High-content cytometry and transcriptomic biomarker profiling of human B-cell activation

• Published in: Journal of allergy and clinical immunology 2014-01, Vol.133 (1), p.172-180.e10
• Main Authors: Hennig, Christian, MD, Ilginus, Claudia, Boztug, Kaan, MD, Skokowa, Julia, PhD, Marodi, Laszlo, MD, Szaflarska, Anna, MD, Sass, Mareike, Pignata, Claudio, MD, Kilic, Sara Sebnem, MD, Caragol, Isabel, MD, Baumann, Ulrich, MD, Klein, Christoph, MD, Welte, Karl, MD, Hansen, Gesine, MD
• Format: Article
• Language: English
• Subjects: Adolescent; Adult; Allergy and Immunology; B-cell immunology; B-Lymphocytes - immunology; Bacterial infections; Biological and medical sciences; Biomarkers; Biomarkers - metabolism; Cell Differentiation; Cells, Cultured; Chemokine CXCL10 - genetics; Chemokine CXCL10 - metabolism; Chemokine CXCL9 - genetics; Chemokine CXCL9 - metabolism; Child; chip cytometry; Defects; Disease; DNA repair; Female; Fundamental and applied biological sciences. Psychology; Fundamental immunology; Gene Expression Profiling; High-Throughput Screening Assays; Humans; Image Cytometry; Immune system; Immunoglobulin Class Switching; Immunologic Deficiency Syndromes - diagnosis; Immunologic Deficiency Syndromes - immunology; Immunopathology; Infant, Newborn; Infections; Lymphocyte Activation - genetics; Male; Medical sciences; Microarray Analysis; Middle Aged; Patients; primary antibody deficiency; Primary immunodeficiency; RNA, Messenger - analysis; Sarcoidosis. Granulomatous diseases of unproved etiology. Connective tissue diseases. Elastic tissue diseases. Vasculitis; Transcriptome - immunology; Young Adult
• ISSN: 0091-6749; 1097-6825
• DOI: 10.1016/j.jaci.2013.06.047

Background Primary antibody deficiencies represent the most prevalent, although very heterogeneous, group of inborn immunodeficiencies, with a puzzling complexity of cellular and molecular processes involved in disease pathogenesis. Objective We aimed to study in detail the kinetics of CD40 ligand/IL-21–induced B-cell differentiation to define new biomarker sets for further research into primary antibody deficiencies. Methods We applied high-content screening methods to monitor B-cell activation on the cellular (chip cytometry) and transcriptomic (RNA microarray) levels. Results The complete activation process, including stepwise changes in protein and RNA expression patterns, entry into the cell cycle, proliferation and expression of activation-induced cytidine deaminase (AID), DNA repair enzymes, and post–class-switch expression of IgA and IgG, was successfully monitored during in vitro differentiation. We identified a number of unknown pathways engaged during B-cell activation, such as CXCL9/CXCL10 secretion by B cells. Finally, we evaluated a deduced set of biomarkers on a group of 18 patients with putative or proved intrinsic B-cell defects recruited from the European Society for Immunodeficiencies database and successfully predicted 2 AID defects and 1 DNA repair defect. Complete absence of class-switched B cells was a sensitive predictor of AID deficiency and should be further evaluated as a diagnostic biomarker. Conclusion The biomarkers found in this study could be used to further study the complex process of B-cell activation and to understand conditions that lead to the development of primary antibody deficiencies.