Cité (USPC)-Institut National de la Santé et de la Recherche Médicale (INSERM), Cardiff University, Centre Hospitalier Universitaire de Toulouse (CHU Toulouse), Centre de Physiopathologie Toulouse Purpan (CPTP), Université Toulouse III - Paul Sabatier (UT3), Université de Toulouse (UT)-Université de Toulouse (UT)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Service de médecine interne et maladies infectieuses, Université Paris-Sud - Paris 11 (UP11)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpital Bicêtre, Service de Médecine Interne - Immunologie Clinique [AP-HP Bicêtre], AP-HP Hôpital Bicêtre (Le Kremlin-Bicêtre), Infections à Vih, Réservoirs, Pharmacologie des Antirétroviraux et Prévention de la Transmission Mère Enfant, Université Paris Descartes - Paris 5 (UPD5), Service de Microbiologie Clinique [Paris], CHU Necker - Enfants Malades [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), Université Paris Cité - UFR Médecine [Santé] (UPCité UFR Médecine), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Passaes, Caroline, HIV, Inflammation et persistance, Institut Pasteur [Paris], Université de Paris - UFR Médecine Paris Centre [Santé] (UP Médecine Paris Centre), Université de Paris (UP), and CHU Toulouse [Toulouse]

Subjects
0303 health sciences, [SDV.IMM] Life Sciences [q-bio]/Immunology, Period (gene), animal diseases, Memory pool, virus diseases, Viremia, Biology, medicine.disease, Phenotype, Macaque, 3. Good health, 03 medical and health sciences, 0302 clinical medicine, biology.animal, Immunology, medicine, Potency, Cytotoxic T cell, [SDV.IMM]Life Sciences [q-bio]/Immunology, CD8, 030304 developmental biology, 030215 immunology
Abstract

Highly efficient virus-specific CD8+ T-cells are associated with immune control of HIV infection, but it remains unclear how these cells are generated and maintained over time. We used a macaque model of spontaneous control of SIVmac251 infection to monitor the development and evolution of potent antiviral CD8+ T-cell responses. SIV-specific CD8+ T-cells emerged during primary infection in all animals. However, the ability of CD8+ T cells to suppress SIV replication was low in early stages but increased after a period of maturation, temporally linked with the establishment of sustained low-level viremia in controller macaques. SIV-specific CD8+ T-cells with a central memory phenotype expressed higher levels of survival markers in controllers versus non-controllers. In contrast, a persistently skewed differentiation phenotype was observed among central memory SIV-specific CD8+ T-cells in non-controllers since primary infection, typified by relatively high expression levels of T-bet.Collectively, these data show that the phenotype of SIV-specific CD8+ T-cells defined early after SIV infection favor the gain of antiviral potency as a function of time in controllers, whereas SIV-specific CD8+ T-cell responses in non-controllers fail to gain antiviral potency due to early defects imprinted in the central memory pool.

Published
2020

4. An ADAM-10 dependent EPCR shedding links meningococcal interaction with endothelial cells to purpura fulminans

Author

Sandrine Bourdoulous, Hervé Lécuyer, Jean-Philippe Barnier, François Saller, Xavier Nassif, Soraya Matczak, Elsa P. Bianchini, Mathieu Coureuil, Zoé Virion, Delphine Borgel, Bodescot, Myriam, Recherche transnationale sur les maladies infectieuses humaines - Bacterial interaction with the microvasculature, a target for therapeutic intervention during septicaemia - - BactInfectERA2014 - ANR-14-IFEC-0006 - IFEC - VALID, Institut Necker Enfants-Malades (INEM - UM 111 (UMR 8253 / U1151)), 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 Paris Descartes - Paris 5 (UPD5), Service de Microbiologie Clinique [Paris], CHU Necker - Enfants Malades [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), Institut Cochin (IC UM3 (UMR 8104 / U1016)), Hémostase, Inflammation, Thrombose (HITH - U1176 Inserm - CHU Bicêtre), Université Paris-Sud - Paris 11 (UP11)-Institut National de la Santé et de la Recherche Médicale (INSERM)-AP-HP Hôpital Bicêtre (Le Kremlin-Bicêtre), Service d'Hématologie Biologique [Paris], This work was supported by an Agence Nationale de la Recherche grant (ANR-14-IFEC-0006-01 call ERANET INFECT-ERA 2014 - grant recipient SB), the grant program EMERGENCE from La Mairie de Paris (grant recipient MC) and a grant from the Meningitis Research Foundation (MRF 1602 - grant recipient XN). The laboratory of XN is supported by INSERM, CNRS, Université Paris Descartes, and the Fondation pour la Recherche Médicale (FRM DEQ20140329533)., ANR-14-IFEC-0006,BactInfectERA,Bacterial interaction with the microvasculature, a target for therapeutic intervention during septicaemia(2014), and Université Paris-Sud - Paris 11 (UP11)-AP-HP Hôpital Bicêtre (Le Kremlin-Bicêtre)-Institut National de la Santé et de la Recherche Médicale (INSERM)

