Antonio A. Freitas

We investigated the ability of monoclonal B cells to restore primary and secondary T-cell dependent antibody responses in adoptive immune-deficient hosts. Priming induced B cell activation and expansion, AID expression, antibody production and the generation of IgM⁺IgG^- and IgM^-IgG⁺ antigen-experienced B-cell subsets that persisted in the lymphopenic environment by cell division. Upon secondary transfer and recall the IgM^-IgG⁺ cells responded by the production of antigen-specific IgG while the IgM⁺ memory cells secreted mainly IgM and little IgG, but generated new B cells expressing germinal center markers. The recall responses were more efficient if the antigenic boost was delayed suggesting that a period of adaptation is necessary before the transferred cells are able to respond. Overall these findings indicate that reconstitution of a functional and complete memory pool requires transfer of all different antigen-experienced B cell subsets. We also found that the size of the memory B cell pool did not rely on the number of the responding naïve B cells, suggesting autonomous homeostatic controls for naïve and memory B cells. By reconstituting a stable memory B cell pool in immune-deficient hosts using a monoclonal high-affinity B cell population we demonstrate the potential value of B cell adoptive immunotherapy.

The ability of CD4+ T cells to change their phenotype and to specialize into different functional subsets may enhance the risk of autoimmune diseases. Here we investigate how a pleiotropic cytokine interleukin (IL)-15 may modify the functional commitment of CD4+ T cells expressing the lineage-associated transcription factors: forkhead box P3 (Foxp3; Treg) and RORγt (Th17) in the context of inflammatory bowel disease (IBD). We demonstrate in mice that impaired delivery of IL-15 to CD4+ T cells in the colon downmodulates Foxp3 expression (diminishing STAT5 phosphorylation) and enhances RORγt expression (by upregulating the expression of Runx1). In consequence, CD4+ T cells deprived of IL-15 rapidly trigger IBD characterized by enhanced production of pro-inflammatory cytokines (interferon-γ, IL-6) and accumulation of Th1/Th17 cells. Overall, our findings indicate a potentially beneficial role of IL-15 in IBD by fine-tuning the balance between Treg and Th17 cells and controlling intestinal inflammation.

Many species of bacteria use quorum sensing to sense the amount of secreted metabolites and to adapt their growth according to their population density. We asked whether similar mechanisms would operate in lymphocyte homeostasis. We investigated the regulation of the size of interleukin-2 (IL-2)–producing CD4+ T cell (IL-2p) pool using different IL-2 reporter mice. We found that in the absence of either IL-2 or regulatory CD4+ T (T reg) cells, the number of IL-2p cells increases. Administration of IL-2 decreases the number of cells of the IL-2p cell subset and, pertinently, abrogates their ability to produce IL-2 upon in vivo cognate stimulation, while increasing T reg cell numbers. We propose that control of the IL-2p cell numbers occurs via a quorum sensing–like feedback loop where the produced IL-2 is sensed by both the activated CD4+ T cell pool and by T reg cells, which reciprocally regulate cells of the IL-2p cell subset. In conclusion, IL-2 acts as a self-regulatory circuit integrating the homeostasis of activated and T reg cells as CD4+ T cells restrain their growth by monitoring IL-2 levels, thereby preventing uncontrolled responses and autoimmunity.

Maintenance of plasma IgM levels is critical for immune system function and homeostasis in humans and mice. However, the mechanisms that control homeostasis of the activated IgM-secreting B cells are unknown. After adoptive transfer into immune-deficient hosts, B lymphocytes expand poorly, but fully reconstitute the pool of natural IgM-secreting cells and circulating IgM levels. By using sequential cell transfers and B cell populations from several mutant mice, we were able to identify novel mechanisms regulating the size of the IgM-secreting B cell pool. Contrary to previous mechanisms described regulating homeostasis, which involve competition for the same niche by cells having overlapping survival requirements, homeostasis of the innate IgM-secreting B cell pool is also achieved when B cell populations are able to monitor the number of activated B cells by detecting their secreted products. Notably, B cell populations are able to assess the density of activated B cells by sensing their secreted IgG. This process involves the FcγRIIB, a low-affinity IgG receptor that is expressed on B cells and acts as a negative regulator of B cell activation, and its intracellular effector the inositol phosphatase SHIP. As a result of the engagement of this inhibitory pathway, the number of activated IgM-secreting B cells is kept under control. We hypothesize that malfunction of this quorum-sensing mechanism may lead to uncontrolled B cell activation and autoimmunity.

