Mac-1-negative B-1b phenotype of natural antibody-producing cells, including those responding to Gal alpha 1,3Gal epitopes in alpha 1,3-galactosyltransferase-deficient mice

Human natural Abs against Galalpha1-3Galbeta1-4GlcNAc (Gal) epitopes are a major barrier to xenotransplantation. Studies in this report, which use combined multiparameter flow cytometric sorting and enzyme-linked immunospot assay, demonstrate that anti-Gal IgM-producing cells are found exclusively in a small B cell subpopulation (i.e., CD21(-/low) IgM(high) B220(low) CD5(-) Mac-1(-) 493(-) cells) in the spleens of alpha1, 3-galactosyltransferase-deficient mice. All IgM-producing cells were detected in a similar splenic subpopulation of alpha1, 3-galactosyltransferase-deficient and wild-type mice. A higher frequency of B cells with anti-Gal surface IgM receptors was observed in the peritoneal cavity than in the spleen, but these did not actively secrete Abs, and showed phenotypic properties of B-1b cells (CD21(-/low) IgM(high) CD5(-) CD43(+) Mac-1(+)). However, these became Mac-1(-) and developed anti-Gal Ab-producing activity after in vitro culture with LPS. The splenic B cells with anti-Gal receptors consisted of both Mac-1(+) B-1b cells and Mac-1(-) B-1b-like cells. The latter comprised most anti-Gal IgM-producing cells. Our studies indicate that anti-Gal natural IgM Abs are produced by a B1b-like, Mac-1(-) splenic B cell population and not by plasma cells or B-1a cells. They are consistent with a model whereby B-1b cells lose Mac-1 expression upon Ag exposure and that these, rather than plasma cells, become the major IgM Ab-producing cell population.     

Peritoneal cavity B cells are precursors of splenic IgM natural antibody-producing cells

Peritoneal cavity B-1 cells are believed to produce IgM natural Abs. We have used alpha1,3-galactosyltransferase-deficient (GalT(-/-)) mice, which, like humans, produce IgM natural Abs against the carbohydrate epitope Galalpha1,3Gal (Gal), to demonstrate that peritoneal cavity B-1b cells with anti-Gal receptors produce anti-Gal IgM Abs only after LPS stimulation. Likewise, peritoneal cavity cells of GalT(-/-) and wild-type mice do not produce IgM Abs of other specificities without LPS stimulation. Development of Ab-secreting capacity is associated with loss of CD11b/CD18 (Mac-1) expression. In contrast, there are large numbers of cells producing anti-Gal and other IgM Abs in fresh splenocyte preparations from GalT(-/-) and (for non-Gal specificities) wild-type mice. These cells are Mac-1(-) but otherwise B-1b-like in their phenotype. We therefore hypothesized a pathway wherein peritoneal cavity B cells migrate into the spleen after activation in vivo and lose Mac-1 expression to become IgM Ab-producing cells. Consistent with this possibility, splenectomy reduced anti-Gal Ab production after immunization of GalT(-/-) mice with Gal-positive rabbit RBC. Furthermore, splenectomized B6 GalT(-/-), Ig micro -chain mutant ( micro (-/-)) (both Gal- and B cell-deficient) mice produced less anti-Gal IgM than nonsplenectomized controls after adoptive transfer of peritoneal cavity cells from B6 GalT(-/-) mice. When sorted GalT(-/-) Mac-1(+) peritoneal cavity B cells were adoptively transferred to B6 GalT(-/-), micro (-/-) mice, IgM Abs including anti-Gal appeared, and IgM-producing and Mac1(-) B cells were present in the spleen 5 wk after transfer. These findings demonstrate that peritoneal cavity Mac-1(+) B-1 cells are precursors of Mac-1(-) splenic IgM Ab-secreting cells.     