Subjects
Bacterial Diseases, 0301 basic medicine, Physiology, ADAM10, Meningococcal Disease, Molting, Cardiovascular Medicine, Neisseria meningitidis, Pathology and Laboratory Medicine, medicine.disease_cause, Biochemistry, Vascular Medicine, Epithelium, Bacterial Adhesion, ADAM10 Protein, Animal Cells, [SDV.MHEP.MI]Life Sciences [q-bio]/Human health and pathology/Infectious diseases, Medicine and Health Sciences, Small interfering RNAs, Biology (General), Cells, Cultured, Endothelial protein C receptor, Thrombin, Endothelial Protein C Receptor, Hematology, Bacterial Pathogens, 3. Good health, [SDV.MHEP.CSC] Life Sciences [q-bio]/Human health and pathology/Cardiology and cardiovascular system, Nucleic acids, Endothelial stem cell, Infectious Diseases, medicine.anatomical_structure, Medical Microbiology, Cardiovascular Diseases, Purpura Fulminans, [SDV.MHEP.MI] Life Sciences [q-bio]/Human health and pathology/Infectious diseases, Cellular Types, Anatomy, Pathogens, Neisseria, Research Article, medicine.drug, Purpura fulminans, Endothelium, QH301-705.5, Immune Cells, Immunology, Antigen-Presenting Cells, Meningococcal disease, Microbiology, 03 medical and health sciences, [SDV.MHEP.CSC]Life Sciences [q-bio]/Human health and pathology/Cardiology and cardiovascular system, Virology, Genetics, medicine, Humans, Non-coding RNA, Microbial Pathogens, Blood Coagulation, Molecular Biology, Bacteria, Coagulation Disorders, business.industry, Organisms, Biology and Life Sciences, Endothelial Cells, Proteins, Membrane Proteins, Epithelial Cells, Thrombosis, Cell Biology, RC581-607, medicine.disease, Gene regulation, Meningococcal Infections, Biological Tissue, 030104 developmental biology, Microvessels, RNA, Parasitology, Gene expression, Endothelium, Vascular, Amyloid Precursor Protein Secretases, Immunologic diseases. Allergy, Physiological Processes, business, Protein C
Abstract

Purpura fulminans is a deadly complication of Neisseria meningitidis infections due to extensive thrombosis of microvessels. Although a Disseminated Intra-vascular Coagulation syndrome (DIC) is frequently observed during Gram negative sepsis, it is rarely associated with extensive thrombosis like those observed during meningococcemia, suggesting that the meningococcus induces a specific dysregulation of coagulation. Another specific feature of N. meningitidis pathogenesis is its ability to colonize microvessels endothelial cells via type IV pili. Importantly, endothelial cells are key in controlling the coagulation cascade through the activation of the potent anticoagulant Protein C (PC) thanks to two endothelial cell receptors among which the Endothelial Protein C Receptor (EPCR). Considering that congenital or acquired deficiencies of PC are associated with purpura fulminans, we hypothesized that a defect in the activation of PC following meningococcal adhesion to microvessels is responsible for the thrombotic events observed during meningococcemia. Here we showed that the adhesion of N. meningitidis on endothelial cells results in a rapid and intense decrease of EPCR expression by inducing its cleavage in a process know as shedding. Using siRNA experiments and CRISPR/Cas9 genome edition we identified ADAM10 (A Disintegrin And Metalloproteinase-10) as the protease responsible for this shedding. Surprisingly, ADAM17, the only EPCR sheddase described so far, was not involved in this process. Finally, we showed that this ADAM10-mediated shedding of EPCR induced by the meningococcal interaction with endothelial cells was responsible for an impaired activation of Protein C. This work unveils for the first time a direct link between meningococcal adhesion to endothelial cells and a severe dysregulation of coagulation, and potentially identifies new therapeutic targets for meningococcal purpura fulminans., Author summary Neisseria meningitidis (meningococcus) is responsible for a severe syndrome called purpura fulminans in which the coagulation system is totally dysregulated, leading to an extensive occlusion of blood microvessels. The pathogenesis of this syndrome is still not understood. Here we show that the meningococcus, when adhering on the apical surface of endothelial cells, induces the activation of membranous protease named ADAM-10, which in turn hydrolyses a cellular receptor called EPCR. The latter is key for the activation of a circulating potent anticoagulant, the Protein C (PC). PC activation is then impaired following meningococcal adhesion on endothelial cells. This work unveils for the first time a specific dysregulation of coagulation induced by the meningococcus and potentially identifies new therapeutic targets for meningococcal purpura fulminans.

