B1 cells: similarities and differences with other B cell subsets

Since their discovery, B1 B cells' origins and developmental pathways have eluded characterization. In the past year, focus on B1 B cells has shifted dramatically from developmental to functional aspects of these cells. Most advances have been made in describing the physiological activities of B1 cells, including their migration, activation by antigen and role in both autoimmunity and malignancy.     

Dendritic cell type 3 arises from Ly6C+ monocyte-dendritic cell progenitors

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脂多糖诱导小鼠内皮损伤早期T细胞亚群的变化

目的:探讨脂多糖(LPS)诱导内皮损伤早期T细胞亚群的变化。方法:采用扫描电镜观察LPS刺激后内膜的形态学变化,流式细胞术检测外周血中内皮细胞表型CD62P+、CD62P+CD62E+改变,采用胞内外因子染色法检测T细胞亚群的肿瘤坏死因子-α(TNF-α)、γ-干扰素(IFN-γ)、白细胞介素17(IL-17)及白细胞介素10(IL-10)的分泌状态。结果:LPS可导致小鼠血管内皮完整性受损,脱落内皮细胞CD62P+、CD62P+CD62E+比例明显高于对照组(P<0.05);内皮损伤的同时,小鼠CD3+CD4+T细胞中,TNF-α、IFN-γ、IL-17分泌细胞比例显著增加(P<0.05),而IL-10的分泌在LPS刺激1小时、4小时时升高较为明显(P<0.05)。结论:炎性CD4+T细胞亚群与脱落内皮细胞CD62P、CD62E的高表达状态是血管内膜损伤早期的重要特征性标志。     

Modulation of B and T cell subsets in mice treated with fractionated total lymphoid irradiation. II. Tolerance susceptibility of B cell subsets

Total lymphoid irradiation (TLI) results in long-lasting changes in the characteristics of both T and B cells suggestive of arrested maturation. A characteristic feature of immature B cells is their high susceptibility to tolerance induction. This study examines the susceptibility to tolerance to bovine serum albumin (BSA) of TLI-treated mice. Two experimental protocols were designed. In the first, tolerance to BSA was induced in TLI-treated adult (BALB/c × C57BL/6)F₁ mice, and the ability of B cells of those mice to respond to BSA was assessed in an adoptive transfer system. In the second experimental protocol, tolerance was induced in adoptive hosts reconstituted with purified B cells originating from TLI-treated mice and with splenic T cells of normal, untreated mice. Results obtained in these two systems clearly demonstrated that splenic B cells of TLI-treated mice are highly susceptible to tolerance induction. This high susceptibility of B cells is linked neither with the elevation of immature T cells nor with induced T suppressor cells which arise due to the long-term malfunction of the thymus. Tolerance could be induced in TLI cells even 4 months after termination of the treatment. Thus, maturation processes of B cells in TLI-treated mice are arrested for long periods of time.     

Mono-allelic Ly49 NK cell receptor expression.

Each cell is equipped with two copies (alleles) of each autosomal gene. While the vast majority use both alleles, occasional genes are expressed from a single allele. The reason for mono-allelic expression is not always evident and can serve distinct purposes. First, it may facilitate the tight control over the dosage of certain gene products such as some growth factors and their receptors or X-linked genes. Second, the differential usage of the two parental alleles may reflect the mechanisms that ensure mono-specificity, e.g. olfactory receptors, T and B cell receptors. The context of allele-specific expression of the murine Ly49 natural killer (NK) cell receptor genes suggests that their allele-specific expression reflects a process that generates clonal variability.     