Association of B-1 B cells with follicular dendritic cells in spleen

Although CD5(+) B-1 B cells have been recognized as an infrequent B cell subset in mice for many years, attempts to identify their histologic location in normal mouse spleen have proven difficult due to both their paucity and low level expression of CD5. In this study we have studied V(H)11/D(H)/J(H) gene-targeted mice, V(H)11t, that develop elevated numbers of CD5(+) V(H)11/V(k)9 B cells with an anti-phosphatidylcholine (anti-PtC) autoreactive specificity, allowing B-1 B cell detection by anti-PtC Id-specific Abs in spleen section staining. Using this approach we found that anti-PtC B-1 cells first appear within the white pulp in neonates, expand in association with follicular dendritic cells (FDC), and localize more centrally than other (non-B-1) IgD(high) follicular B cells in adults. Among neonatal B cells, CD5(+) B-1 cells in both normal and V(H)11t mouse spleen and peritoneal cavity express the highest levels of CXCR5, which is important for FDC development. Injection of purified spleen or peritoneal B-1 cells into RAG knockout mice resulted in B-1 cell follicle formation in spleen, inducing FDC development and plasma cell generation. These results indicate that B-1 B cells are the first B cells to express fully mature levels of CXCR5, thereby promoting the development of FDC.     

Functional activity of natural antibody is altered in Cr2-deficient mice

The major source of natural IgM Abs are B-1 cells, which differ from conventional B cells in their anatomic location, cell surface phenotype, restricted usage of particular V(H) genes and limited use of N-region addition during V-D-J rearrangement. The origin of B-1 cells is unclear. However, they are capable of self-renewal and their development is sensitive to signaling via the B cell receptor, as genetic defects that impair the strength of the signal often result in limited development. These findings suggest that B-1 cells require either an intrinsic signal, or contact with Ag, for positive selection and expansion and/or maintenance in the periphery. In support of interaction with cognate Ag, deficiency in the complement receptors CD21/CD35 results in a 30-40% decrease in the CD5(+) B-1 population. To determine whether this reduction reflects a loss of certain specificities or simply a proportional decline in the repertoire, we examined peritoneal B cells isolated from Cr2(+) and Cr2(def) mice for recognition of a B-1 cell Ag, i.e., phosphatidylcholine, and assayed for injury in an IgM natural Ab-dependent model of reperfusion injury. We found a similar frequency of phosphatidylcholine-specific CD5(+) B-1 cells in the two strains of mice. By contrast, the Cr2(def) mice have reduced injury in the IgM-dependent model of reperfusion injury. Reconstitution of the deficient mice with pooled IgM or adoptive transfer of Cr2(+) peritoneal B cells restored injury. These results suggest that complement receptors CD21/CD35 are important in maintenance of the B-1 cell repertoire to some, but not all, specificities.     

Characterization of the immunodeficiency of RIIS/J [corrected] mice. I. Association with the CD5 (LY-1) [corrected] B cell lineage

RIIIS/J mice produce low antibody responses to several polysaccharide Ag of bacterial origin. They have low levels of serum IgM and IgG3 and high levels of serum IgG2a and IgG2b. Low serum IgM and IgG3 have been attributed to a low frequency of CD5 (Ly-1) B cells, which play an important role in the production of natural antibodies. Indeed, RIIIS/J mice have a low frequency of CD5 (Ly-1)+, IgM bright+, Ly-5 (B220)dull+ (i.e., CD5 (Ly-1) B) cells in their peritoneum. RIIIS/J mice treated with LPS produce a low anti-bromelain-treated mouse RBC splenic plaque-forming cell response and a normal anti-mouse transferrin splenic PFC response. Those data are compatible with the fact that CD5 (Ly-1) B cells contain the precursors of B lymphocytes secreting anti-bromelain-treated mouse RBC antibody. However, they have a higher frequency of IgM bright+, Mac-1+ cells in their peritoneum. These cells represent the CD5 (Ly-1) "sister population" of CD5 (Ly-1) B cells described by others. This suggests that characteristics usually associated with the CD5 (Ly-1) lineage are applicable only to the CD5 (Ly-1)+ Mac-1+ IgM+ population, but not the related CD5 (Ly-1)- Mac-1+ IgM+ population. RIIIS/J mice should thus prove a valuable model to study the CD5 (Ly-1) B cell lineage.     