Published
2018

5. Quelles mesures pour maîtriser le risque infectieux chez les patients immunodéprimés ? Recommandations formalisées d’experts

Author

Zahar, R., Jolivet, S., Adam, H., Dananché, C., Lizon, J., Alfandari, S., Boulestreau, H., Baghdadi, N., Bay, J.-O., Bénéteau, M., Bougnoux, M., Brenier-Pinchart, P., Dalle, J.-H., Fournier, Sébastien, Fuzibet, J.-G., Kauffmann-Lacroix, C., Le Guinche, I., Lepelletier, D., Loukili, N., Lory, A., Morvan, M., Oumedaly, R., Ribaud, P., Rohrlich, P., Vanhems, Philippe, Aho, S., Vanjak, D., Gangneux, J.-P., Infection, Anti-microbiens, Modélisation, Evolution (IAME (UMR_S_1137 / U1137)), Université Paris 13 (UP13)-Université Paris Diderot - Paris 7 (UPD7)-Université Sorbonne Paris Cité (USPC)-Institut National de la Santé et de la Recherche Médicale (INSERM), CIC - CHU Bichat, Institut National de la Santé et de la Recherche Médicale (INSERM), Service d'épidémiologie et d'hygiène hospitalières (CHU de Dijon), Centre Hospitalier Universitaire de Dijon - Hôpital François Mitterrand (CHU Dijon), Laboratoire des pathogènes émergents -- Emerging Pathogens Laboratory (LPE-Fondation Mérieux), Centre International de Recherche en Infectiologie - UMR (CIRI), École normale supérieure - Lyon (ENS Lyon)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-École normale supérieure - Lyon (ENS Lyon)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Centre Hospitalier Régional Universitaire de Nancy (CHRU Nancy), Service de Réanimation polyvalente et des Maladies infectieuses, Centre Hospitalier Gustave Dron, Service d'Hygiène Hospitalière, CHU Bordeaux [Bordeaux]-Groupe hospitalier Pellegrin, Centre Hospitalier Régional Universitaire [Lille] (CHRU Lille), CHU Clermont-Ferrand, Institut Curie [Saint-Cloud], Biologie et Pathogénicité fongiques, Institut Pasteur [Paris]-Institut National de la Recherche Agronomique (INRA), Service de Microbiologie Clinique [Paris], CHU Necker - Enfants Malades [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), Institute for Advanced Biosciences / Institut pour l'Avancée des Biosciences (Grenoble) (IAB), Centre Hospitalier Universitaire [Grenoble] (CHU)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Etablissement français du sang - Auvergne-Rhône-Alpes (EFS)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019]), Hôpital Robert Debré Paris, Hôpital Robert Debré, Centre Hospitalier Universitaire de Nice (CHU Nice), Centre hospitalier universitaire de Poitiers (CHU Poitiers), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), Service de bactériologie et hygiène hospitalière [Nantes], Université de Nantes (UN)-Centre hospitalier universitaire de Nantes (CHU Nantes), Hôpital Sainte-Marguerite [CHU - APHM] (Hôpitaux Sud ), Centre Hospitalier Régional Universitaire [Montpellier] (CHRU Montpellier), Institut d'Hématologie de Basse-Normandie (IHBN), Université de Caen Normandie (UNICAEN), Normandie Université (NU)-Normandie Université (NU)-CHU Caen, Normandie Université (NU)-Tumorothèque de Caen Basse-Normandie (TCBN)-Tumorothèque de Caen Basse-Normandie (TCBN)-Centre Régional de Lutte contre le Cancer François Baclesse [Caen] (UNICANCER/CRLC), UNICANCER-Tumorothèque de Caen Basse-Normandie (TCBN)-Normandie Université (NU)-UNICANCER, Service greffe de moelle osseuse, Université Paris Diderot - Paris 7 (UPD7)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Groupe Hospitalier Saint Louis - Lariboisière - Fernand Widal [Paris], Centre méditerranéen de médecine moléculaire (C3M), 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)-Institut National de la Santé et de la Recherche Médicale (INSERM), Institut Curie [Paris], Service de Parasitologie-Mycologie [Rennes], Université de Rennes 1 (UR1), Université de Rennes (UNIV-RENNES)-Université de Rennes (UNIV-RENNES)-Hôpital Pontchaillou-CHU Pontchaillou [Rennes], Centre International de Recherche en Infectiologie (CIRI), École normale supérieure de Lyon (ENS de Lyon)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Université Jean Monnet - Saint-Étienne (UJM)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-École normale supérieure de Lyon (ENS de Lyon)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Université Jean Monnet - Saint-Étienne (UJM)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Institut Curie - Saint Cloud (ICSC), Biologie et Pathogénicité fongiques (BPF), Institut National de la Recherche Agronomique (INRA)-Institut Pasteur [Paris] (IP), Normandie Université (NU)-UNICANCER-Tumorothèque de Caen Basse-Normandie (TCBN)-UNICANCER, Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Université Paris Diderot - Paris 7 (UPD7)-Groupe Hospitalier Saint Louis - Lariboisière - Fernand Widal [Paris], 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)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Côte d'Azur (UCA), Université de Rennes (UR)-Hôpital Pontchaillou-CHU Pontchaillou [Rennes], Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Paris 13 (UP13)-Université Paris Diderot - Paris 7 (UPD7)-Université Sorbonne Paris Cité (USPC), Service d'Epidémiologie et Hygiène Hospitalière, Institut National de la Santé et de la Recherche Médicale (INSERM)-École normale supérieure - Lyon (ENS Lyon)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-École normale supérieure - Lyon (ENS Lyon)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS), Institut National de la Recherche Agronomique (INRA)-Institut Pasteur [Paris], Centre Hospitalier Universitaire [Grenoble] (CHU)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Etablissement français du sang - Auvergne-Rhône-Alpes (EFS)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes (UGA), Normandie Université (NU)-Tumorothèque de Caen Basse-Normandie (TCBN)-Tumorothèque de Caen Basse-Normandie (TCBN)-Centre Régional de Lutte contre le Cancer François Baclesse (CRLC François Baclesse ), Normandie Université (NU)-Tumorothèque de Caen Basse-Normandie (TCBN), Assistance publique - Hôpitaux de Paris (AP-HP) (APHP)-Université Paris Diderot - Paris 7 (UPD7)-Groupe Hospitalier Saint Louis - Lariboisière - Fernand Widal [Paris], Assistance publique - Hôpitaux de Paris (AP-HP) (APHP), Université Côte d'Azur (UCA)-Université Côte d'Azur (UCA)-Institut National de la Santé et de la Recherche Médicale (INSERM), and Institut Curie

