B cells emerge from the bone marrow as transitional (TS) B cells that differentiate through T1, T2, and T3 stages to become naive B cells. We have identified a bifurcation of human B cell maturation from the T1 stage forming IgMhi and IgMlo developmental trajectories. IgMhi T2 cells have higher expression of α4β7 integrin and lower expression of IL-4 receptor (IL4R) compared with the IgMlo branch and are selectively recruited into gut-associated lymphoid tissue. IgMhi T2 cells also share transcriptomic features with marginal zone B cells (MZBs). Lineage progression from T1 cells to MZBs via an IgMhi trajectory is identified by pseudotime analysis of scRNA-sequencing data. Reduced frequency of IgMhi gut-homing T2 cells is observed in severe SLE and is associated with reduction of MZBs and their putative IgMhi precursors. The collapse of the gut-associated MZB maturational axis in severe SLE affirms its existence in health.

No abstract

B cells generate antibodies that are essential for immune protection, but their subgroups are poorly defined. Here, we perform undirected deep profiling of B cells in matched human lymphoid tissues from deceased transplant organ donors and blood. In addition to identifying unanticipated features of tissue-based B cell differentiation, we resolve two subsets of marginal zone B (MZB) cells differing in cell surface and transcriptomic profiles, clonal relationships to other subsets, enrichment of genes in the NOTCH pathway, distribution bias within splenic marginal zone microenvironment, and immunoglobulin repertoire diversity and hypermutation frequency. Each subset is present in spleen, gut-associated lymphoid tissue, mesenteric lymph nodes, and blood. MZB cells and the lineage from which they are derived are depleted in lupus nephritis. Here, we show that this depletion is of only one MZB subset. The other remains unchanged as a proportion of total B cells compared with health. Thus, it is important to factor MZB cell heterogeneity into studies of human B cell responses and pathology.

B cells emerge from the bone marrow as transitional (TS) B cells that differentiate through T1, T2 and T3 stages to become naïve B cells. We have identified a bifurcation of human B cell maturation from the T1 stage forming IgMhi and IgMlo developmental trajectories. IgMhi T2 cells have higher expression of α4β7 integrin and lower expression of IL4 receptor (IL4R) compared to the IgMlo branch and are selectively recruited into gut-associated lymphoid tissue. IgMhi T2 cells also share transcriptomic features with marginal zone B cells (MZB). Lineage progression from T1 cells to MZB via an IgMhi trajectory is identified by pseudotime analysis of scRNA-sequencing data. Reduced frequency of IgMhi gut homing T2 cells is observed in severe SLE and is associated with reduction of MZB and their putative IgMhi precursors. The collapse of the gut-associated MZB maturational axis in severe SLE affirms its existence and importance for maintaining health.

Multiplexed imaging technologies enable the study of biological tissues at single-cell resolution while preserving spatial information. Currently, high-dimension imaging data analysis is technology-specific and requires multiple tools, restricting analytical scalability and result reproducibility. Here we present SIMPLI (Single-cell Identification from MultiPLexed Images), a flexible and technology-agnostic software that unifies all steps of multiplexed imaging data analysis. After raw image processing, SIMPLI performs a spatially resolved, single-cell analysis of the tissue slide as well as cell-independent quantifications of marker expression to investigate features undetectable at the cell level. SIMPLI is highly customisable and can run on desktop computers as well as high-performance computing environments, enabling workflow parallelisation for large datasets. SIMPLI produces multiple tabular and graphical outputs at each step of the analysis. Its containerised implementation and minimum configuration requirements make SIMPLI a portable and reproducible solution for multiplexed imaging data analysis. Software is available at “SIMPLI [https://github.com/ciccalab/SIMPLI]”.