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.

Our understanding of the B cell subsets found in human blood and their functional significance has advanced significantly in the past decade. This has been aided by the evolution of high dimensional phenotypic tools such as mass cytometry and single cell RNA sequencing which have revealed heterogeneity in populations that were previously considered homogenous. Despite this, there is still uncertainty and variation between studies as to how B cell subsets are identified and named. This review will focus on the most commonly encountered subsets of B cells in human blood and will describe gating strategies for their identification by flow and mass cytometry. Important changes to population frequencies and function in common inflammatory and autoimmune diseases will also be described.

Background: Complement FH (FH) is a key regulator of complement activity whereas thrombin (FIIa) is central to hemostasis with both pro- and anticoagulant functions. Both have separately been shown to have auxiliary activities across the two systems. The purpose of this study was to determine the effect of FH on pro- and anti-coagulant functions and investigate the interaction between FH and thrombin. Methods: Tail bleeding time and hemolysis were measured in FH-deficient mice (CFH-/-). Activated partial thromboplastin time (aPTT) was determined in FH-depleted human plasma. FH effect on fibrin clot generation was investigated in turbidity assays and on activated protein C (APC) generation. Binding affinity of thrombin with FH was determined using surface plasmon resonance (SPR). Results: Tail bleeding time in CFH-/- mice was significantly prolonged compared to wild type mice. The aPTT in FH-depleted human plasma was elevated compared to normal plasma and restored by adding back FH to depleted plasma. Accordingly, FH enhanced thrombin-mediated fibrin clot generation by shortening lag time, increasing rate of clot formation and maximum turbidity, and affected clot structure. Despite this, FH also increased the rate of thrombin-mediated protein C (PC) activation, both in the presence and absence of soluble recombinant thrombomodulin (TM). Nanomolar affinity binding of FH with thrombin, but not prothrombin, was confirmed. Conclusion: Complement FH binds thrombin with strong affinity and acts as a novel cofactor that enhances both pro- and anticoagulant actions of thrombin. These data highlight an important role for FH in hemostasis.