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1. Standardized whole blood stimulation improves immunomonitoring of induced immune responses in multi-center study

Author

Alain Savenay, Matthew L. Albert, Delphine Louis, Manfred Schmolz, Nina Salabert-Le Guen, Olivier Lantz, Régis Josien, Darragh Duffy, Françoise Mascart, Antoine Toubert, Marie Noelle Ungeheuer, Vincent Rouilly, Raouf Djebali, Lydia Redjah, José J.G. Ruiz de Morales, Catherine Ottone, Véronique Corbière, Eva Martínez-Cáceres, Cécile Braudeau, Samuel S.T. LaBrie, Immunobiologie des Cellules dendritiques, Institut Pasteur [Paris]-Institut National de la Santé et de la Recherche Médicale (INSERM), Centre de Recherche Translationnelle (CRT), Institut Pasteur [Paris], Laboratoire d’Immunologie, Centre d’Immunomonitorage Nantes Atlantique (CIMNA), Centre hospitalier universitaire de Nantes (CHU Nantes), Centre de Recherche en Transplantation et Immunologie (U1064 Inserm - CRTI), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Nantes - UFR de Médecine et des Techniques Médicales (UFR MEDECINE), Université de Nantes (UN)-Université de Nantes (UN), laboratoire de vaccinologie et immunité mucosale, Université Libre de Bruxelles [Bruxelles] (ULB), Investigation Clinique et d’Accès aux Ressources Biologiques (Plate-forme) - Clinical Investigation and Access to BioResources (ICAReB), Myriad RBM, Immunité et cancer (U932), Université Paris Descartes - Paris 5 (UPD5)-Institut Curie-Institut National de la Santé et de la Recherche Médicale (INSERM), Hospital Universitari Germans Trias I Pujol, Immunobiology Clinic, Hôpital Erasme, Complejo Asistencial Universitario de León, Alloimmunité-Autoimmunité-Transplantation (A2T), Institut Universitaire d'Hématologie (IUH), Université Paris Diderot - Paris 7 (UPD7)-Université Paris Diderot - Paris 7 (UPD7)-Institut National de la Santé et de la Recherche Médicale (INSERM), HOT Screen GmbH, Genentech, Inc. [San Francisco], TruCulture tubes, stimuli, and protein Luminex XMAP analysis were provided by Myriad RBM, Inc (Austin, USA) and TruCulture tubes were manufactured at HOT Screen (Reutlingen, Germany). Donor recruitment costs were covered by each participating FOCIS center of excellence. Author contributions: DD, VR, VC, SL, OL, EMC, RP, MS, AT & MLA designed the study. CB, RD, MNU, DL, FM, JGRM, CO, LR, AS performed experiments. DD, VR, & MLA performed analysis and wrote the paper. All authors reviewed the manuscript., Institut Pasteur [Paris] (IP)-Institut National de la Santé et de la Recherche Médicale (INSERM), Centre de Recherche Translationnelle - Center for Translational Science (CRT), Institut Pasteur [Paris] (IP), Université libre de Bruxelles (ULB), Université Paris Descartes - Paris 5 (UPD5)-Institut Curie [Paris]-Institut National de la Santé et de la Recherche Médicale (INSERM), Hôpital Erasme [Bruxelles] (ULB), Faculté de Médecine [Bruxelles] (ULB), Université libre de Bruxelles (ULB)-Université libre de Bruxelles (ULB)-Faculté de Médecine [Bruxelles] (ULB), Université libre de Bruxelles (ULB)-Université libre de Bruxelles (ULB), Institut Pasteur [Paris]-Institut National de la Santé et de la Recherche Médicale ( INSERM ), Centre de Recherche Translationnelle ( CRT ), Centre Hospitalier Universitaire de Nantes, Centre de Recherche en Transplantation et Immunologie ( U1064 Inserm - CRTI - CHU Nantes ), Université de Nantes ( UN ) -Hôtel-Dieu de Nantes-Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Centre hospitalier universitaire de Nantes ( CHU Nantes ), Université Libre de Bruxelles [Bruxelles] ( ULB ), Investigation Clinique et d’Accès aux Ressources Biologiques (Plate-forme) - Clinical Investigation and Access to BioResources ( ICAReB ), Immunité et cancer ( U932 ), Université Paris Descartes - Paris 5 ( UPD5 ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Institut Curie, Alloimmunité-Autoimmunité-Transplantation ( A2T ), and Université Paris Diderot - Paris 7 ( UPD7 ) -Institut National de la Santé et de la Recherche Médicale ( INSERM )

