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4 results on '"Nina Salabert-Le Guen"'

1. An easy and reliable whole blood freezing method for flow cytometry immuno-phenotyping and functional analyses

Author

Chevreuil Justine, Nina Salabert-Le Guen, Régis Josien, Jerome Martin, Rimbert Marie, and Cécile Braudeau

Subjects
0301 basic medicine, Histology, medicine.medical_treatment, Biology, Peripheral blood mononuclear cell, T-Lymphocytes, Regulatory, Cryopreservation, Pathology and Forensic Medicine, Flow cytometry, Immunophenotyping, Andrology, 03 medical and health sciences, 0302 clinical medicine, Freezing, medicine, Humans, B cell, Whole blood, medicine.diagnostic_test, FOXP3, Cell Biology, Flow Cytometry, 030104 developmental biology, medicine.anatomical_structure, Cytokine, 030220 oncology & carcinogenesis, Leukocytes, Mononuclear, Ex vivo
Abstract

BackgroundImmune profiling by flow cytometry is not always possible on fresh blood samples due to time and/or transport constraints. Besides, the cryopreservation of peripheral blood mononuclear cells (PBMC) requires on-site specialized lab facilities, thus severely restricting the extent by which blood immune monitoring can be applied to multicenter clinical studies. These major limitations can be addressed through the development of simplified whole blood freezing methods.MethodsIn this report, we describe an optimized easy protocol for rapid whole blood freezing with the CryoStor® CS10 solution. Using flow cytometry, we compared cellular viability and composition on cryopreserved whole blood samples to matched fresh blood, as well as fresh and frozen PBMC.ResultsThough partial loss of neutrophils was observed, leucocyte viability was routinely >75% and we verified the preservation of viable T cells, NK cells, monocytes, dendritic cells and eosinophils in frequencies similar to those observed in fresh samples. A moderate decrease in B cell frequencies was observed. Importantly, we validated the possibility to analyze major intracellular markers, such as FOXP3 and Helios in regulatory T cells. Finally, we demonstrated good functional preservation of CS10-cryopreserved cells through the analysis of intracellular cytokine production in ex vivo stimulated T cells (IFNg, IL-4, IL-17A,) and monocytes (IL-1b, IL-6, TNFa).ConclusionsIn conclusion, our protocol provides a robust method to apply reliable immune monitoring studies to cryopreserved whole blood samples, hence offering new important opportunities for the design of future multicenter clinical trials.

Published
2020

2. 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

3. Thymic stromal lymphopoietin does not activate human basophils

Author

Aurélie Fantou, Agathe Delbove, Luc Colas, Cécile Braudeau, Caroline Hémont, Vassili Soumelis, Caroline Poli, Gaelle Beriou, Nina Salabert-Le Guen, Régis Josien, Karine Amouriaux, Jerome Martin, Degauque, Nicolas, 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), Institut de transplantation urologie-néphrologie (ITUN), Université de Nantes (UN)-Centre hospitalier universitaire de Nantes (CHU Nantes), Innate Immunity and Immunotherapy (CRCINA-ÉQUIPE 7), Centre de Recherche en Cancérologie et Immunologie Nantes-Angers (CRCINA), Université d'Angers (UA)-Université de Nantes (UN)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Centre hospitalier universitaire de Nantes (CHU Nantes)-Université d'Angers (UA)-Université de Nantes (UN)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Centre hospitalier universitaire de Nantes (CHU Nantes), Laboratoire d’Immunologie, Centre d’Immunomonitorage Nantes Atlantique (CIMNA), Centre hospitalier universitaire de Nantes (CHU Nantes), Unité de recherche de l'institut du thorax (ITX-lab), Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Université de Nantes - UFR de Médecine et des Techniques Médicales (UFR MEDECINE), Immunité et cancer (U932), Université Paris Descartes - Paris 5 (UPD5)-Institut Curie [Paris]-Institut National de la Santé et de la Recherche Médicale (INSERM), Université de Nantes (UN)-Université de Nantes (UN)-Centre hospitalier universitaire de Nantes (CHU Nantes)-Centre National de la Recherche Scientifique (CNRS)-Université d'Angers (UA)-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)-Centre hospitalier universitaire de Nantes (CHU Nantes)-Centre National de la Recherche Scientifique (CNRS)-Université d'Angers (UA), unité de recherche de l'institut du thorax UMR1087 UMR6291 (ITX), Université de Nantes - UFR de Médecine et des Techniques Médicales (UFR MEDECINE), and Université de Nantes (UN)-Université de Nantes (UN)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)