TCL1 oncogene expression in B cell subsets from lymphoid hyperplasia and distinct classes of B cell lymphoma

Activation of the TCL1 oncogene has been implicated in T cell leukemias/lymphomas and recently was associated with AIDS diffuse large B cell lymphomas (AIDS-DLBCL). Also, in nonmalignant lymphoid tissues, antibody staining has shown that mantle zone B cells expressed abundant Tcl1 protein, whereas germinal center (GC; centrocytes and centroblasts) B cells showed markedly reduced expression. Here, we analyze isolated B cell subsets from hyperplastic tonsil to determine a more precise pattern of Tcl1 expression with development. We also examine multiple B cell lines and B lymphoma patient samples to determine whether different tumor classes retain or alter the developmental pattern of expression. We show that TCL1 expression is not affected by Epstein-Barr virus (EBV) infection and is high in na?ve B cells, reduced in GC B cells, and absent in memory B cells and plasma cells. Human herpesvirus-8 infected primary effusion lymphomas (PEL) and multiple myelomas are uniformly TCL1 negative, whereas all other transformed B cell lines tested express moderate to abundant TCL1. This observation supports the hypothesis that PEL, like myeloma, usually arise from post-GC stages of B cell development. Tcl1 protein is also detected in most na?ve/GC-derived B lymphoma patient samples (23 of 27 [85%] positive), whereas most post-GC-derived B lymphomas lack expression (10 of 41 [24%] positive). These data indicate that the pattern of Tcl1 expression is distinct between na?ve/GC and post-GC-derived B lymphomas (P < 0.001) and that the developmental pattern of expression is largely retained. However, post-GC-derived AIDS-DLBCL express TCL1 at a frequency equivalent to na?ve/GC-derived B lymphomas in immune-competent individuals (7 of 9 [78%] positive), suggesting that TCL1 down-regulation is adversely affected by severe immune system dysfunction. These findings demonstrate that TCL1 expression in B cell lymphoma usually reflects the stage of B cell development from which they derive, except in AIDS-related lymphomas.     

Ly49A expression on T cells alters T cell selection

Ly49 receptors are inhibitory receptors expressed on subsets of both NK cells and NK1.1(+) T cells. The function of these receptors on NK cells is believed to be important in maintaining self-tolerance, yet their role on T cells is unclear. In this report we investigated how an Ly49A transgene alters T and NK cell development in an in vivo environment, where a ligand for Ly49A is expressed. Ly49A transgenic mice that co-expressed an MHC ligand for Ly49A, H-2D(d), developed a severe inflammatory disorder that resulted in death within the first weeks of age. T cells expressing forbidden TCR V(beta) chains were found both in the thymus and periphery of transgenic mice, while non-transgenic littermates had successfully deleted these T cell subsets. These data indicate that the expression of Ly49A on T cells could alter T cell selection and allow survival of potentially self-reactive T cells.     

Hyaline glomerulopathy in B6C3F1 mice.

Hyaline glomerulopathy is a spontaneous disease of undetermined etiology that occurs sporadically in various strains of aging mice. In our laboratory, this disease was observed with unusual ultrastructural features as an incidental finding in 2 female B6C3F1 mice from 2 carcinogenicity bioassays. Microscopically, renal lesions were characterized by marked diffuse enlargement and prominent hyalinization of the glomeruli, equally affecting both kidneys. Affected glomeruli were PAS positive, but were negative for amyloid by the Congo red method. Immunocytochemical staining revealed weakly positive glomerular deposits with polyclonal anti-mouse IgG-IgM-IgA cocktail. Ultrastructurally, there were characteristic subendothelial osmiophilic deposits composed of loosely-packed linear structures in the glomeruli. Lamellae, which appeared as fibrils in perpendicular sections, were relatively uniform, measured 6.1-17.01 nm in diameter, and formed single or double-layered structures. The ultrastructural and immunocytochemical characteristics are suggestive of a spontaneous immune-mediated mechanism in a strain of mouse commonly used in toxicology studies.     

CD4+ lymphocytes are extracted from blood by peripheral lymph nodes at different rates from other T cell subsets and B cells

The circulation of lymphocyte subsets through prescapular lymph nodes in sheep has been quantified using a panel of monoclonal antibodies against sheep lymphocyte surface antigens. Differences in the extraction of lymphocyte subsets from blood by the lymph node were found with CD4+ lymphocytes being extracted at a faster rate (1/2) than CD8+, SBU-T19+, major histocompatibility complex class II+ and B cells (1/4 to 1/5). In order to accommodate existing data on organ-specific adhesion molecules, one subset specific and one tissue specific, expressed on vascular endothelium could act jointly to regulate the migration of recirculating lymphocytes.     