Subjects
0301 basic medicine, Lipopolysaccharides, Whole blood stimulation, CD3 Complex, medicine.medical_treatment, CD8 Antigens, Point-of-Care Systems, Immunology, Stimulation, Blood Donors, Immunologic Tests, Peripheral blood mononuclear cell, Antibodies, Functional immune responses, 03 medical and health sciences, Immune system, medicine, [ SDV.IMM ] Life Sciences [q-bio]/Immunology, Immunology and Allergy, Humans, Whole blood, biology, business.industry, Immunotherapy, Multi-center clinical studies, 3. Good health, Vaccination, Immunomonitoring, 030104 developmental biology, Gene Expression Regulation, PBMCs, biology.protein, [SDV.IMM]Life Sciences [q-bio]/Immunology, Cytokines, Antibody, business, Biomarkers
Abstract

Functional immune responses are increasingly important for clinical studies, providing in depth biomarker information to assess immunotherapy or vaccination. Incorporating functional immune assays into routine clinical practice has remained limited due to challenges in standardizing sample preparation. We recently described the use of a whole blood syringe-based system, TruCulture (R), which permits point-of-care standardized immune stimulation. Here, we report on a multi-center clinical study in seven FOCIS Centers of Excellence to directly compare TruCulture to conventional PBMC methods. Whole blood and PBMC5 from healthy donors were exposed to LPS, anti-CD3 anti-CD28 antibodies, or media alone. 55 protein analytes were analyzed centrally by Luminex multi-analyte profiling in a CLIA-certified laboratory. TruCulture responses showed greater reproducibility and improved the statistical power for monitoring differential immune response activation. The use of TruCulture addresses a major unmet need through a robust and flexible method for immunomonitoring that can be reproducibly applied in multi-center clinical studies. One sentence summary: A multi-center study revealed greater reproducibility from whole blood stimulation systems as compared to PBMC stimulation for studying induced immune responses. (C) 2017 Published by Elsevier Inc.

Published
2017

2. Immunotherapy reduces allergen-mediated CD66b expression and myeloperoxidase levels on human neutrophils from allergic patients

Author

Antonio Vega, Miguel Blanca, Cristina Chamorro, Javier Monteseirín, Inmaculada Ventura, Elisa Gomez, Rocío Aroca, Ramón Pérez-Cano, [Aroca,R, Chamorro,C, Vega,A, Ventura,A, Gómez,E, Monteseirín,J] Servicio Regional de Inmunología y Alergia, Hospital Universitario Virgen Macarena, Sevilla, Spain. [Pérez-Cano,R, Monteseirín,J] Departamento de Medicina, Facultad de Medicina, Universidad de Sevilla, Sevilla, Spain. [Blanca,M] Research Laboratory, Carlos Haya Hospital-Fundacion IMABIS, Málaga, Spain., and Conceived and designed the experiments: JM. Performed the experiments: RA CC AV IV EG. Analyzed the data: JM. Wrote the paper: JM MB AV RPC.

