Germinal centers (GCs) were described more than 125 years ago as compartments within secondary lymphoid organs that contained mitotic cells. Since then, it has become clear that this structure is the site of B cell clonal expansion, somatic hypermutation, and affinity-based selection, the combination of which results in the production of high-affinity antibodies. Decades of anatomical and functional studies have led to an overall model of how the GC reaction and affinity-based selection operate. More recently, the introduction of intravital imaging into the GC field has opened the door to direct investigation of certain key dynamic features of this microanatomic structure, sparking renewed interest in the relationship between cell movement and affinity maturation. We review these and other recent advances in our understanding of GCs, focusing on cellular dynamics and on the mechanism of selection of high-affinity B cells.

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SUMMARY The germinal center (GC) reaction produces high-affinity antibodies by random mutation and selective clonal expansion of B cells with high-affinity receptors. However, the mechanism by which B cells are selected remains unclear, as does the role of the two anatomically-defined areas of the GC, light zone (LZ) and dark zone (DZ). We combined a new transgenic photoactivatable green fluorescent protein (PA-GFP) tracer with multiphoton laser-scanning microscopy and flow cytometry to examine anatomically defined LZ and DZ B cells and GC selection. We find that B cell division is restricted to the DZ, and that there is a net vector of B cell movement from the DZ to the LZ. The decision to return from the LZ to the DZ and undergo clonal expansion is controlled by T cells, which discern between LZ B cells based on the amount of antigen captured, providing a mechanism for GC selection.

Dendritic cells (DCs) in lymphoid tissue arise from precursors that also produce monocytes and plasmacytoid DCs (pDCs). Where DC and monocyte lineage commitment occurs and the nature of the DC precursor that migrates from the bone marrow to peripheral lymphoid organs is unknown. We show that DC development progresses from the macrophage and DC precursor (MDP), to common DC precursors (CDPs) that give rise to pDCs and classical spleen DCs (cDCs), but not monocytes, and finally to committed precursors of cDCs (pre-cDCs). Pre-cDCs enter lymph nodes through and migrate along HEVs and later disperse and integrate into the DC network. Further cDC development involves cell division, controlled in part by regulatory T cells (Treg) and fms-related tyrosine kinase-3 (Flt3).Dendritic cells (DCs) are immune cells specialized to capture, process and present antigens to T lymphocytes to induce immunity or tolerance (1). Where commitment to DC development takes place, at what stage the monocyte lineage diverges from DCs, and the precise nature of the migrating DC precursor that moves from the bone marrow to the peripheral lymphoid organs is not known. These questions have been difficult to resolve in part because DC subsets are functionally and phenotypically diverse (2). For example, classical spleen DCs (cDCs) include two major functionally distinct subsets distinguished by the expression of a variety of C-type lectins and CD8 (2-4). Spleen and other tissues also contain plasmacytoid DCs (pDCs) that primarily initiate immune responses to nucleic acids (5,6).Lymphoid tissue cDCs, pDCs, and monocytes share a common progenitor called the macrophage and DC precursor (MDP) identified by its surface phenotype (Lin − cKit hi CD115 + CX 3 CR1 + Flt3 + ) (7,8), whereas a distinct progenitor called the common DC precursor (CDP) (Lin − cKit lo CD115 + Flt3 + ) is restricted to producing cDCs and pDCs (9,10). Although monocytes can develop many of the phenotypic features of DCs under inflammatory conditions (11-13), the cDC, pDC and monocyte lineages separate by the time they reach tissues and neither monocytes nor pDCs develop into cDCs under steady state conditions (8,14). Unlike monocytes and pDCs, cDCs in lymphoid tissues are thought to emerge from the bone marrow as immature cells that must further differentiate and divide in lymphoid organs (15,16 We searched for MDPs and CDPs in the blood and spleen by flow cytometry but could only detect them in the bone marrow ( Fig. 1A and Fig. S1). Although pre-cDCs can be identified in the spleen by combining density centrifugation and flow cytometry (18), we speculated that these cells could be identified directly by expression of Flt3 and the chemokine receptor, CX 3 CR1, which are expressed on other DC progenitors and also on mature cDCs (7,10,19). Indeed, we found a small but distinct population of lineage negative CD11c + MHC class II − SIRP-α int Flt3 + cells (pre-cDCs) in the bone marrow (0.2%), blood (0.03%), spleen (0.05%) and lymph nodes (LN) (0.03%) (Fig. 1B).To determine ...