Isolation of plasma membranes from rat C6 glioma cells cultivated on microcarriers.

We have studied the feasibility of rat C6 glioma cell cultivation on microcarrier beads and the isolation of their plasma membranes from the beads. Cells were cultivated on Cytodex-1 microcarrier beads and the plasma membranes were subsequently isolated from confluent cell monolayers on the beads. This approach yielded approximately 4 x 10(6) cells/ml in a 1 L spinner vessel. Enzymatic assays indicated an 18-fold enrichment of plasma membranes isolated from the beads with minor contamination by other cell organelles. Assay for IGF-I receptor binding capacity revealed that 70% of the total receptor binding capacity could be recovered in the plasma membrane fraction isolated from the beads as compared with the receptor binding capacity of intact cells, demonstrating the functional integrity of the isolated membranes. Electron microscopy and immunofluorescence analysis indicated that the isolated plasma membranes were highly homogeneous with the majority exposing the cytoplasmic surface. Our procedure of C6 glioma cell cultivation on microcarriers and subsequent plasma membrane isolation, provides large quantities of homogeneous and metabolically active membranes which can be used to study receptor-mediated effects on cell proliferation and differentiation.     

The expression of B cell surface receptors. III. The murine low-affinity IgE Fc receptor is not expressed on Ly 1 or 'Ly 1-like' B cells.

The distribution of IgE FcR (Fc epsilon R)-positive and -negative B cells was examined in normal adult mice. Using three-color flow cytometry, the expression of the Fc epsilon R was analyzed on various B-cell subsets present in the peritoneum and spleen. The results demonstrate that in the peritoneal cavity, the Fc epsilon R is not expressed on the large majority of Ly 1+ B cells and Ly 1-, Mac 1+ sister B cells. The receptor is present, however, on the small number of conventional B cells residing in the peritoneum. Although interleukin 4 (IL-4) can increase the levels of the Fc epsilon R on conventional B cells, incubation of Ly 1 and sister B cells with IL-4 did not result in the expression of the Fc epsilon R. When examining B cells present in the spleen, a small subset of B cells was consistently found to be Fc epsilon R-. These Fc epsilon R- cells were IgM-bright, IgD-dull and largely Ly 1- and Mac 1-negative. Staining of splenic tissue sections revealed that the Fc epsilon R- B cells were primarily localized to the marginal zones, whereas the Fc epsilon R+ B cells were found in the follicles. Taken together, the results indicate that the Fc epsilon R may be a useful marker in delineating the various B-cell subsets. In the peritoneum, the Fc epsilon R appears to discriminate conventional B cells from those of the Ly 1/sister lineage, and in the spleen it is likely to distinguish resting follicular B cells from Ly 1/sister and marginal zone B cells.     

Ly108 expression distinguishes subsets of invariant NKT cells that help autoantibody production and secrete IL-21 from those that secrete IL-17 in lupus prone NZB/W mice

Lupus is a systemic autoimmune disease characterized by anti-nuclear antibodies in humans and genetically susceptible NZB/W mice that can cause immune complex glomerulonephritis. T cells contribute to lupus pathogenesis by secreting pro-inflammatory cytokines such as IL-17, and by interacting with B cells and secreting helper factors such as IL-21 that promote production of IgG autoantibodies. In the current study, we determined whether purified NKT cells or far more numerous conventional non-NKT cells in the spleen of NZB/W female mice secrete IL-17 and/or IL-21 after TCR activation in vitro, and provide help for spontaneous IgG autoantibody production by purified splenic CD19⁺ B cells. Whereas invariant NKT cells secreted large amounts of IL-17 and IL-21, and helped B cells, non-NKT cells did not. The subset of IL-17 secreting NZB/W NKT cells expressed the Ly108^loCD4⁻NK1.1⁻ phenotype, whereas the IL-21 secreting subset expressed the Ly108^hiCD4⁺NK1.1⁻ phenotype and helped B cells secrete a variety of IgG anti-nuclear antibodies. α-galactocylceramide enhanced the helper activity of NZB/W and B6.Sle1b NKT cells for IgG autoantibody secretion by syngeneic B cells. In conclusion, different subsets of iNKT cells from mice with genetic susceptibility to lupus can contribute to pathogenesis by secreting pro-inflammatory cytokines and helping autoantibody production.     

