Differential effects of IL-27 on human B cell subsets

IL-27 is a novel heterodimeric cytokine of the IL-12 family that plays an important role in the regulation of T cell responses. Its role on human B cells has not been previously studied. In this study, we show that both chains of the IL-27 receptor complex, IL-27R and gp130, are constitutively expressed at the surface of naive and memory human tonsillar B cells, and are induced on germinal center B cells following CD40 stimulation. In naive B cells, IL-27 induced strong STAT1 and STAT3 phosphorylation, whereas it induced moderate STAT1 and low STAT3 activation in memory B cells. IL-27 induced T-bet expression in naive and memory B cells stimulated by CD40 or surface Ig engagement, but induced significant IL-12Rbeta2 surface expression in anti-Ig-stimulated naive B cells only. In anti-Ig-stimulated naive or memory B cells, IL-27 also induced CD54, CD86, and CD95 surface expression. In addition, IL-27 increased proliferation of anti-Ig-activated naive B cells and of anti-CD40-activated naive and germinal center B cells, but not of CD40-activated memory B cells. These data indicate that the B cell response to IL-27 is modulated during B cell differentiation and varies depending on the mode of B cell activation.     

Novel diversity in IL-4-mediated responses in resting human naive B cells versus germinal center/memory B cells

Recent studies have defined several phenotypic and molecular changes associated with the maturation of naive human B cells within the milieu of germinal centers. Although naive B cells serve as natural precursors to germinal center (GC)/memory (M) subpopulations, little is known about the physiological requirements for the survival of the naive B cell pool in the absence of cell-cell contact or Ag-mediated activation. Because IL-4 induces expression of several membrane receptors such as CD23 which are uniquely present on resting human naive B lymphocytes, we hypothesized that these cells might be intrinsically programmed to respond to IL-4 in the absence of cell division. Using buoyant density-dependent isolation and further enrichment by negative/positive selection of human naive and GC/M subpopulations, we characterized cytokine receptor moieties on these cells and analyzed their survival and growth in the presence of IL-4 or IL-10. Resting naive B cells expressed significantly higher IL-4 receptor alpha-chain on their cell surface than the combined GC/M subpopulation. The IL-10 receptor and the IL-2 receptor gammac chain were almost equally expressed on both subpopulations. When cultured in vitro, the addition of IL-4, but not IL-10, protected naive B cells from apoptosis in the absence of activation and growth. However, IL-4 exerted no such effect on resting GC/M B cells. These data support the hypothesis that IL-4 plays a pivotal role in the survival and maintenance of resting human naive B cells.     

Cytokine control of memory B cell homing machinery

The germinal center (GC) is a pivotal site for the development of B cell memory. Whereas GC B cells do not chemotax to most chemokines and do not express the adhesion receptors L-selectin, alpha(4)beta(7), and cutaneous lymphocyte Ag (CLA), memory B cells respond to various chemotactic signals and express adhesion receptors. In this study, we show that CD40 ligand, IL-2, and IL-10 together drive this transition of GC B cells to memory phenotype in vitro, up-regulating memory B cell markers, chemotactic responses to CXC ligand (CXCL)12, CXCL13, and CCL19, and expression of adhesion receptors L-selectin, alpha(4)beta(7), and CLA. Moreover, addition of IL-4 modulates this transition, preventing chemotactic responses to CXCL12 and CXCL13 (but not to CCL19), and inhibiting the re-expression of L-selectin, but not of CLA or alpha(4)beta(7). CCR7 expression, responsiveness to CCL19, and L-selectin/alpha(4)beta(7) phenotype are coordinately regulated. Thus, IL-2/IL-10 and IL-4 play important and distinctive roles in developing the migratory capacities of memory B cells.     

Subpopulations of B cells in germinal centers. III. HJ6, a monoclonal antibody, binds globoside and a subpopulation of germinal center B cells.