Germinal centers (GCs) are the site of antibody diversification and affinity maturation, and as such are vitally important for humoral immunity. The study of GC biology has undergone a renaissance in the past 10 years, with a succession of findings that have transformed our understanding of the cellular dynamics of affinity maturation. In this review, we discuss recent developments in the field, with special emphasis on how GC cellular and clonal dynamics shape antibody affinity and diversity during the immune response.

Antibodies somatically mutate to attain high affinity in germinal centers (GCs). There, competition between B cell clones and among somatic mutants of each clone drives an increase in average affinity across the population. The extent to which higher-affinity cells eliminating competitors restricts clonal diversity is unknown. By combining multiphoton microscopy and sequencing, we show that tens to hundreds of distinct B cell clones seed each GC, and that GCs lose clonal diversity at widely disparate rates. Furthermore, efficient affinity maturation can occur in the absence of homogenizing selection, ensuring that many clones can mature in parallel within the same GC. Our findings have implications for development of vaccines in which antibodies with non-immunodominant specificities must be elicited, as is the case for HIV-1 and influenza.

Upon antigenic challenge, B cells enter the dark-zone (DZ) of germinal-centers (GC) to proliferate and hypermutate their immunoglobulin genes. Mutants with increased affinity are positively selected in the light-zone (LZ) to either differentiate into plasma and memory cells, or re-enter the DZ. The molecular circuits governing GC positive selection are not known. We show that the GC reaction requires the biphasic regulation of c-MYC expression, involving its transient induction during early GC commitment, its repression by BCL6 in DZ B cells, and its re-induction in B cells selected for DZ re-entry. Inhibition of MYC in vivo leads to GC collapse, indicating an essential role in GCs. These results have implications for the mechanism of GC selection and the role of MYC in lymphomagenesis.

Background Population-based data on COVID-19 are essential for guiding policies. There are few such studies, particularly from low or middle-income countries. Brazil is currently a hotspot for COVID-19 globally. We aimed to investigate severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) antibody prevalence by city and according to sex, age, ethnicity group, and socioeconomic status, and compare seroprevalence estimates with official statistics on deaths and cases. Methods In this repeated cross-sectional study, we did two seroprevalence surveys in 133 sentinel cities in all Brazilian states. We randomly selected households and randomly selected one individual from all household members. We excluded children younger than 1 year. Presence of antibodies against SARS-CoV-2 was assessed using a lateral flow point-of-care test, the WONDFO SARS-CoV-2 Antibody Test (Wondfo Biotech, Guangzhou, China), using two drops of blood from finger prick samples. This lateral-flow assay detects IgG and IgM isotypes that are specific to the SARS-CoV-2 receptor binding domain of the spike protein. Participants also answered short questionnaires on sociodemographic information (sex, age, education, ethnicity, household size, and household assets) and compliance with physical distancing measures. Findings We included 25 025 participants in the first survey (May 14–21) and 31 165 in the second (June 4–7). For the 83 (62%) cities with sample sizes of more than 200 participants in both surveys, the pooled seroprevalence increased from 1·9% (95% CI 1·7–2·1) to 3·1% (2·8–3·4). City-level prevalence ranged from 0% to 25·4% in both surveys. 11 (69%) of 16 cities with prevalence above 2·0% in the first survey were located in a stretch along a 2000 km of the Amazon river in the northern region. In the second survey, we found 34 cities with prevalence above 2·0%, which included the same 11 Amazon cities plus 14 from the northeast region, where prevalence was increasing rapidly. Prevalence levels were lower in the south and centre-west, and intermediate in the southeast, where the highest level was found in Rio de Janeiro (7·5% [4·2–12·2]). In the second survey, prevalence was similar in men and women, but an increased prevalence was observed in participants aged 20–59 years and those living in crowded conditions (4·4% [3·5–5·6] for those living with households with six or more people). Prevalence among Indigenous people was 6·4% (4·1–9·4) compared with 1·4% (1·2–1·7) among White people. Prevalence in the poorest socioeconomic quintile was 3·7% (3·2–4·3) compared with 1·7% (1·4–2·2) in the wealthiest quintile. Interpretation Antibody prevalence was highly heterogeneous by country region, with rapid initial escalation in Brazil's north and northeast. Prevalence is strongly associated with Indigenous ancestry and low socioeconomic status. These population subgroups are unlikely to be protected if the policy response to the pandemic by th...