Subjects
Immunocompromised Host, Infection Control, Risk Factors, Practice Guidelines as Topic, Air Microbiology, Humans, [SDV.MHEP.HEM]Life Sciences [q-bio]/Human health and pathology/Hematology, Disease Susceptibility, France, Infections
Abstract

International audience; The increase use of immunosuppressive treatments in patients with solid cancer and/or inflammatory diseases requires revisiting our practices for the prevention of infectious risk in the care setting. A review of the literature by a multidisciplinary working group at the beginning of 2014 wished to answer the following 4 questions to improve healthcare immunocompromised patients: (I) How can we define immunocompromised patients with high, intermediate and low infectious risk, (II) which air treatment should be recommended for this specific population? (III) What additional precautions should be recommended for immunocompromised patients at risk for infection? (IV) Which global environmental control should be recommended? Based on data from the literature and using the GRADE method, we propose 15 recommendations that could help to reduce the risk of infection in these exposed populations.; L’intensification des traitements immunosuppresseurs et la généralisation de leur utilisation aux patients atteints de cancer solide et/ou de maladies inflammatoires nécessitent de revisiter nos pratiques de prévention du risque infectieux en milieu de soin. Cette révision est justifiée aussi par les modifications de nos pratiques avec une intensification des prises en charge en structures de soins de suites réhabilitation et en ambulatoire. Une revue de la littérature menée par un groupe de travail multidisciplinaire réuni début 2014 a souhaité répondre aux 4 questions suivantes : (i) Quelle définition des patients immunodéprimés à risque infectieux élevé, intermédiaire et faible, (ii) Quel traitement d’air recommander pour les patients immunodéprimés à risque d’infections ? (iii) Quelles précautions complémentaires recommander pour les patients immunodéprimés à risque d’infections ? (iv) Quelle maîtrise globale de l’environnement recommander pour les patients immunodéprimés à risque d’infections ? À partir des données de la littérature et en utilisant la méthode GRADE, nous proposons 15 recommandations qui permettront de réduire le risque infectieux dans ces populations exposées.

