These

findings lead us to suggest that commensal microbiota contribute to the development of MZ B cells. With the exception of mice, little is known about the origin and development of marginal zone (MZ) B cells in mammals, including humans. The gradual decline in the CAL-101 mw number of circulating MZ B cells in splenoctomized patients suggests that the spleen may play a role in the development and/or maintenance of MZ B cells in humans [1, 2]. Because MZ B cells with somatically diversified Ig genes are found in young children [2-4] and in patients unable to form T-dependent germinal centers [2, 3, 5, 6], Weill et al. [7] proposed that human MZ B cells develop in gut-associated lymphoid tissue (GALT) in a T-independent manner, analogous to the strategy used by B cells in sheep and rabbits [7, 8]. Unlike the spleen, however, the requirement and/or role of GALT for peripheral B-cell development cannot be directly addressed in humans. Because we previously used rabbits in which organized GALT (comprised

of appendix, sacculus rotundus, and Peyer’s patches) Y-27632 mw was surgically removed at birth (GALTless) [9], we thought to use these GALTless rabbits to investigate the role of GALT in the development of B-cell subsets. These GALTless rabbits appeared healthy, with no apparent signs of infection, and exhibited a growth rate that was similar to control littermates. The frequency of peripheral IgM+ B cells in these GALTless rabbits was however, significantly reduced, but the identity of the B cells that were reduced or missing Baf-A1 could not be determined. In this study, we analyzed frozen tissues preserved from the spleens of these GALTless rabbits and found that both the follicular (FO) and MZ B-cell compartments were perturbed. MZ B cells are identified by expression (or lack thereof) of surface Ig, CD23, and CD27 [7, 10]. Because IgD is not found in rabbits [11], we tested if the expression of IgM, CD23, and CD27 can

be used to distinguish rabbit MZ B cells from FO B cells, that we previously [12, 13] described as CD23+ (Fig. 1A). Using a cross-reactive anti-human CD27 mAb and anti-rabbit L chain Ab, we found B cells in the margin of B-cell follicles (Fig. 1A) and these were IgMhi (Fig. 1B, left). We used anti-L chain Ab instead of anti-IgM for immunohistology, because the polyclonal anti-L chain Ab provides a stronger signal than does the mAb anti-μ chain Ab. The CD27+ B cells expressed higher levels of complement receptor, CD21 than CD27− B cells (Fig. 1B, middle), and most CD27+ B cells expressed CD1b (Fig. 1B, right), similar to the expression of CD1 isoforms on human (CD1c) and murine (CD1d) MZ B cells [7, 14]. We conclude that MZ B cells in the spleen can be identified as a CD27+CD23−IgMhi phenotype. Essentially all CD27+ cells in the spleen were B cells, and most of them were IgM+ (Fig. 1B); a few were class-switched B cells (Table 1).