SUMMARY B cells are activated in vivo following the B-cell receptors (BCRs) binding to antigens captured on the surfaces of antigen presenting cells. Antigen binding results in BCR microclustering and signaling, however, the molecular nature of the signaling-active BCR clusters is not well understood. Using new single molecule imaging techniques we provide evidence that within microclusters, the binding of monovalent membrane antigens results in the assembly of immobile signaling-active BCR oligomers. The oligomerization depends on interactions between the membrane-proximal Cμ4 domains of the mIg that are both necessary and sufficient for assembly. Antigen-bound BCRs that lacked the Cμ4 domain failed to cluster and signal and conversely, Cμ4 domain alone clustered spontaneously and activated B cells. These results support a novel mechanism for the initiation of BCR signaling in which antigen binding induces a conformational change in the Fc portion of the BCR revealing an interface that promotes BCR clustering.

It has been speculated that heart valve interstitial cells (VICs) maintain valvular tissue homeostasis through regulated extracellular matrix (primarily collagen) biosynthesis. VICs appear to be phenotypically plastic, inasmuch as they transdifferentiate into myofibroblasts during valve development, disease, and remodeling. Under normal physiological conditions, transvalvular pressures (TVPs) on the right and left side of the heart are vastly different. Hence, we hypothesize that higher left-side TVPs impose larger local tissue stress on VICs, which increases their stiffness through cytoskeletal composition, and that this relation affects collagen biosynthesis. To evaluate this hypothesis, isolated ovine VICs from the four heart valves were subjected to micropipette aspiration to assess cellular stiffness, and cytoskeletal composition and collagen biosynthesis were quantified by using the surrogates smooth muscle alpha-actin (SMA) and heat shock protein 47 (HSP47), respectively. VICs from the aortic and mitral valves were significantly stiffer (P < 0.001) than those from the pulmonary and tricuspid valves. Left-side isolated VICs contained significantly more (P < 0.001) SMA and HSP47 than right-side VICs. Mean VIC stiffness correlated well (r = 0.973) with TVP; SMA and HSP47 also correlated well (r = 0.996) with one another. Assays were repeated for VICs in situ, and, as with in vitro results, left-side VIC protein levels were significantly greater (P < 0.05). These findings suggest that VICs respond to local tissue stress by altering cellular stiffness (through SMA content) and collagen biosynthesis. This functional VIC stress-dependent biosynthetic relation may be crucial in maintaining valvular tissue homeostasis and also prove useful in understanding valvular pathologies.

While both infections and vaccines induce memory B cell (MBC) populations that participate in secondary immune responses, the MBCs generated in each case can differ. Here we compare SARS-CoV-2 spike receptor binding domain (S1-RBD)-specific primary MBCs that form in response to infection or a single mRNA vaccination. Both primary MBC populations have similar frequencies in the blood, and respond to a second S1-RBD exposure by rapidly producing plasmablasts with an abundant IgA+ subset, and secondary MBCs that are mostly IgG+ and cross-react on the B.1.351 variant. However, infection-induced primary MBCs have better antigen-binding capacity and generate more plasmablasts and secondary MBCs of the classical and atypical subsets than vaccine-induced primary MBCs. Our results suggest that infection-induced primary MBCs have undergone more affinity maturation than vaccine-induced primary MBCs and produce more robust secondary responses.

Non-alcoholic steatohepatitis (NASH) affects 3–5% of the U. S. population having severe clinical complications to the development of fibrosis and end-stage liver diseases such as cirrhosis and hepatocellular carcinoma. A critical cause of NASH is chronic systemic inflammation promoted by innate immune cells such as liver macrophages (Mϕ) and natural killer (NK) cells. However, little is known about how the crosstalk between Mϕ and NK cells contributes to regulate NASH progression to fibrosis. In this report, we demonstrate that NKp46+ cells play an important role in preventing NASH progression to fibrosis by regulating M1/M2 polarization of liver Mϕ. Using a murine model of NASH, we demonstrate that DX5+NKp46+ NK cells are increased during disease and play a role in polarizing Mϕ toward M1-like phenotypes. This NK’s immunoregulatory function depends on the production of IFN-γ but not by granzyme-mediated cytolytic activity. Notably, depletion of NKp46+ cells promote the development of fibrosis with increased expression of profibrogenic genes as well as skewed M2 Mϕ phenotypes in hepatic tissues. Conclusion NK cell-derived IFN-γ may be essential for maintaining a balanced inflammatory environment that promotes tissue integrity and limiting NASH progression to fibrosis.