To identify surface Ag uniquely expressed on human germinal center B cells, we produced a mouse mAb, HJ6. When tonsillar lymphocytes were examined, HJ6 did not label T cells and labeled only about half of PNA+ B cells that were HK23-. HJ6 did not label mononuclear cells from peripheral blood, splenocytes, and any of 29 cell lines including 23 B cell lines. This binding pattern of HJ6 was very similar to that of a mAb named 5B5. It was shown previously that 5B5 bound a glycolipid named CTH (CD77) and its Ag was expressed on HK23- PNA+ tonsillar lymphocytes and Burkitt's lymphoma cell lines. Despite the similarity, HJ6 differed from 5B5: HJ6 did not stain Burkitt's lymphoma cell lines and stained PNA+ tonsillar lymphocytes in the presence of a large concentration of galactose. When its binding to isolated glycolipids was studied, HJ6 was found to bind globoside and Forssman Ag and not to other glycolipids including CTH. When its binding to neutral glycolipids extracted from tonsillar lymphocytes was studied, HJ6 bound only globoside; Forssman Ag was not detected in tonsillar lymphocytes. Taken together, we conclude that globoside is a B cell Ag expressed on a subpopulation of germinal center B cells.     

CD40 stimulation of human peripheral B lymphocytes: distinct response from naive and memory cells

During secondary immune response, memory B lymphocytes proliferate and differentiate into Ig-secreting cells. In mice, the binding of CD40 by CD154 clearly enhances the activation and differentiation of memory B lymphocytes. In humans, the role of CD40-CD154 in the stimulation of memory B lymphocytes is not as obvious since in vitro studies reported positive and negative effects on their proliferation and differentiation in Ig-secreting cells. In this study, we examine the response of peripheral memory and naive cells in relation to the duration of CD40-CD154 interaction. We measured the proliferation and differentiation of both subsets stimulated with CD154 and IL-4 for short- (4-5 days) and long-term (>7 days) periods. Following short-term stimulation, memory B lymphocytes did not expand but represented the only subset differentiating into IgG- and IgM-secreting cells. A longer stimulation of this population led to cell death, while promoting naive B lymphocyte proliferation, expansion, and differentiation into IgM- or IgG-secreting cells. This prolonged CD40 stimulation also triggered naive B lymphocytes to switch to IgG and to express CD27 even in absence of somatic hypermutation, suggesting that these latter events could be independent. This study suggests that naive and memory B lymphocytes have distinct requirements to engage an immune response, reflecting their different roles in humoral immunity.     

Expression, regulation, and function of B cell-expressed CD154 in germinal centers.

Activated B cells and T cells express CD154/CD40 ligand in vitro. The in vivo expression and function of B cell CD154 remain unclear and therefore were examined. Tonsillar B and T cells expressed CD154 at a similar density both in situ and immediately ex vivo, whereas a significantly higher percentage of the former expressed CD154. CD154-expressing B cells were most frequent in the CD38positiveIgD+ pre-germinal center (GC)/GC founder, CD38positive GC and CD38-IgD- memory populations, and were also found in the CD38-IgD+ naive and CD38brightIgD+ plasmablast subsets, but not in the CD38brightIgD- plasma cell subset. B cell expression of CD154 was induced by engaging surface Ig or CD40 by signals that predominantly involved activation of AP-1/NF-AT and NF-kappaB, respectively. The functional importance of CD154-mediated homotypic B cell interactions in vivo was indicated by the finding that mAb to CD154 inhibited differentiation of CD38positiveIgD- GC B cells to CD38-IgD- memory cells. In addition, mAb to CD154 inhibited proliferation induced by engaging sIg or CD40, indicating the role of up-regulation of this molecule in facilitating B cell responsiveness. Of note, CD154 itself not only functioned as a ligand but also as a direct signaling molecule as anti-CD154-conjugated Sepharose beads costimulated B cell responses induced by engaging surface Ig. These results indicate that CD154 is expressed by human B cells in vivo and plays an important role in mediating B cell responses.     

Bm1-Bm5 classification of peripheral blood B cells reveals circulating germinal center founder cells in healthy individuals and disturbance in the B cell subpopulations in patients with primary Sjögren's syndrome

