Understanding mechanisms underlying the commitment to a particular B cell fate has clinical importance. Memory cells can proliferate extensively upon secondary challenge, and long-lived plasma cells can secrete antibody for long periods of time without antigen contact. Therefore, the prevalence of autoreactivity in the memory population may result in a chronic antibody-mediated disease. In contrast, short-lived plasma cells may generate large amounts of a pathogenic antibody that, as shown in this study, does not necessarily need to be of the highest affinity to cause organ damage.
Through the analysis of Fcγ−/−
mice, we have demonstrated that DCs participate in determining B cell differentiation in vivo. Fcγ−/−
B cells became predominantly splenic plasma cells upon activation instead of forming the more heterogeneous population of short-lived and long-lived plasma cells and memory cells seen in WT mice. Current data suggest that the majority of splenic plasma cells are short lived, whereas plasma cells in the BM can be either short lived or long lived (15
). In our study, Fcγ−/−
mice had a transient surge of plasma cells in the spleen, while displaying a diminished rise in plasma cells in the BM and a reduced memory B cell response. In contrast, WT mice had a smaller population of splenic plasma cells and an increase in BM plasma cells after immunization and demonstrated B cell memory upon secondary challenge with antigen. Fcγ−/−
mice produced significantly higher serum titers of specific antibody than WT mice and displayed class switching to IgG2a as well as IgG1. The increase in serum antipeptide and anti-DNA response appears to be an FcR-dependent phenomenon because FcγRI−/−
immunized with MAP peptide showed a similar phenotype (Fig. S5, available at http://www.jem.org/cgi/content/full/jem.20070731/DC1
). These features of the response may account for the increased glomerular dysfunction. Interestingly, antibodies from Fcγ−/−
displayed less affinity maturation. This serologic data were confirmed by the demonstration of reduced GL-7–positive cells in splenic B cells of Fcγ−/−
mice (unpublished data) and less somatic mutation in antigen-specific hybridomas. Because some studies suggest that short-lived and long-lived plasma cells arise from completely separate precursors (25
), we are currently studying the B cells responding to antigen in WT and Fcγ−/−
mice to determine if the same clones are activated or whether the plasma cell precursors differ between the strains.
DCs have previously been shown to mediate a critical function in B cell activation by transporting antigen from the periphery to T and B cells (26
). The altered B cell phenotype we observed in Fcγ−/−
could also be traced to the influence of Fcγ−/−
DCs and IL-12. Fcγ−/−
DCs had a more mature phenotype and produced at least twofold more IL-12 than WT DCs. Several studies have established a role for IL-12 in B cell differentiation and function. IL-12 can induce plasma cell formation and B cell heavy chain class switching to IgG1 and IgG2a. The switch to IgG2 is IFN-γ–dependent, whereas the promotion of differentiation to plasma cells and the switch to IgG1 does not require IFN-γ (27
). Studies in human primary B cells have also shown that IL-12 up-regulates IL-12Rβ2 expression and IFN-γ production by B cells, possibly allowing them to function as B effector cells and promote the activation of Th1 cells (28
). Notably, although IL-12 promotes short-lived plasma cells, perhaps in synergy with CD40L (27
), memory B cell differentiation occurs independently of IL-12 (22
). In our study, when IL-12 was administered to WT mice, the mice responded to antigen immunization with the generation of more splenic plasma cells and fewer BM plasma cells, a pattern mimicking that seen in immunized Fcγ−/−
mice. Therefore, IL-12 produced by DCs seems to be highly influential in directing B cell differentiation. It is possible that this difference in differentiation pathway reflects a change in the environment in which the B cell resides rather than a direct effect on the B cell. We do not favor this explanation, as chemokine expression in the spleen is similar in both strains (Fig. S6, available at http://www.jem.org/cgi/content/full/jem.20070731/DC1
). Furthermore, IL-12 has been shown by others to act directly on B cells (21
). Interestingly, CR2 (CD21/CD35) has also been reported to diminish the development of long-lived plasma cells while leaving intact the generation of short-lived plasmablasts, germinal center, and memory B cells. This effect is mediated through the regulation of Blimp-1 and XBP-1, which are critical transcription factors in plasma cell differentiation (29
). CR2, like FcR, appears to function through a B cell–intrinsic pathway that specifically arrests the differentiation of long-lived plasma cells. We should note, however, that antigen-specific T cell proliferation was enhanced in Fcγ−/−
mice. The enhanced number of antigen-specific T cells expressing CD40L and available to interact with antigen-specific B cells may also contribute to the preferential generation of short-lived plasma cells. Thus, both the direct action of DCs on B cells as well as the action of T cells in Fcγ−/−
mice may contribute to skewing the B cell response.