Maintenance of plasma IgM levels is critical for immune system function and homeostasis in humans and mice. However, the mechanisms that control homeostasis of the activated IgM-secreting B cells are unknown. After adoptive transfer into immune-deficient hosts, B-lymphocytes expand poorly but fully reconstitute the pool of natural IgM-secreting cells and circulating IgM levels. By using sequential cell transfers and B cell populations from several mutant mice, we were able to identify novel mechanisms regulating the size of the IgM-secreting B cell pool. Contrary to previous mechanisms described regulating homeostasis, which involve competition for the same niche by cells having overlapping survival requirements, homeostasis of the innate IgM-secreting B cell pool is also achieved when B cells populations are able to monitor the number of activated B cells by detecting their secreted products. Notably, B cell populations are able to assess the density of activated B cells by sensing their secreted IgG. This process involves the Fc{\gamma}RIIB, a low-affinity IgG receptor that is expressed on B cells and acts as a negative regulator of B cell activation, and its intracellular effector the inositol phosphatase SHIP. As a result of the engagement of this inhibitory pathway the number of activated IgM-secreting B cells is kept under control. We hypothesize that malfunction of this quorum-sensing mechanism may lead to uncontrolled B cell activation and autoimmunity.

Homeostasis of lymphocyte numbers is believed to be due to competition between cellular populations for a common niche of restricted size, defined by the combination of interactions and trophic factors required for cell survival. Here we propose a new mechanism: homeostasis of lymphocyte numbers could also be achieved by the ability of lymphocytes to perceive the density of their own populations. Such a mechanism would be reminiscent of the primordial quorum-sensing systems used by bacteria, in which some bacteria sense the accumulation of bacterial metabolites secreted by other elements of the population, allowing them to “count” the number of cells present and adapt their growth accordingly. We propose that homeostasis of CD4⁺ T cell numbers may occur via a quorum-sensing-like mechanism, where IL-2 is produced by activated CD4⁺ T cells and sensed by a population of CD4⁺ Treg cells that expresses the high-affinity IL-2Rα-chain and can regulate the number of activated IL-2-producing CD4⁺ T cells and the total CD4⁺ T cell population. In other words, CD4⁺ T cell populations can restrain their growth by monitoring the number of activated cells, thus preventing uncontrolled lymphocyte proliferation during immune responses. We hypothesize that malfunction of this quorum-sensing mechanism may lead to uncontrolled T cell activation and autoimmunity. Finally, we present a mathematical model that describes the key role of IL-2 and quorum-sensing mechanisms in CD4⁺ T cell homeostasis during an immune response.

We here describe novel aspects of CD8⁺ and CD4⁺ T cell subset interactions that may be clinically relevant and provide new tools for regulating the reconstitution of the peripheral CD8⁺ T cell pools in immune-deficient states. We show that the reconstitution capacity of transferred isolated naïve CD8⁺ T cells and their differentiation of effector functions is limited, but both dramatically increase upon the co-transfer of CD4⁺ T cells. This helper effect is complex and determined by multiple factors. It was directly correlated to the number of helper cells, required the continuous presence of the CD4⁺ T cells, dependent on host antigen-presenting cells (APCs) expressing CD40 and on the formation of CD4/CD8/APC cell clusters. By comparing the recovery of (CD44⁺CD62L^high) T_CM and (CD44⁺CD62L^low) T_EM CD8⁺ T cells, we found that the accumulation of T_CM and T_EM subsets is differentially regulated. T_CM-cell accumulation depended mainly on type I interferons, interleukin (IL)-6, and IL-15, but was independent of CD4⁺ T-cell help. In contrast, T_EM-cell expansion was mainly determined by CD4⁺ T-cell help and dependent on the expression of IL-2Rβ by CD8 cells, on IL-2 produced by CD4⁺ T-cells, on IL-15 and to a minor extent on IL-6.