Analyses of B cells in the bone marrow and secondary lymphoid tissues have revealed a broad range of cell surface markers defining B cell subpopulations, but only a few of these have been used to analyze B cell subpopulations in peripheral blood (PB). We report here the delineation of circulating PB B cell subpopulations by staining for CD19, CD38, and IgD in combination with CD10, CD44, CD77, CD95, CD23, IgM, and the B cell memory marker CD27. The utility of this approach is shown by the demonstration of disturbances of circulating B cell subpopulations in patients with autoimmune disease. Five mature B cell (Bm) subpopulations were identified in normal PB that were comparable with the tonsillar Bm1, Bm2, early Bm5, Bm5 subpopulations and, surprisingly, to the germinal center (GC) founder cell subpopulation (Bm2' and Bm3delta-4delta), suggesting that some GC founder cells are circulating. No PB B cells resembled the Bm3 and Bm4 GC cells. Remarkably, some cells with the CD38-IgD+ phenotype, previously known as naive Bm1 cells, expressed CD27. The CD38-IgD+ subpopulation therefore includes both naive Bm1 cells and IgD+ memory B cells. This new classification of B cell developmental stages reveals disturbances in the proportions of B cell subpopulations in primary Sj?gren's syndrome (pSS) patients compared with healthy donors and rheumatoid arthritis patients. Patients with pSS contained a significantly higher percentage of B cells in two activated stages, which might reflect a disturbance in B cell trafficking and/or alteration in B cell differentiation. These findings could be of diagnostic significance for pSS.     

Loss of IL-7 receptor alpha on CD4+ T cells defines terminally differentiated B cell-helping effector T cells in a B cell-rich lymphoid tissue

IL-7 plays important roles in development and homeostatic proliferation of lymphocytes. IL-7 uses a receptor composed of IL-7Ralpha (CD127) and the common gamma-chain (CD132) to transmit its signal. It has been unknown how CD127 is regulated during Th cell differentiation to the B cell-helping T cell lineage. In this study, we report that loss of CD127 defines terminally differentiated B cell-helping effector T cells in human tonsils. Although naive CD4(+) T cells uniformly express CD127, the memory/effector (non-FOXP3(+)) CD4(+) T cells are divided into CD127(+) and CD127(-) cells. The CD127(-) T cells are exclusively localized within the germinal centers where B cells become plasma and memory B cells, whereas CD127(+) T cells are found in T cell areas and the area surrounding B cell follicles. Consistently, the CD127(-) T cells highly express the B cell zone homing receptor CXCR5 with concomitant loss of CCR7. Compared with CD127(+) memory T cells, CD127(-) T cells have considerably shorter telomeres, do not proliferate in response to IL-7, and are prone to cell death. The CD127(-) T cells produce a large amount of the B cell follicle-forming chemokine CXCL13 upon stimulation with B cells and Ags. Most importantly, they are highly efficient in helping B cells produce Igs of all isotypes in a manner dependent on CD40L and ICOS and inducing activation-induced cytidine deaminase and Ig class switch recombination. The selective loss of CD127 on the B cell-helping effector T cells would have implications in regulation and termination of Ig responses.     

EBV persistence involves strict selection of latently infected B cells

EBV is found preferentially in IgD- B cells in the peripheral blood. This has led to the proposal that the recirculating memory B cell pool is the site of long-lived persistent infection. In this paper we have used CD27, a newly identified specific marker for memory B cells, to test this hypothesis. We show that EBV is tightly restricted in its expression. Less than 1 in 1000 of the infected cells in the peripheral blood are naive (IgD+, CD27-) and <1 in 250 are IgD+ memory cells. Furthermore, EBV was undetectable in the self-renewing peripheral CD5+ or B1 cells, a subset that has not been through a germinal center. No such restriction was observed in tonsillar B cells. Therefore, the virus has access to a range of B cell subsets in the lymph nodes but is tightly restricted to a specific long-lived compartment of B cells, the IgD-, CD27+, and CD5- memory B cells, in the periphery. We suggest that access to this compartment is essential to allow the growth-promoting latent genes to be switched off to create a site of persistent infection that is neither pathogenic nor a target for immunosurveillance.     

Follicular dendritic cells in vitro modulate the expression of Fas and Bcl-2 on germinal center B cells

Germinal center (GC) B cells are highly susceptible to apoptosis. The cellular mechanism regulating this sensitivity, however, has not yet been fully delineated. To investigate whether follicular dendritic cells (FDC) are capable of regulating the susceptibility to apoptosis of GC B cells, we constructed a GC model in vitro: emperipolesis of tonsillar B cells by FDC. We then analyzed the expressions of apoptosis-related proteins (Bcl-2 and Fas) on the cells by three-color flow cytometry. B cells nonentrapped by FDC decreased rapidly in number owing to early apoptosis in vitro, whereas entrapped B cells were rescued for at least 18 h and showed peculiar regulation of Fas and Bcl-2. GC founder cells (CD38+, IgD+; GCFC) and GC B cells (CD38+, IgD-) showed approximately a twofold increased expression of Fas; in contrast, mantle zone B cells (CD38-, IgD+) and memory B cells (CD38-, IgD-) showed no changes. Bcl-2 expression in mantle zone and memory B cells was reduced by approximately one-half; however, GCFC and GC B cells continued to express little Bcl-2 and this did not change. Our findings strongly suggest that FDC play a part in the modulation of the susceptibility to apoptosis on B cells within GC.     