Hepatitis C virus drives the unconstrained monoclonal expansion of VH1-69-expressing memory B cells in type II cryoglobulinemia: a model of infection-driven lymphomagenesis

Chronic hepatitis C virus infection causes B cell lymphoproliferative disorders that include type II mixed cryoglobulinemia and lymphoma. This virus drives the monoclonal expansion and, occasionally, the malignant transformation of B cells producing a polyreactive natural Ab commonly encoded by the V(H)1-69 variable gene. Owing to their property of producing natural Ab, these cells are reminiscent of murine B-1 and marginal zone B cells. We used anti-Id Abs to track the stages of differentiation and clonal expansion of V(H)1-69(+) cells in patients with type II mixed cryoglobulinemia. By immunophenotyping and cell size analysis, we could define three discrete stages of differentiation of V(H)1-69(+) B cells: naive (small, IgM(high)IgD(high)CD38(+)CD27(-)CD21(high)CD95(-)CD5(-)), "early memory" (medium-sized, IgM(high)IgD(low)CD38(-)CD27(+)CD21(low)CD95(+)CD5(+)), and "late memory" (large-sized, IgM(low)IgD(low-neg)CD38(-)CD27(low)CD21(low-neg)CD5(-)CD95(-)). The B cells expanded in cryoglobulinemia patients have a "memory" phenotype; this fact, together with the evidence for intraclonal variation, suggests that antigenic stimulation by hepatitis C virus causes the unconstrained expansion of activated V(H)1-69(+) B cells. In some cases, these cells replace the entire pool of circulating B cells, although the absolute B cell number remains within normal limits. Absolute monoclonal V(H)1-69(+) B lymphocytosis was seen in three patients with cryoglobulinemia and splenic lymphoma; in two of these patients, expanded cells carried trisomy 3q. The data presented here indicate that the hepatitis C virus-driven clonal expansion of memory B cells producing a V(H)1-69(+) natural Ab escapes control mechanisms and subverts B cell homeostasis. Genetic alterations may provide a further growth advantage leading to an overt lymphoproliferative disorder.     

IL-9-induced expansion of B-1b cells restores numbers but not function of B-1 lymphocytes in xid mice

Mice expressing the X-linked immunodeficiency (xid) mutation lack functional Bruton's tyrosine kinase and were shown to be specifically deficient in peritoneal B-1 lymphocytes. We have previously shown that IL-9, a cytokine produced by TH2 lymphocytes, promotes B-1 cell expansion in vivo. To determine whether IL-9 overexpression might compensate the xid mutation for B-1 lymphocyte development, we crossed xid mice with IL-9-transgenic mice. In this model, IL-9 restored normal numbers of mature peritoneal B-1 cells that all belonged to the CD5(-) B-1b subset. Despite this normal B-1 lymphocyte number, IL-9 failed to restore classical functions of B-1 cells, namely, the production of natural IgM Abs, the T15 Id Ab response to phosphorylcholine immunization, and the antipolysaccharide humoral response against Streptococcus pneumoniae. By using bromelain-treated RBC, we showed that the antigenic repertoire of these IL-9-induced B-1b lymphocytes was different from the repertoire of classical CD5(+) B-1a cells, indicating that the lack of B-1 function by B-1b cells is associated with distinct Ag specificities. Taken together, our data show that B-1b cell development can restore the peritoneal B-1 population in xid mice but that these B-1b cells are functionally distinct from CD5(+) B-1a lymphocytes.     

TLR9 in peritoneal B-1b cells is essential for production of protective self-reactive IgM to control Th17 cells and severe autoimmunity