Published
2017

6. Comparative In Vitro Activities of Caspofungin and Micafungin, Determined Using the Method of the European Committee on Antimicrobial Susceptibility Testing, against Yeast Isolates Obtained in France in 2005-2006

Author

Dorothée Raoux, Damien Hoinard, D. Moissenet, G. Galeazzi, I. Poilane, C. Lawrence, Françoise Dromer, Marie-Elisabeth Bougnoux, Olivier Lortholary, Eric Dannaoui, Unité de Parasitologie-Mycologie [CHU HEGP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpital Européen Georges Pompidou [APHP] (HEGP), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO)-Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO), Centre d'infectiologie Necker-Pasteur [CHU Necker], Institut Pasteur [Paris] (IP)-CHU Necker - Enfants Malades [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), Biologie et Pathogénicité fongiques (BPF), Institut National de la Recherche Agronomique (INRA)-Institut Pasteur [Paris] (IP), Service de Microbiologie Clinique [Paris], CHU Necker - Enfants Malades [AP-HP], Hôpital Louis Mourier - AP-HP [Colombes], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), Laboratoire de microbiologie, Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpital Raymond Poincaré [AP-HP], CHU Trousseau [APHP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU), Université Pierre et Marie Curie - Paris 6 - UFR de Médecine Pierre et Marie Curie (UPMC), Université Pierre et Marie Curie - Paris 6 (UPMC), Centre National de Référence des Mycoses invasives et antifongiques - Mycologie moléculaire (CNRMA), Institut Pasteur [Paris] (IP)-Centre National de la Recherche Scientifique (CNRS), and CHU Bordeaux [Bordeaux]

Subjects
Antifungal Agents, Lipoproteins, [SDV]Life Sciences [q-bio], Microbial Sensitivity Tests, Candida parapsilosis, Microbiology, Candida tropicalis, Echinocandins, Lipopeptides, chemistry.chemical_compound, Caspofungin, Drug Resistance, Fungal, Yeasts, Candida krusei, polycyclic compounds, medicine, Humans, Pharmacology (medical), Candida albicans, Candida, Antibacterial agent, Pharmacology, biology, Candida glabrata, Micafungin, biochemical phenomena, metabolism, and nutrition, bacterial infections and mycoses, biology.organism_classification, stomatognathic diseases, Infectious Diseases, Mycoses, chemistry, Susceptibility, lipids (amino acids, peptides, and proteins), France, medicine.drug
Abstract

The in vitro activities of caspofungin and micafungin against 1,038 yeast isolates have been determined. The caspofungin and micafungin MICs were lower for Candida albicans , Candida glabrata , and Candida tropicalis than for Candida parapsilosis , Candida guilliermondii , and Candida krusei . A clear correlation was seen between the MICs for the two drugs.

Published
2008

7. Metagenomics and metabolomics approaches in the study of Candida albicans colonization of host niches: a framework for finding microbiome-based antifungal strategies.

Author

Delavy M, Sertour N, d'Enfert C, and Bougnoux ME

Subjects
Female, Humans, Candida albicans genetics, Antifungal Agents pharmacology, Metabolomics, Bacteria, Gastrointestinal Microbiome, Microbiota
Abstract

In silico and experimental approaches have allowed an ever-growing understanding of the interactions within the microbiota. For instance, recently acquired data have increased knowledge of the mechanisms that support, in the gut and vaginal microbiota, the resistance to colonization by Candida albicans, an opportunistic fungal pathogen whose overgrowth can initiate severe infections in immunocompromised patients. Here, we review how bacteria from the microbiota interact with C. albicans. We show how recent OMICs-based pipelines, using metagenomics and/or metabolomics, have identified bacterial species and metabolites modulating C. albicans growth. We finally discuss how the combined use of cutting-edge OMICs-based and experimental approaches could provide new means to control C. albicans overgrowth within the microbiota and prevent its consequences., Competing Interests: Declaration of interests No interests are declared., (Copyright © 2023 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Published
2023
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8. Evaluation of ceftolozane-tazobactam susceptibility on a French nationwide collection of Enterobacterales.

