Tviblindi algorithm identifies branching developmental trajectories of human B‐cell development and describes abnormalities in RAG‐1 and WAS patients

Detailed knowledge of human B‐cell development is crucial for the proper interpretation of inborn errors of immunity and malignant diseases. It is of interest to understand the kinetics of protein expression changes during development, but also to properly interpret the major and possibly alternative developmental trajectories. We have investigated human samples from healthy individuals with the aim of describing all B‐cell developmental trajectories. We validated a 30‐parameter mass cytometry panel and demonstrated the utility of “vaevictis” visualization of B‐cell developmental stages. We used the trajectory inference tool “tviblindi” to exhaustively describe all trajectories leading to all developmental ends discovered in the data. Focusing on Natural Effector B cells, we demonstrated the dynamics of expression of nuclear factors (PAX‐5, TdT, Ki‐67, Bcl‐2), cytokine and chemokine receptors (CD127, CXCR4, CXCR5) in relation to the canonical B‐cell developmental stage markers. We observed branching of the memory development, where follicular memory formation was marked by CD73 expression. Lastly, we performed an analysis of two example cases of abnormal B‐cell development caused by mutations in RAG‐1 and Wiskott–Aldrich syndrome gene in patients with primary immunodeficiency. In conclusion, we developed, validated, and presented a comprehensive set of tools for the investigation of B‐cell development in the bone marrow compartment. Novel computational framework tviblindi identifies branching B cell developmental trajectories in healthy (HD) and abnormal (RAG‐1 and Wiskott–Aldrich syndrome primary immunodeficiency) bone marrow and peripheral blood samples using 30 parameter mass cytometry. Integrated vaevictis projection allows interpretation of trajectories to κ (grey arrow) and λ (black arrow) light chain expressing B cells, memory development branching trajectories (solid vs. dashed arrows), and abnormal trajectories caused by RAG‐1 and WAS gene mutations. The pseudotime line plot shows the dynamics of selected marker expressions along the trajectories. Created with BioRender.com.