The role of TLR9 in the development of the autoimmune disease systemic lupus erythematosus is controversial. In different mouse models of the disease, loss of TLR9 abolishes the generation of anti-nucleosome IgG autoantibodies but at the same time exacerbates lupus disease. However, the TLR9-dependent tolerance mechanism is unknown. In this study, we show that loss of TLR9 is associated with low peritoneal B-1b cell numbers and low levels of protective self-reactive IgM serum autoantibodies in lupus-prone FcγRIIB-deficient mice leading to the uncontrolled accumulation of proinflammatory CD4(+) cells and exacerbated autoimmunity. TLR7 signaling was not able to compensate for the loss of TLR9 signaling in peritoneal B-1b cells to induce IgM Abs. Transfer of TLR9-expressing peritoneal B-1b cells from FcγRIIB-deficient mice or of recombinant monoclonal self-reactive IgM Abs was sufficient to reduce the frequency of proinflammatory Th17 cells and lupus disease in FcγRIIB/TLR9 double-deficient mice. Taken together, these data provide evidence for a TLR9-dependent tolerance mechanism of peritoneal B-1b cells generating protective self-reactive IgM in lupus-prone mice to control Th17 cell development and severe autoimmunity.     

Identification of initiator B cells, a novel subset of activation-induced deaminase-dependent B-1-like cells that mediate initiation of contact sensitivity

Contact sensitivity (CS) is related to delayed-type hypersensitivity and is a well-characterized prototype of T cell-mediated inflammation. However, the inflammatory response associated with CS is additionally dependent on Ag-specific IgM produced by a subpopulation of B cells in response to sensitization. Upon re-exposure to hapten, this IgM mediates rapid vascular activation and subsequent recruitment of proinflammatory T cells to the local site. Interference with this pathway prevents the full development of the classic delayed inflammatory response and is therefore termed the "CS initiation" pathway. In this study, we show that CS initiation is defective in mice deficient in activation-induced deaminase, an enzyme central to the process of somatic hypermutation. Using adoptive transfer experiments, we demonstrate that the defect is specific to a B-1-like population of B cells and that transfer of WT cells reconstitutes CS initiation mechanisms in deficient recipients. We went on to identify a novel subpopulation of Ag-binding B cells in the spleens of sensitized mice that possess initiation activity (CD19(+)CD5(+)Thy-1(int)IgM(high)IgD(high)) that we name "initiator B cells." Analysis of BCR H chain genes isolated from these cells revealed evidence of activation-induced deaminase-mediated somatic hypermutation. The sensitivity of CS initiation to very low amounts of sensitizing hapten suggests that the responsible B cells have increased IgM receptor gene mutations enabling selection to generate Abs with sufficient affinity to mediate the response.     

Transgenic expression of Bcl-xL or Bcl-2 by murine B cells enhances the in vivo antipolysaccharide, but not antiprotein, response to intact Streptococcus pneumoniae

IgG antipolysaccharide (PS) and antiprotein responses to Streptococcus pneumoniae (Pn) are both CD4(+) T cell dependent. However, the primary IgG anti-PS response terminates more quickly, uses a shorter period of T cell help, fails to generate memory, and is more dependent on membrane Ig (mIg) signaling. We thus determined whether this limited anti-PS response to Pn reflected a greater propensity of PS-specific B cells to undergo apoptosis. We used mice that constitutively expressed the antiapoptotic protein Bcl-x(L) or Bcl-2 as a B cell-specific transgene. Both transgenic (Tg) mice exhibited increased absolute numbers of splenic B-1 and peritoneal B-1b and B-2 cells, subsets implicated in anti-PS responses, but not in marginal zone B (MZB) cells. Both Tg mouse strains elicited, in an apparently Fas-independent manner, a more prolonged and higher peak primary IgM and IgG anti-PS, but not antiprotein, response to Pn, but without PS-specific memory. A similar effect was not observed using purified PS or pneumococcal conjugate vaccine. In vitro, both splenic MZB and follicular Tg B cells synthesized DNA at markedly higher levels than their wild-type counterparts, following mIg cross-linking. This was associated with increased clonal expansion and decreased apoptosis. Using Lsc(-/-) mice, the Pn-induced IgG response specific for the capsular PS was found to be almost entirely dependent on MZB cells. Collectively, these data suggest that apoptosis may limit mIg-dependent clonal expansion of PS-specific B cells during a primary immune response to an intact bacterium, as well as decrease the pool of PS-responding B cell subsets.     