Author

Jousset AB, Bernabeu S, Emeraud C, Bonnin RA, Lomont A, Zahar JR, Merens A, Isnard C, Soismier N, Farfour E, Fihman V, Yin N, Barraud O, Jacquier H, Ranc AG, Laurent F, Corvec S, d'Epenoux LR, Bille E, Degand N, Plouzeau C, Guillard T, Cattoir V, Mizrahi A, Grillon A, Janvier F, Brun CL, Amara M, Bastide M, Lemonnier A, and Dortet L

Subjects
Humans, Prospective Studies, Enterobacteriaceae genetics, Pseudomonas aeruginosa, Cephalosporins pharmacology, Cephalosporins therapeutic use, Tazobactam pharmacology, Tazobactam therapeutic use, Escherichia coli, beta-Lactamases genetics, Anti-Bacterial Agents pharmacology, Anti-Bacterial Agents therapeutic use, Pseudomonas Infections drug therapy
Abstract

Objectives: Ceftolozane-tazobactam (C/T) proved its efficacy for the treatment of infections caused by non-carbapenemase producing Pseudomonas aeruginosa and Enterobacterales. Here, we aimed to provide susceptibility data on a large series of Enterobacterales since the revision of EUCAST categorization breakpoints in 2020., Methods: First, C/T susceptibility was determined on characterized Enterobacterales resistant to third generation cephalosporins (3GCs) (extended spectrum β-lactamase [ESBL] production or different levels of AmpC overexpression) (n = 213) and carbapenem-resistant Enterobacterales (CRE) (n = 259), including 170 carbapenemase producers (CPE). Then, 1632 consecutive clinical Enterobacterales responsible for infection were prospectively collected in 23 French hospitals. C/T susceptibility was determined by E-test® (biomérieux) and broth microdilution (BMD) (Sensititre™, Thermo Scientific) to perform method comparison., Results: Within the collection isolates, 88% of 3GC resistant strains were susceptible to C/T, with important variation depending on the resistance mechanism: 93% vs. 13% susceptibility for CTX-M and SHV-ESBL producers, respectively. Only 20% of the CRE were susceptible to C/T. Among CPE, 80% of OXA-48-like producers were susceptible to C/T, whereas all metallo-β-lactamase producers were resistant. The prospective study revealed that 95.6% of clinical isolates were susceptible to C/T. Method comparison performed on these 1632 clinical isolates demonstrated 99% of categorization agreement between MIC to C/T determined by E-test® in comparison with the BMD (reference) and only 74% of essential agreement., Conclusion: Overall, C/T showed good activity against wild-type Enterobacterales, AmpC producers, and ESBL-producing Escherichia coli but is less active against ESBL-producing Klebsiella pneumoniae, and CRE. E-test® led to an underestimation of the MICs in comparison to the BMD reference., (Copyright © 2023 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Published
2023
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9. Novel role of UHRF1 in the epigenetic repression of the latent HIV-1.

Author

Verdikt R, Bendoumou M, Bouchat S, Nestola L, Pasternak AO, Darcis G, Avettand-Fenoel V, Vanhulle C, Aït-Ammar A, Santangelo M, Plant E, Douce VL, Delacourt N, Cicilionytė A, Necsoi C, Corazza F, Passaes CPB, Schwartz C, Bizet M, Fuks F, Sáez-Cirión A, Rouzioux C, De Wit S, Berkhout B, Gautier V, Rohr O, and Van Lint C

Subjects
Acquired Immunodeficiency Syndrome, DNA Methylation, Decitabine metabolism, Humans, CCAAT-Enhancer-Binding Proteins genetics, CCAAT-Enhancer-Binding Proteins metabolism, Epigenetic Repression, HIV Infections genetics, HIV-1 physiology, Ubiquitin-Protein Ligases genetics, Ubiquitin-Protein Ligases metabolism, Virus Latency genetics
Abstract

