The ability of the gut immune system to protect the mucosa against pathogens by long-term production of antigen-specific IgA antibodies is pivotal to mucosal vaccine development. A recent study by Macpherson and colleagues addressed the importance of mucosal memory after mono-colonization of germ-free mice with a commensal bacterial strain dependent on nutrients not present in the mammalian hosts (Hapfelmeier et al., 2010
). Using this reversible colonization model, the authors found that repeated bacterial exposures gave rise to increasing specific SIgA titers in an additive rather than a synergistic manner, failing to exhibit a classical prime-boost effect. Interestingly, in the absence of competing antigens, the half-life of the LP plasma cells was very long, whereas in the presence of other bacteria that triggered the formation of new IgA plasma cells, the lifespan of the IgA plasma cells in the gut LP was dramatically reduced (Hapfelmeier et al., 2010
). Based on these observations, the authors concluded that memory B cells did not develop, and that, although LP plasma cells had the potential for longevity, they were short-lived in conventionally reared mice.
These conclusions may be relevant for gut B cell responses driven by the microbiota, but they contrast with findings after oral immunizations with TD-antigens, as discussed below. Because GC and SHM play a critical role for both types of IgA responses we believe it will be important to understand if there are fundamentally different regulatory principles that operate in the PP GC depending on the type of antigen that drive the B cell response (Bemark et al., 2012
). Whereas memory B cells can form independently of GC, such memory B cells, mostly recognizing TI-antigens, are of a different quality and harbor less mutations and affinity maturation than GC-dependent memory responses (Toyama et al., 2002
). To what extent specific anti-bacterial gut IgA responses are more or less mutated still need further investigation, but recent data suggest that they are more mutated and carry GC-dependent features (Lindner et al., 2012
). Hence, to understand why commensal bacteria do not stimulate memory B cell development, but TD-antigens, such as CT, do, we will have to predict that memory B cell development in PP is not an intrinsic property of the GC, but rather a quality that is under additional regulatory control. In this context, interactions with Tfh cells may be critical.
Tfh cells have been shown to induce a memory B cell phenotype in GC B cells cultured in vitro
and can produce IL-21, IL-10, IL-6, and IL-4, which all have been implicated as instructive signals in GC plasma cell and memory B cell differentiation (Casamayor-Palleja et al., 1996
; Hashiguchi et al., 2011
; Shlomchik and Weisel, 2012
). Given that PP Tfh appear to be different from archetypical Tfh in systemic secondary lymphoid tissues, we propose that they may provide instructive signals to B cell memory development even in the absence of cognate interactions. Hence, failure to develop memory B cells, concomitant with a strong induction of long-lived plasma cells, as reported by Hapfelmeier et al. (2010)
may be a consequence of what type of, or lack of, Tfh activity that the commensal bacteria induced. In conventionally reared mice this situation may not have occurred as a multitude of commensal species are present in the intestine and potentially can stimulate a broadly functional Tfh population in PP GC. Of note, the composition of the microbiota plays an important role in shaping gut T cell functions as evident in studies showing, for example, that segmented filamentous bacteria stimulate the development of TH
17 cells, which dramatically could influence activation of mucosal B cells (Ivanov et al., 2009
; Datta et al., 2010
). In addition, we must consider that transiently colonizing germ-free mice with E. coli
K-12 bacteria most probably will lead to a TLR-mediated hyper-responsive state, which could facilitate BCR signaling and result in a TI-type of response, lacking memory development. Nevertheless, future studies are much needed to resolve this important and puzzling question.
Humans indeed maintain a sizable proportion of blood B cells that appear to be circulating IgA+
memory cells (Klein et al., 1998
; Harris et al., 2009
; Tengvall et al., 2010
). Furthermore, IgA-producing plasma cells reactive to rotaviral antigens were isolated from the duodenum of 9 out of 10 human adults that had not recently experienced rotavirus, arguing for the presence of long-lived plasma cells in the gut LP (Di Niro et al., 2010
). In mice, we know from previous studies that anti-toxin IgA plasma cells can reside in the gut LP more than 6 months after oral immunizations with CT (Lycke and Holmgren, 1987
). However, after 12 months specific plasma cells had largely disappeared, but memory B cells in the GALT could easily be triggered by a challenge-immunization with oral CT, giving rise to a vigorous anti-toxin IgA plasma cell response in the gut LP within 3 days upon re-challenge (Lycke and Holmgren, 1987
). The duration of gut anti-toxin protection, thus, clearly reflected the ability of long-lived IgA memory B cells to elicit a rapid recall response to a renewed exposure to the antigen.
We have previously demonstrated that memory B cells from the GALT can be adoptively transferred to naїve syngeneic recipient mice, and upon an oral immunization with CT, elicit a strong gut IgA response in the LP (Lycke and Holmgren, 1989
). More recently we have observed that these transferable memory B cells are present exclusively among CD80+
B cells from spleen, MLN, and PP even 1 year after oral priming immunizations (Andersson et al., 2007
; Bemark et al., 2011
; Bemark et al., unpublished observation). Others have demonstrated that adoptively transferred rotavirus-specific memory B cells can effectively clear the infection from rotavirus-infected RAG-deficient mice (Williams et al., 1998
). Interestingly, in the latter study splenic memory B cells with the ability to clear rotavirus infection through SIgA production could be identified on the basis α4
integrin expression, whereas B cells lacking this integrin did not clear the infection. Taken together, it is clear that mucosal IgA responses against TD-antigens following infection or oral vaccination also effectively stimulate the development of antigen-specific memory B cells and long-lived plasma cells.
The generation of memory B cells after oral immunization raises several questions with regard to if, and in that case, how they differ from memory cells that are generated through systemic immunization. This question is not only important from a mechanistic point of view, but may also have important implications for the development of more effective mucosal vaccines. Moreover, if, as discussed above, B cells recognizing antigens from the microbiota enter PP GC, will they give rise to memory B cells and long-lived plasma cells, or are cognate B–T cell interactions needed? Given that the mucosal immune system holds several unique traits compared to the systemic immune system we would argue that it could also apply to the ability to support memory B cell and long-lived plasma cell development. There are indeed data to suggest that oral immunization can be more efficient than systemic immunization for the generation of long-term immune protection, especially against enteropathogenic infections (Lycke, 2012
). Whether this is because of the expression of α4
integrin on mucosal memory B cells, as suggested by the rotavirus studies, or could be a function of other properties of these memory B cells compared to systemic memory B cells is poorly understood. Moreover, we know that oral immunization stimulate mucosal and systemic memory B cell development, whereas systemic immunization fails to stimulate mucosal memory responses (Lycke, 2012
; Bemark et al., unpublished observation). Understanding what the difference is between the immunization routes is critical for vaccine development, in general, and for mucosal vaccine development, in particular. Presently, we lack information about the relationship between systemic memory IgG and mucosal memory IgA cells following oral immunization. Are these memory B cells clonally related and do they emanate from the same or different inductive sites? We are currently addressing these pressing questions using the NP-CT system and the B1-8hi
NP-specific IgH knock-in B cell adoptive transfer model described above, with the hope to defining which Tfh functions and molecular pathways that are involved in memory B cell development in the PP GC.