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.
Subject(s)
B-Lymphocytes/immunology , Immunologic Deficiency Syndromes/diagnosis , Lymphocyte Activation/genetics , Adolescent , Adult , Biomarkers/metabolism , Cell Differentiation , Cells, Cultured , Chemokine CXCL10/genetics , Chemokine CXCL10/metabolism , Chemokine CXCL9/genetics , Chemokine CXCL9/metabolism , Child , Female , Gene Expression Profiling , High-Throughput Screening Assays , Humans , Image Cytometry , Immunoglobulin Class Switching , Immunologic Deficiency Syndromes/immunology , Infant, Newborn , Male , Microarray Analysis , Middle Aged , RNA, Messenger/analysis , Transcriptome/immunology , Young AdultABSTRACT
The heterogeneous group of primary immunodeficiencies requires personalized diagnosis and therapy to acheive an optimal outcome for each patient. This was exemplified by two patients with intrinsic B-cell class-switch defects (subclass of Hyper-IgM syndromes), where lymphoproliferation and autoimmunity determined the clinical course for many years due to lack of exact diagnosis. Based on genetics or a novel functional diagnostic approach, a definite individual diagnosis was established for each patient and they started Rituximab therapy. Autoimmune phenomena and generalized lymphadenopathy disappeared and remained well controlled during the observation period (3-4 years) without adverse effects. Quality of life increased remarkably in both patients.