Subjects
0301 basic medicine, Adult, Male, Thymic stromal lymphopoietin, [SDV.IMM] Life Sciences [q-bio]/Immunology, Immunology, 03 medical and health sciences, Young Adult, Thymic Stromal Lymphopoietin, Team1, Hypersensitivity, Immunology and Allergy, Medicine, Humans, CRTI, ComputingMilieux_MISCELLANEOUS, business.industry, Middle Aged, Basophils, Eosinophils, 030104 developmental biology, [SDV.IMM]Life Sciences [q-bio]/Immunology, Cytokines, Female, business, Signal Transduction
Abstract

International audience

Published
2017

4. Dysregulated Responsiveness of Circulating Dendritic Cells to Toll-Like Receptors in ANCA-Associated Vasculitis

Author

Régis Josien, Jerome Martin, Karine Amouriaux, Marie Rimbert, Audrey Besançon, Stéphanie Giraudet, Mohamed Hamidou, Marine Aliaga, Caroline Hémont, Cécile Braudeau, Nina Salabert-Le Guen, Caroline Terrien, Antoine Néel, Le Bihan, Sylvie, Laboratoires d'excellence - Immunothérapies Grand Ouest - - IGO2011 - ANR-11-LABX-0016 - LABX - VALID, 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), Institut de transplantation urologie-néphrologie (ITUN), Université de Nantes (UN)-Centre hospitalier universitaire de Nantes (CHU Nantes), Laboratoire d’Immunologie [CHU Nantes] (Centre d’Immunomonitorage Nantes Atlantique - CIMNA), Centre hospitalier universitaire de Nantes (CHU Nantes), Faculté de Médecine - Université de Nantes, Service de médecine interne [Nantes], Université de Nantes (UN)-Hôtel-Dieu-Centre hospitalier universitaire de Nantes (CHU Nantes), LabEX IGO Immunothérapie Grand Ouest, Nantes Université (Nantes Univ), A grant from the Direction de la Recherche Clinique et de l’Innovation (DRCI), CHU Nantes (#BRD/09/06) to RJ. The CIMNA was supported by the IMBIO-DC program supported by the 'Région des Pays de la Loire'. AN was supported by the Société Nationale Française de Médecine Interne (Bourse Marcel Simon)., ANR-11-LABX-0016,IGO,Immunothérapies Grand Ouest(2011), LabEx IGO 'Immunotherapy, Graft, Oncology' [Nantes], and ANR: ANR11-LABX-0016-01

Subjects
dysregulation, medicine.medical_treatment, T cell, anca-associated vasculitis, Immunology, Stimulation, 03 medical and health sciences, 0302 clinical medicine, medicine, Immunology and Allergy, dendritic cells, Original Research, Whole blood, 030203 arthritis & rheumatology, Toll-like receptor, [SDV.MHEP] Life Sciences [q-bio]/Human health and pathology, business.industry, medicine.disease, 3. Good health, Cytokine, medicine.anatomical_structure, il-12/il-23p40, toll-like receptor, Cytokine secretion, Granulomatosis with polyangiitis, Microscopic polyangiitis, business, [SDV.MHEP]Life Sciences [q-bio]/Human health and pathology, 030215 immunology
Abstract

International audience; Objective: Dendritic cells (DCs) are critical effectors of innate and adaptive immunity playing crucial roles in autoimmune responses. We previously showed that blood DC numbers were reduced in autoimmune antineutrophil cytoplasmic autoantibody-associated vasculitis (AAV). Here, we assessed toll-like receptor (TLR) responsiveness of blood DCs from patients with granulomatosis with polyangiitis (GPA) or microscopic polyangiitis (MPA).Methods: Blood samples from healthy controls (HCs), GPA, or MPA patients, without treatment, during acute phase (AP) or remission phase (RP) were analyzed. Cytokine production by DCs and T cells was assessed on whole blood by flow cytometry after TLRs or polyclonal stimulation, respectively.Results: We first showed that GPA and MPA are associated with a decreased blood DC number during AP. Conventional DCs (cDCs) from patients with GPA and MPA in AP exhibited a profound decrease of IL-12/IL-23p40 production after TLR3, 4, or 7/8 stimulation compared to patients in remission and HC, with a return to normal values in RP. TNFα secretion was also affected, with a decrease in cDCs from GPA patients in AP after TLR3 stimulation but an increase after TLR7/8 stimulation. By contrast, the responsiveness of plasmacytoid DCs to TLR7 and 9 was only marginally affected. Finally, we observed that IFNγ-producing CD4 + T cell frequency was significantly lower in AP-GPA patients than in HC.Conclusion: We describe, for the first time, a dysregulated response to TLRs of circulating DCs in AAV patients mostly affecting cDCs that exhibit an unexpected reduced inflammatory cytokine secretion possibly contributing to an altered Th cell response.

Published
2017

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