Although the differences in IL-12 production between the mouse strains may seem small, we have previously shown in studies of hormonal effects on B cells that a 20% change in expression of CD22 can affect B cell receptor signaling and alter negative selection (30
). Recently, McGaha et al. (31
) have shown that a 50% increase in FcγRIIb expression in B cells can prevent disease in lupus-prone mice. Thus, it should not be surprising that a twofold increase in IL-12 production by DCs can affect B cell differentiation. In fact, a study of experimental myasthenia gravis in mice also demonstrated that exogenous IL-12 given in a similar protocol led to increased serum antibody (27
A previous study reported no alteration in humoral response to the hapten NP in Fcγ−/− mice. This study was performed on C57BL/6 mice deficient in the FcR γ chain and may reveal a strain-dependent difference in FcR γ chain function. It is also possible, however, that the force of selection for high-affinity antibodies in the NP model is so strong that differences in B cell differentiation were obscured. Some effects were seen that were similar to those we report. For example, in the reported study that Fcγ−/− mice had fewer splenic germinal centers after immunization.
We do not yet fully understand why the absence of the γ chain leads to the observed DC phenotype. Studies on γ chain–deficient mice have focused on alterations in expression and function of FcRI, FcRIII, and FcRIV (32
). To confirm that the serological changes were mediated through FcR engagement, we have immunized FcRIII-deficient mice with MAP peptide. These mice show increased serum antipeptide antibodies (fivefold increase) and an activated phenotype of BMDCs. Similarly, DCs derived from BM cells under serum-free conditions also display an activated phenotype, which is, again, consistent with FcR engagement modulating DC function (26
). The activated phenotype was abrogated by addition of purified IgG to the culture. Although γ chain–containing Fc receptors have been demonstrated to be activating receptors, we speculated that engagement of these receptors may diminish the response to other activating receptors. Such reciprocal regulation of toll-like receptor 9 and IgE receptor signaling has already been demonstrated. Production of type I IFN by plasmacytoid DC is inhibited by preexposure of FcεRI to IgG (33
). These two signaling pathways counterregulate the development of Th1 and Th2 immune response. Park-Min et al. (34
) recently demonstrated that immune complexes binding to FcγRIII suppresses LPS-mediated IFNγ signaling in both human peripheral blood mononuclear cells derived macrophages and murine peritoneal macrophages. A recent publication reported that human peripheral monocyte-derived DC cultures exposed to immune complexes show diminished differentiation. Furthermore, in response to TLR-mediated stimulation, they produced less IL-12 (35
). This inhibition was mediated mainly through FcγRI but, to a lesser extent by FcγRII. Thus, there is precedent for an absence of engagement of activating FcRs permitting an enhanced response in other activation pathways. Although the mechanism for the altered function of Fcγ−/−
DC requires further confirmation, it is apparent that the difference in IL-12 production by DCs affects B cell differentiation. This study reports the first in vivo manipulation that leads to differential formation of short-lived and long-lived plasma cells. Understanding how to modulate B cell differentiation will facilitate an appreciation of the role of both short-lived and long lived plasma cells. Furthermore, knowledge of the B cell phenotype responsible for the production of antiself or antimicrobial antibodies will help in the design of therapeutic strategies in autoimmune disease, as well as in optimizing vaccine development.