Subjects
Hipersensibilidad, Lipopolysaccharides, Male, Allergy, Chemicals and Drugs::Amino Acids, Peptides, and Proteins::Proteins::Glycoproteins::Membrane Glycoproteins::GPI-Linked Proteins [Medical Subject Headings], Pulmonology, Neutrophils, Physiology, medicine.medical_treatment, lcsh:Medicine, medicine.disease_cause, Immunoglobulin E, Biochemistry, Organisms::Eukaryota::Animals::Chordata::Vertebrates::Mammals::Primates::Haplorhini::Catarrhini::Hominidae::Humans [Medical Subject Headings], Allergen, Animal Cells, Immune Physiology, Molecular Cell Biology, Allergies, Medicine and Health Sciences, Chemicals and Drugs::Amino Acids, Peptides, and Proteins::Proteins::Glycoproteins::Membrane Glycoproteins::Cell Adhesion Molecules [Medical Subject Headings], lcsh:Science, Immune Response, CEACAM8 protein, human, Chemicals and Drugs::Amino Acids, Peptides, and Proteins::Proteins::Blood Proteins::Immunoproteins::Immunoglobulins::Antibodies::Immunoglobulin Isotypes::Immunoglobulin E [Medical Subject Headings], Cells, Cultured, Analytical, Diagnostic and Therapeutic Techniques and Equipment::Investigative Techniques::Therapeutic Irrigation::Nasal Lavage::Nasal Lavage Fluid [Medical Subject Headings], Multidisciplinary, Immune System Proteins, biology, Chemicals and Drugs::Enzymes and Coenzymes::Enzymes::Oxidoreductases::Peroxidases::Peroxidase [Medical Subject Headings], Flow Cytometry, Up-Regulation, Analytical, Diagnostic and Therapeutic Techniques and Equipment::Therapeutics::Biological Therapy::Immunomodulation::Immunotherapy::Immunosuppression::Desensitization, Immunologic [Medical Subject Headings], Inmunoglobulina E, Myeloperoxidase, Analytical, Diagnostic and Therapeutic Techniques and Equipment::Diagnosis::Diagnostic Techniques and Procedures::Clinical Laboratory Techniques::Cytological Techniques::Cell Separation::Flow Cytometry [Medical Subject Headings], Female, Nasal Lavage Fluid, Immunotherapy, medicine.symptom, Cellular Types, Research Article, Adult, Neutrófilos, Immune Cells, Immunology, Diseases::Immune System Diseases::Hypersensitivity [Medical Subject Headings], Check Tags::Male [Medical Subject Headings], Inflammation, GPI-Linked Proteins, Antibodies, Chemicals and Drugs::Carbohydrates::Glycoconjugates::Lipopolysaccharides [Medical Subject Headings], Antígenos CD, Antigens, CD, medicine, Named Groups::Persons::Age Groups::Adult [Medical Subject Headings], Hypersensitivity, Cell Adhesion, Humans, Asthma, Peroxidase, Phenomena and Processes::Chemical Phenomena::Biochemical Phenomena::Biochemical Processes::Up-Regulation [Medical Subject Headings], lcsh:R, Immunity, Biology and Life Sciences, Proteins, Cell Biology, medicine.disease, Check Tags::Female [Medical Subject Headings], Desensitization, Immunologic, Anatomy::Cells::Cells, Cultured [Medical Subject Headings], Immunoglobulin G, biology.protein, Moléculas de Adhesión Celular, Chemicals and Drugs::Biological Factors::Antigens::Antigens, Surface::Antigens, Differentiation::Antigens, CD [Medical Subject Headings], Clinical Immunology, lcsh:Q, Anatomy::Cells::Blood Cells::Leukocytes::Granulocytes::Neutrophils [Medical Subject Headings], Cell Adhesion Molecules, Chemicals and Drugs::Amino Acids, Peptides, and Proteins::Proteins::Blood Proteins::Immunoproteins::Immunoglobulins::Antibodies::Immunoglobulin Isotypes::Immunoglobulin G [Medical Subject Headings]
Abstract

Journal Article; Research Support, Non-U.S. Gov't; CD66b is a member of the carcinoembryonic antigen family, which mediates the adhesion between neutrophils and to endothelial cells. Allergen-specific immunotherapy is widely used to treat allergic diseases, and the molecular mechanisms underlying this therapy are poorly understood. The present work was undertaken to analyze A) the in vitro effect of allergens and immunotherapy on cell-surface CD66b expression of neutrophils from patients with allergic asthma and rhinitis and B) the in vivo effect of immunotherapy on cell-surface CD66b expression of neutrophils from nasal lavage fluid during the spring season. Myeloperoxidase expression and activity was also analyzed in nasal lavage fluid as a general marker of neutrophil activation. RESULTS CD66b cell-surface expression is upregulated in vitro in response to allergens, and significantly reduced by immunotherapy (p

Published
2014