Human interdigitating dendritic cells induce isotype switching and IL-13-dependent IgM production in CD40-activated naive B cells

Interdigitating dendritic cells (IDC) represent a mature progeny of dendritic cells (DC) in vivo and are exhibiting a strong lymphocyte stimulatory potential. Because of the restricted localization to secondary lymphoid organs where decisive cellular interactions take place in the initial events of immunity, IDC regulatory function was addressed in relation to naive B cells. In this study, we demonstrate that human tonsillar IDC induce a dual response from CD40-activated IgD+/CD38- naive B lymphocytes. IDC direct naive B cells toward either isotype switching or an IL-13-dependent IgM secretion. IDC-dependent proliferation, isotype switching, and Ig production are all strictly mediated by soluble factors, suggesting that such skewing in B cell activation is the result of differential cytokine expression. Moreover, IDC-expressed IL-13 represents a novel source of a cytokine with recently established effects in Th2 induction as well as in immunological disorders resulting in allergic reactions.     

Distinct response of human B cell subpopulations in recognition of an innate immune signal,CpG DNA

Innate immunity has recently gained renewed interest in its ability to regulate adaptive immunity. Among the innate immune signals, CpG DNA has revealed its potential as a vaccine adjuvant. However, the cellular mechanism for the effect of CpG DNA on the humoral immune response is not well understood. Here, we investigated the effects of CpG DNA on human B cell differentiation using highly purified B cell subsets: naive, germinal center (GC), and memory B cells. In the in vitro culture system that mimics the primary or secondary immune response in vivo, CpG DNA markedly augmented the proliferation and generation of plasma cells from naive and memory B cells. CpG DNA dramatically increased plasma cell generation from GC B cells. However, CpG DNA did not have effect on memory B cell generation from GC B cells. These results suggest that CpG DNA potentiates the B cell adaptive immune response by enhancing terminal differentiation, but does not affect the generation of memory B cells.     

IL-7R alpha gene expression is inversely correlated with cell cycle progression in IL-7-stimulated T lymphocytes

IL-7 plays a major role in T lymphocyte homeostasis and has been proposed as an immune adjuvant for lymphopenic patients. This prospect is based, at least in part, on the short-term expansion of peripheral T cells in rIL7-treated mice and primates. Nevertheless, in vivo, following initial increases in T cell proliferation and numbers, lymphocytes return to a quiescent state. As the bases for this cell cycle exit have not yet been elucidated, it is important to assess the long-term biological effects of IL-7 on quiescent human T lymphocyte subsets. In this study, we find that IL-7-stimulated CD4+ naive lymphocytes enter into cell cycle with significantly delayed kinetics as compared with the memory population. Importantly though, these lymphocytes exit from the cell cycle despite the continuous replenishment of rIL-7. This response is distinct in memory and naive CD4+ lymphocytes with memory cells starting to exit from cycle by day 10 vs day 18 for naive cells. Return to quiescence is associated with a cessation in IL-7R signaling as demonstrated by an abrogation of STAT-5 phosphorylation, despite an up-regulation of surface IL-7Ralpha. Indeed, an initial 10-fold decrease in IL-7Ralpha mRNA levels is followed by increased IL-7Ralpha expression in naive as well as memory T cells, with kinetics paralleling cell cycle exit. Altogether, our data demonstrate that IL-7 promotes the extended survival of both naive and memory CD4+ T cells, whereas cycling of these two subsets is distinct and transient. Thus, IL-7 therapy should be designed to allow optimal responsiveness of naive and memory T cell subsets.     