Human B cell development. II. Subpopulations in the human fetus

In man, during fetal development the B cell populations show distinct phenotypes at different tissue sites. The pre-B and B lymphocytes of the fetal liver and bone marrow express IgM and B cell markers, B1 (CD20) and BA-1 (CD24). These "early" cells are negative with a number of other reagents, anti-IgD, RFB4 (CD22), RFB6 (CD21), and RFA-2, which on the other hand recognize peripheral B cells. These peripheral B lymphocytes in the developing fetus are heterogeneous. The diffusely distributed B cells in the earliest lymph node samples, 16 to 17 wk of gestational age, and from 16 to 21 wk in the spleen, are strongly IgM+ (IgD+,RFB4+,RFB6+, and RFA-2+) but lack T cell-associated markers such as T1 (CD5, p 67,000 dalton equivalent of murine Ly-1) and Tü-33. In fetal lymph nodes, primary nodules develop around the follicular dendritic (FD) cells from 17 wk onward, and contain a virtually pure population of B cells; B1+,BA1+,RFB4+,RFB6+,RFA-2+, which simultaneously express IgM,IgD together with T1 (CD5), a T cell-associated antigen. A sizeable subpopulation of these IgM+,T1+ cells are also positive for Tü-33, another T cell-associated marker. In the spleen, the B cells of the IgM+,IgD+,T1+ type appear in smaller numbers and only relatively late around wk 22. These cells are diffusely distributed at first, and start accumulating around the small FD cell clusters as soon as these emerge about the 23rd gestational wk. At that time, the IgM+,T1+B cells can also be washed out from the peritoneal and pleural cavities. The T1+,IgM+B cells may represent the normal equivalent cells of B chronic lymphoid leukemia and centrocytic lymphoma, and appear to be the counterpart of Ly-1+,IgM+B cells in the mouse.     

A population of CD19highCD45R-/lowCD21low B lymphocytes poised for spontaneous secretion of IgG and IgA antibodies

Ab responses to selected Ags are produced by discrete B cell populations whose presence and functional relevance vary along the ontogeny. The earliest B lineage-restricted precursors in gestational day 11 mouse embryos display the CD19(+)CD45R/B220(-) phenotype. Phenotypically identical cells persist throughout gestation and in postnatal life, in parallel to the later-arising, CD19(+)CD45R(+) B cells. Very early after birth, the CD19(+)CD45R(-) B cell subset included high frequencies of spontaneously Ig-secreting cells. In the adult spleen, a small subset of CD19(high)CD45R(-/low)IgM(+/-)IgD(-)CD21/Cr2(-/low) cells, which was detected in perifollicular areas, displayed genetic and phenotypical traits of highly differentiated B cells, and was enriched in IgG- and IgA-secreting plasma cells. In vitro differentiation and in vivo adoptive transfer experiments of multipotent hemopoietic progenitors revealed that these CD19(high)CD45R(-/low) B cells were preferentially regenerated by embryo-, but not by adult bone marrow-, derived progenitors, except when the latter were inoculated into newborn mice. Both the early ontogenical emergence and the natural production of serum Igs, are shared features of this CD19(high)CD45R(-/low) B cell population with innate-like B lymphocytes such as B1 and marginal zone B cells, and suggest that the new population might be related to that category.     

Combination of 3' and 5' IgH regulatory elements mimics the B-specific endogenous expression pattern of IgH genes from pro-B cells to mature B cells in a transgenic mouse model

To ensure the B cell differentiation stage specificity of the intronic Emu element and of the locus control region (LCR) that lies downstream of the IgH chain locus, we generated transgenic mice harboring a V(H) promoter-GFP reporter gene linked to the 3'LCR region and the Emu element. By flow cytometry, GFP(+) lymphocytes were observed amongst pro-B cells (B220(+)CD43(+)CD117(+)) and at all stages of differentiation up to mature B cells (B220(+)IgM(+)IgD(+)). Expression was strictly confined to cells committed to the B lymphocyte lineage as judged by the lack of GFP(+)Thy1,2(+) cells (T lymphocytes) and GFP(+)B220(-)CD117(+)CD43(+) cells (uncommitted lymphohematopoietic progenitors). Therefore, the Emu-GFP-3'LCR transgene is not expressed by hematopoietic stem cells, begins its expression in pro-B cells and is specifically active at all stages of B cell maturation. The combination of 3' and 5' IgH regulatory elements thus appears as a potentially useful cassette in transgenes that require a stringent and early B lineage-specific expression.     

