Christian Hennig1, Claudia Ilginus2, Kaan Boztug3, Julia Skokowa4, Laszlo Marodi5, Anna Szaflarska6, Mareike Sass7, Claudio Pignata8, Sara Sebnem Kilic9, Isabel Caragol10, Ulrich Baumann2, Christoph Klein11, Karl Welte4, Gesine Hansen2. 1. Department of Pediatric Pneumology, Allergy and Neonatology, Hannover Medical School, Hannover, Germany. Electronic address: hennig.christian@mh-hannover.de. 2. Department of Pediatric Pneumology, Allergy and Neonatology, Hannover Medical School, Hannover, Germany. 3. Department of Pediatric Oncology and Hematology, Hannover Medical School, Hannover, Germany; Research Center for Molecular Medicine (CeMM) of the Austrian Academy of Sciences, Vienna, Austria; Department of Pediatrics and Adolescent Medicine, Medical University of Vienna, Vienna, Austria. 4. Department of Pediatrics and Molecular Hematopoiesis, Hannover Medical School, Hannover, Germany. 5. University of Debrecen and Department of Infectious and Pediatric Immunology, Medical and Health Science Center, Debrecen, Hungary. 6. Department of Clinical Immunology, Polish-American Institute of Pediatrics, Jagiellonian University, Medical College, Krakow, Poland. 7. Department of Pediatric Oncology and Hematology, Hannover Medical School, Hannover, Germany. 8. Department of Pediatrics, 'Federico II' University, Naples, Italy. 9. Department of Pediatrics, Uludag University Medical Faculty, Bursa, Turkey. 10. Pediatric Infectious Diseases and Immunodeficiencies Unit, Vall d'Hebron Hospital, Barcelona, Spain. 11. Department of Pediatric Oncology and Hematology, Hannover Medical School, Hannover, Germany; Department of Pediatrics, Ludwig-Maximilians-Universität, Munich, Germany.
Abstract
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.
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.