Intrinsic differences in the proliferation of naive and memory human B cells as a mechanism for enhanced secondary immune responses

Humoral immune responses elicited after secondary exposure to immunizing Ag are characterized by robust and elevated reactivity of memory B cells that exceed those of naive B cells during the primary response. The mechanism underlying this difference in responsiveness of naive vs memory B cells remains unclear. We have quantitated the response of naive and memory human B cells after in vitro stimulation with T cell-derived stimuli. In response to stimulation with CD40 ligand alone or with IL-10, both IgM-expressing and Ig isotype-switched memory B cells entered their first division 20-30 h earlier than did naive B cells. In contrast, the time spent traversing subsequent divisions was similar. Consistent with previous studies, only memory cells differentiated to CD38(+) blasts in a manner that increased with consecutive division number. These differentiated CD38(+) B cells divided faster than did CD38(-) memory B cell blasts. Proliferation of CD40 ligand-stimulated naive B cells as well as both CD38(+) and CD38(-) cells present in cultures of memory B cells was increased by IL-10. In contrast, IL-2 enhanced proliferation of CD38(-) and CD38(+) memory B cell blasts, but not naive cells. Thus, memory B cells possess an intrinsic advantage over naive B cells in both the time to initiate a response and in the division-based rate of effector cell development. These differences help explain the accelerated Ab response exhibited by memory B cells after secondary challenge by an invading pathogen, a hallmark of immunological memory.     

Kinetics of human B cell behavior and amplification of proliferative responses following stimulation with IL-21

Although recent studies indicated that IL-21 is an important regulator of human B cell activation, detailed comparison of the effects of IL-21 on distinct B cell subsets have not been performed. Our studies revealed that IL-21R is expressed by naive and germinal center B cells, but not memory or plasma cells. IL-21R was increased on naive and memory B cells following in vitro activation. Investigation into the kinetics and magnitude of responses of human B cells to IL-21 revealed that IL-21 potently augmented proliferation of CD40L-stimulated neonatal, splenic naive, and memory and tonsil germinal center B cells. This response exceeded that induced by IL-4, IL-10, and IL-13, cytokines that also induce B cell proliferation. Remarkably, CD40L/IL-21-stimulated naive B cells underwent the same number of divisions as memory cells and exhibited a greater enhancement in their response compared with CD40L alone than memory B cells. Therefore, IL-21 is a powerful growth factor for naive B cells. This may result from the higher expression of IL-21R on naive, compared with memory, B cells. Stimulation of human B cells with CD40L/IL-21 also induced IL-10 production and activation of STAT3. We propose that IL-21 may have therapeutic application in conditions of immunodeficiency where it could expand naive B cells, the predominant B cell subset in such patients. Conversely, because IL-21 is increased in murine models of lupus, dysregulated IL-21 production may contribute to perturbed B cell homeostasis observed in systemic lupus erythematosus. Thus, antagonizing IL-21 may be a novel strategy for treating Ab-mediated autoimmune diseases.     

Cytokine-mediated regulation of human B cell differentiation into Ig-secreting cells: predominant role of IL-21 produced by CXCR5+ T follicular helper cells

Differentiation of B cells into Ig-secreting cells (ISC) is critical for the generation of protective humoral immune responses. Because of the important role played by secreted Ig in host protection against infection, it is necessary to identify molecules that control B cell differentiation. Recently, IL-21 was reported to generate ISC from activated human B cells. In this study, we examined the effects of IL-21 on the differentiation of all human mature B cell subsets--neonatal, transitional, naive, germinal center, IgM-memory, and isotype-switched memory cells--into ISC and compared its efficacy to that of IL-10, a well-known mediator of human B cell differentiation. IL-21 rapidly induced the generation of ISC and the secretion of vast quantities IgM, IgG and IgA from all of these B cell subsets. Its effect exceeded that of IL-10 by up to 100-fold, highlighting the potency of IL-21 as a B cell differentiation factor. Strikingly, IL-4 suppressed the stimulatory effects of IL-21 on naive B cells by reducing the expression of B-lymphocyte induced maturation protein-1 (Blimp-1). In contrast, memory B cells were resistant to the inhibitory effects of IL-4. Finally, the ability of human tonsillar CD4+CXCR5+CCR7- T follicular helper (TFH) cells, known to be a rich source of IL-21, to induce the differentiation of autologous B cells into ISC was mediated by the production of IL-21. These findings suggest that IL-21 produced by TFH cells during the primary as well as the subsequent responses to T cell-dependent Ag makes a major contribution to eliciting and maintaining long-lived humoral immunity.     