Isolation, characterization, and expression of cDNA clones encoding the mouse Fc receptor for IgE (Fc epsilon RII)1

We have isolated cDNA clones encoding a mouse low affinity receptor for IgE (Fc epsilon RII) from a cDNA library of BALB/c splenic B cells activated with LPS and IL-4. The 2.2-kb cDNA clone encodes a 331 amino acid membrane glycoprotein that is homologous to human Fc epsilon RII (CD23) and a family of carbohydrate-binding proteins. COS7 cells transfected with the cDNA clones expressed a 45,000 m.w. protein that bound IgE and the anti-Fc epsilon RII mAb, B3B4. Fc epsilon RII mRNA was up-regulated in mouse B cells by culture with IL-4, but not in B cells cultured with IgE. Fc epsilon RII mRNA was detected in IgM+/IgD+ B cell lines, but not in pre-B cell lines or in B cell lines which have undergone differentiation to secrete Ig. The monocyte line P388D1, mast cell lines MC/9 and PT18, and peritoneal macrophages stimulated with IL-4 lacked detectable Fc epsilon RII mRNA, as did Thy-1.2+, CD4+, and CD8+ normal T cells and Thy-1.2+ T cells from Nippostrongylus brasiliensis-infected mice.     

Mechanisms of peritoneal B-1a cells accumulation induced by murine lupus susceptibility locus Sle2

The abundance of B-1a cells found in the peritoneal cavity of mice is under genetic control. The lupus-prone mouse New Zealand Black and New Zealand White (NZB x NZW)F(1) and its derivative NZM2410 are among the strains with the highest numbers of peritoneal B1-a cells. We have previously identified an NZM2410 genetic locus, Sle2, which is associated with the production of large numbers of B-1a cells. In this paper, we examined the mechanisms responsible for this phenotype by comparing congenic C57BL/6 mice with or without Sle2. Fetal livers generated more B-1a cells in B6.Sle2 mice, providing them with a greater starting number of B-1a cells early in life. Sle2-expressing B1-a cells proliferated significantly more in vivo than their B6 counterparts, and reciprocal adoptive transfers showed that this phenotype is intrinsic to Sle2 peritoneal B cells. The rate of apoptosis detected was significantly lower in B6.Sle2 peritoneal cavity B-1a cells than in B6, with or without exogenous B cell receptor cross-linking. Increased proliferation and decreased apoptosis did not affect Sle2 peritoneal B-2 cells. In addition, a significant number of peritoneal cavity B-1a cells were recovered in lethally irradiated B6.Sle2 mice reconstituted with B6.Igh(a) bone marrow, showing radiation resistance in Sle2 B-1a cells or its precursors. Finally, B6.Sle2 adult bone marrow and spleen were a significant source of peritoneal B-1a cells when transferred into B6.Rag2(-/-) mice. This suggests that peritoneal B-1a cells are replenished throughout the animal life span in B6.Sle2 mice. These results show that Sle2 regulates the size of the B-1a cell compartment at multiple developmental checkpoints.     

B-1 B cells mediate required early T cell recruitment to elicit protein-induced delayed-type hypersensitivity