Background: The multiplicity, heterogeneity, and dynamic nature of human immunodeficiency virus type-1 (HIV-1) latency mechanisms are reflected in the current lack of functional cure for HIV-1. Accordingly, all classes of latency-reversing agents (LRAs) have been reported to present variable ex vivo potencies. Here, we investigated the molecular mechanisms underlying the potency variability of one LRA: the DNA methylation inhibitor 5-aza-2'-deoxycytidine (5-AzadC)., Methods: We employed epigenetic interrogation methods (electrophoretic mobility shift assays, chromatin immunoprecipitation, Infinium array) in complementary HIV-1 infection models (latently-infected T-cell line models, primary CD4 + T-cell models and ex vivo cultures of PBMCs from HIV + individuals). Extracellular staining of cell surface receptors and intracellular metabolic activity were measured in drug-treated cells. HIV-1 expression in reactivation studies was explored by combining the measures of capsid p24 Gag protein, green fluorescence protein signal, intracellular and extracellular viral RNA and viral DNA., Findings: We uncovered specific demethylation CpG signatures induced by 5-AzadC in the HIV-1 promoter. By analyzing the binding modalities to these CpG, we revealed the recruitment of the epigenetic integrator Ubiquitin-like with PHD and RING finger domain 1 (UHRF1) to the HIV-1 promoter. We showed that UHRF1 redundantly binds to the HIV-1 promoter with different binding modalities where DNA methylation was either non-essential, essential or enhancing UHRF1 binding. We further demonstrated the role of UHRF1 in the epigenetic repression of the latent viral promoter by a concerted control of DNA and histone methylations., Interpretation: A better understanding of the molecular mechanisms of HIV-1 latency allows for the development of innovative antiviral strategies. As a proof-of-concept, we showed that pharmacological inhibition of UHRF1 in ex vivo HIV + patient cell cultures resulted in potent viral reactivation from latency. Together, we identify UHRF1 as a novel actor in HIV-1 epigenetic silencing and highlight that it constitutes a new molecular target for HIV-1 cure strategies., Funding: Funding was provided by the Belgian National Fund for Scientific Research (F.R.S.-FNRS, Belgium), the « Fondation Roi Baudouin », the NEAT (European AIDS Treatment Network) program, the Internationale Brachet Stiftung, ViiV Healthcare, the Télévie, the Walloon Region (« Fonds de Maturation »), « Les Amis des Instituts Pasteur à Bruxelles, asbl », the University of Brussels (Action de Recherche Concertée ULB grant), the Marie Skodowska Curie COFUND action, the European Union's Horizon 2020 research and innovation program under grant agreement No 691119-EU4HIVCURE-H2020-MSCA-RISE-2015, the French Agency for Research on AIDS and Viral Hepatitis (ANRS), the Sidaction and the "Alsace contre le Cancer" Foundation. This work is supported by 1UM1AI164562-01, co-funded by National Heart, Lung and Blood Institute, National Institute of Diabetes and Digestive and Kidney Diseases, National Institute of Neurological Disorders and Stroke, National Institute on Drug Abuse and the National Institute of Allergy and Infectious Diseases., Competing Interests: Declaration of interests The authors declare no conflicts of interest., (Copyright © 2022 The Authors. Published by Elsevier B.V. All rights reserved.)

Published
2022
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10. A rapid, automatic and accurate assay for quantifying temocillin in human serum and CSF using turbulent flow liquid chromatography coupled to high-resolution mass spectrometry. Clinical application.

Author

Petitjean G, Aman-Pommier F, Coudoré F, El Helali N, and Le Monnier A

Subjects
Anti-Bacterial Agents blood, Anti-Bacterial Agents cerebrospinal fluid, Anti-Bacterial Agents pharmacology, Humans, Limit of Detection, Linear Models, Microbial Sensitivity Tests, Penicillins pharmacology, Pseudomonas aeruginosa drug effects, Reproducibility of Results, Chromatography, Liquid methods, Penicillins blood, Penicillins cerebrospinal fluid, Spectrometry, Mass, Electrospray Ionization methods
Abstract