Differential surface expression and possible function of 9-O- and 7-O-acetylated GD3 (CD60 b and c) during activation and apoptosis of human tonsillar B and T lymphocytes

The disialoganglioside GD3 (CD60 a) and its O-acetylated variants have previously been described as surface molecules of human T lymphocytes of the peripheral blood system. Here we report the expression of the 9-O-, and 7-O-acetylated disialoglycans of GD3 (CD60 b and CD60 c respectively) on human tonsillar lymphocytes. CD60 b and c are surface-expressed on activated germinal centre B cells and colocalize in raft-like structures on the cell surface together with the cytoplasmic tyrosine kinase Lyn and Syk. Addition of CD60 b and c mAb together with anti-IgM/IL-4 to in vitro cultivated tonsillar B cells resulted in a costimulatory effect. During spontaneous and staurosporine-induced apoptosis a distinct population of activated annexin V+/CD60 b+/CD60 c- B cells was observed. CD60 b and c are also found on cells of the extrafollicular T cell area. On tonsillar T cells, CD60 b mAb had a costimulatory effect together with PHA while CD60 c mAb alone was sufficient to induce proliferation. In further contrast to B cells, during apoptosis a distinct CD60 b+ T cell subpopulation was not observed. Together, surface-expressed CD60 b and c are differently expressed on tonsillar B and T cells and may be involved in the regulation of activation and apoptosis of lymphocytes in secondary lymphatic tissue.     

Regulation of Adhesion and Migration in the Germinal Center Microenvironment

T cell dependent humoral immune responses are initiated by the activation of naive B cells in the T cell areas of the secondary lymphoid tissues. This primary B cell activation leads to migration of germinal center (GC) cell precursors into B cell follicles where they engage follicular dendritic cells (FDC) and T cells, and differentiate into memory B cells or plasma cells. Both B cell homing to the GC and interaction with FDC critically depend on integrin-mediated adhesion. We have recently indentified the c-met-encoded receptor tyrosine kinase and its ligand, the growth and motility factor hepatocyte growth factor/scatter factor (HGF/SF), as a novel paracrine signalling pathway regulating B cell adhesion (van der Voort et al., 1997, J. Exp. Med. 185, 2121–2131). The c-Met protein is expressed on B cells localized in the dark zone of the GC (centroblasts) and is induced by CD40 plus BCR ligation. Stimulation of c-Met with HGF/SF. which is produced at high levels by tonsillar stromal cells and FDC, leads to receptor phosphorylation and to enhanced integrin-mediated adhesion of B cells to both VCAM-l and fibronectin. Interestingly, these responses to HGF/SF are promoted by heparan-sulfate proteoglycan forms of CD44 (CD44-HS). Like c-Met, CD44-HS is induced on B cells by CD40 ligation. It efficiently binds HGF/SF and strongly promotes signalling through c-Met. We conclude that integrin regulation during antigen specific B cell differentiation involves cross-talk between the HGF/SF-c-Met pathway and CD44-HS.     

The VP6 protein of rotavirus interacts with a large fraction of human naive B cells via surface immunoglobulins

Immunity to human group A rotavirus (RV), a major cause of viral gastroenteritis in infants, involves B lymphocytes that provide RV-specific antibodies. Additionally, some arguments suggest that naive B cells could be implicated in the first steps of the immune response against RV. The aim of our study was to analyze the interaction of VP6 and VP7 RV capsid proteins with human B cells depending on the immune status of the individual, i.e., naive or RV experienced. For this purpose, a two-color virus-like particle flow cytometry assay was devised to evaluate the blood B-lymphocyte reactivity to VP6 and VP7 proteins from healthy RV-exposed adults, recently infected infants, and neonates at birth. Both VP6 and VP7 interactions with B cells were mediated by surface immunoglobulins and probably by their Fab portions. VP7-reactive B lymphocytes were mainly detected from RV-experienced patients and almost exclusively in the CD27-positive memory cell fraction. Conversely, VP6-reactive B lymphocytes were detected at similar and high frequencies in adult, infant, and neonate samples. In adult samples, VP6 reacted with about 2% of the CD27-negative (CD27(neg)) naive B cells. These results demonstrated that the VP6 RV protein interacted with a large fraction of naive B lymphocytes from both adults and neonates. We propose that naive B cell-VP6 interaction might influence the strength and quality of the acquired immune response and should be considered for elaborating RV vaccine strategies.