We define the initiation of elicited delayed-type hypersensitivity (DTH) as a series of processes leading to local extravascular recruitment of effector T cells. Responses thus have two sequential phases: 1) 2-h peaking initiation required for subsequent recruitment of T cells, and 2) the late classical 24-h component mediated by the recruited T cells. We analyzed DTH initiation to protein Ags induced by intradermal immunization without adjuvants. Ag-spceific initiating cells are present by 1 day in spleen and lymph nodes. Their phenotypes, determined by depletion of cell transfers by mAb and complement, are CD5(+), CD19(+), CD22(+), B220(+), Thy1(+), and Mac1(+), suggesting that they are B-1 B cells. DTH initiation is absent in micro MT B cell and xid B-1 cell deficient mice, is impaired in mice unable to secrete IgM, and is reconstituted with 1 day immune serum, suggesting that early B-1 cell-derived IgM is responsible. Study of complement C5a receptor-deficient mice, anti-C5 mAb neutralization, or mast cell deficiency suggests that DTH initiation depends on complement and mast cells. ELISPOT assay confirmed production of Ag-specific IgM Abs at days 1 and 4 in wild-type mice, but not in B-1 cell-deficient xid mice. We conclude that rapidly activated B-1 cells produce specific IgM Abs which, after local secondary skin challenge, form Ag-Ab complexes that activate complement to generate C5a. This stimulates C5a receptors on mast cells to release vasoactive substances, leading to endothelial activation for the 2-h DTH-initiating response, allowing local recruitment of DTH-effector T cells.     

The resolution of relapsing fever borreliosis requires IgM and is concurrent with expansion of B1b lymphocytes

The rate of pathogen clearance is a critical determinant of morbidity and mortality. We sought to characterize the immune response responsible for the remarkably rapid clearance of individual episodes of bacteremia caused by the relapsing fever bacterium, Borrelia hermsii. SCID or Rag(-/-) mice were incapable of resolving B. hermsii infection, indicating a critical role for T and/or B cells. TCR(-/-) mice, which lack T cells, and IL-7(-/-) mice, which are deficient in both T cells and follicular B cells, but not in B1 cells and splenic marginal zone (MZ) B cells, efficiently cleared B. hermsii. These findings suggested that B1 cells and/or MZ B cells, two B cell subsets that are known to participate in rapid, T-independent responses, might be involved. The efficient resolution of the episodes of moderate level bacteremia by splenectomized mice suggested that MZ B cells do not play the primary role in clearance of this bacterium. In contrast, xid mice, which are deficient in B1 cells, suffered more severe episodes of bacteremia than wild-type mice. The hypothesis that B1 cells are critical for clearance of B. hermsii was further supported by a selective expansion of the B1b (i.e., IgM(high), IgD(-/low), Mac1(+) CD23(-), and CD5(-)) cell subset in infected xid mice, which coincided with the eventual resolution of infection. Finally, mice selectively incapable of secreting IgM, the dominant isotype produced by B1 cells, were completely unable to clear B. hermsii. Together these results support the model that B1b cells generate the T-independent IgM required for the control and resolution of relapsing fever borreliosis.     

Transgenic overexpression of G5PR that is normally augmented in centrocytes impairs the enrichment of high-affinity antigen-specific B cells, increases peritoneal B-1a cells, and induces autoimmunity in aged female mice

To investigate signals that control B cell selection, we examined expression of G5PR, a regulatory subunit of the serine/threonine protein phosphatase 2A, which suppresses JNK phosphorylation. G5PR is upregulated in activated B cells, in Ki67-negative centrocytes at germinal centers (GCs), and in purified B220(+)Fas(+)GL7(+) mature GC B cells following Ag immunization. G5PR rescues transformed B cells from BCR-mediated activation-induced cell death by suppression of late-phase JNK activation. In G5PR-transgenic (G5PR(Tg)) mice, G5PR overexpression leads to an augmented generation of GC B cells via an increase in non-Ag-specific B cells and a consequent reduction in the proportion of Ag-specific B cells and high-affinity Ab production after immunization with nitrophenyl-conjugated chicken γ-globulin. G5PR overexpression impaired the affinity-maturation of Ag-specific B cells, presumably by diluting the numbers of high-affinity B cells. However, aged nonimmunized female G5PR(Tg) mice showed an increase in the numbers of peritoneal B-1a cells and the generation of autoantibodies. G5PR overexpression did not affect the proliferation of B-1a and B-2 cells but rescued B-1a cells from activation-induced cell death in vitro. G5PR might play a pivotal role in B cell selection not only for B-2 cells but also for B-1 cells in peripheral lymphoid organs.