Temocillin is a β-lactamase-resistant penicillin used for the treatment of multiple drug-resistant Gram-negative bacteria. To maximize efficacy and avoid adverse effects, the dose regimen has to be quickly adjusted to the clinical situations. This necessitates the development of a rapid, reliable and accurate analytical method. Temocillin and the stable isotopically labeled internal standard ([ 13 C 6 ]-amoxicillin) were extracted from either serum or cerebrospinal fluid by a turbulent flow liquid chromatographic method and eluted onto an octadecyl-silica phase with polar endcapping. Mass spectrometry was conducted using an exact mass determination method by electrospray positive ionization high-resolution mass spectrometry. The LLOQ and ULOQ of the present method were determined to be 0.4 and 200 μg/ml for serum and cerebrospinal fluid samples, respectively. The total analysis time was <7 min. The recovery ranged from 87.7 to 120.8%. Intra- and inter-day precision and trueness were tested at four concentration levels: 0.4, 8, 40 and 160 μg/ml. Values were 6.33 ± 1.53, 8.8 ± 1.3, 8.8 ± 0.36 and 2.1 ± 0.76%, and 5.0 ± 0.54, 9.9 ± 1.0, 5.8 ± 1.6 and 0.1 ± 1.1%, for inter- and intra-day analysis, respectively. Temocillin was found to be stable under all relevant laboratory conditions. The method was cross-validated with a microbiological assay. This method is suitable for accurate measurement of temocillin concentration in small volumes of serum or cerebrospinal fluid. Thanks to the online extraction procedure, the overall analytical time is compatible with high-throughput analysis for clinical application., (© 2019 John Wiley & Sons, Ltd.)

Published
2020
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11. [Meningococcus purpura fulminans, a partially solved mystery].

Author

Lécuyer H, Nassif X, and Coureuil M

Subjects
Blood Coagulation physiology, Diagnosis, Differential, Endothelial Protein C Receptor physiology, Humans, Protein C physiology, Meningitis, Meningococcal complications, Meningitis, Meningococcal diagnosis, Neisseria meningitidis physiology, Purpura Fulminans diagnosis, Purpura Fulminans microbiology
Published
2018
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12. An ADAM-10 dependent EPCR shedding links meningococcal interaction with endothelial cells to purpura fulminans.

Author

Lécuyer H, Virion Z, Barnier JP, Matczak S, Bourdoulous S, Bianchini E, Saller F, Borgel D, Nassif X, and Coureuil M

Subjects
ADAM10 Protein genetics, Amyloid Precursor Protein Secretases genetics, Bacterial Adhesion, Blood Coagulation physiology, Cells, Cultured, Endothelial Protein C Receptor genetics, Endothelium, Vascular metabolism, Endothelium, Vascular microbiology, Humans, Membrane Proteins genetics, Meningococcal Infections microbiology, Microvessels metabolism, Microvessels microbiology, Neisseria meningitidis physiology, Protein C genetics, Purpura Fulminans metabolism, Purpura Fulminans pathology, ADAM10 Protein metabolism, Amyloid Precursor Protein Secretases metabolism, Endothelial Protein C Receptor metabolism, Endothelium, Vascular pathology, Membrane Proteins metabolism, Meningococcal Infections complications, Microvessels pathology, Protein C metabolism, Purpura Fulminans etiology
Abstract

Purpura fulminans is a deadly complication of Neisseria meningitidis infections due to extensive thrombosis of microvessels. Although a Disseminated Intra-vascular Coagulation syndrome (DIC) is frequently observed during Gram negative sepsis, it is rarely associated with extensive thrombosis like those observed during meningococcemia, suggesting that the meningococcus induces a specific dysregulation of coagulation. Another specific feature of N. meningitidis pathogenesis is its ability to colonize microvessels endothelial cells via type IV pili. Importantly, endothelial cells are key in controlling the coagulation cascade through the activation of the potent anticoagulant Protein C (PC) thanks to two endothelial cell receptors among which the Endothelial Protein C Receptor (EPCR). Considering that congenital or acquired deficiencies of PC are associated with purpura fulminans, we hypothesized that a defect in the activation of PC following meningococcal adhesion to microvessels is responsible for the thrombotic events observed during meningococcemia. Here we showed that the adhesion of N. meningitidis on endothelial cells results in a rapid and intense decrease of EPCR expression by inducing its cleavage in a process know as shedding. Using siRNA experiments and CRISPR/Cas9 genome edition we identified ADAM10 (A Disintegrin And Metalloproteinase-10) as the protease responsible for this shedding. Surprisingly, ADAM17, the only EPCR sheddase described so far, was not involved in this process. Finally, we showed that this ADAM10-mediated shedding of EPCR induced by the meningococcal interaction with endothelial cells was responsible for an impaired activation of Protein C. This work unveils for the first time a direct link between meningococcal adhesion to endothelial cells and a severe dysregulation of coagulation, and potentially identifies new therapeutic targets for meningococcal purpura fulminans.

Published
2018
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