Hyper-Ia antigen expression on B cells from B6-lpr/lpr mice correlates with manifestations of the autoimmune state

To investigate the state of activation of B cells from mice with the lpr gene defect, membrane Ia antigen (mIa) expression was analyzed on B cells from B6-lpr/lpr (lpr) and control B6- +/-/+/- mice. B cells from lpr mice exhibited marked increases in levels of mIa as determined by flow cytometry using a monoclonal anti-I-Ab,d reagent. This increase, which was progressive with age, suggests that phenotypic alteration of B-cell mIa expression is a consequence of lpr gene action. Since B-cell activation manifest by elevated mIa expression may promote productive interactions with helper T cells, these observations suggest an important role for B-cell abnormalities in the etiology of lpr-induced autoimmune disease.     

Abrogation of tumor induced Ly49 expression on mouse spleen cells by mitomycin C

Mouse spleen cells were activated with IL2 for 4 days in the presence or absence of paraformaldehyde fixed YAC (PFY) tumor cells (spleen cell: PFY ratio 100:1). Fresh spleen cells had poor expression of Ly49A but the expression was significantly upregulated by IL2 activation. Addition of PFYs resulted in a further boosting of Ly49A expression. Besides Ly49A, the Ly49C receptors were also upregulated by PFYs. Upregulated Ly49 expression was not restricted to NK cells (NK1.1 positive cells) but was also seen on T cells (TCRbeta positive cells). Time kinetics studies indicated that maximum upregulation of Ly49 in response to PFY cells occurred on day 3 and 4 and the expression declined thereafter. Ly49 expression in response to PFYs was completely blocked if spleen cells were pre-treated with Mitomycin C, an inhibitor of DNA synthesis. These results show that the addition of a small number of paraformaldehyde fixed YAC cells to spleen cell cultures undergoing IL2 activation, resulted in a significant upregulation of Ly49 receptors and this process was dependent upon cell proliferative activity.     

Modulation of B and T cell subsets in mice treated with fractionated total lymphoid irradiation. I. Blockage of differentiating B cell pathways

The effect of fractionated total lymphoid irradiation (TLI) on the maturation of B cells was examined in mice. At various times after irradiation of the mice, their spleen cells were tested for the mitogenic responses to dextran sulfate and lipopolysaccharide. In parallel the cells were stained with fluorescent anti-Thy-1 and various anti-Ig antisera, and were analyzed on a fluorescence-activated cell sorter (FACS). By comparison with normal spleen cells the cells of TLI-treated mice gave a high response to dextran sulfate and a low response to lipopolysaccharide. FACS analysis revealed that TLI-treated spleen contains elevated numbers of B cells bearing high IgM and low IgD on the surface. The B cells of TLI-treated mice retained these immature characteristics even two to four months after the last irradiation. These findings indicate that TLI causes a longlasting blockage in B cell maturation processes.     

T-bet-expressing B cells contribute to the autoreactive plasma cell pool in Lyn-/- mice

Systemic Lupus Erythematosus (SLE) is characterized by pathogenic autoantibodies against nucleic acid-containing antigens. Understanding which B-cell subsets give rise to these autoantibodies may reveal therapeutic approaches for SLE that spare protective responses. Mice lacking the tyrosine kinase Lyn, which limits B and myeloid cell activation, develop lupus-like autoimmune diseases characterized by increased autoreactive plasma cells (PCs). We used a fate-mapping strategy to determine the contribution of T-bet⁺ B cells, a subset thought to be pathogenic in lupus, to the accumulation of PCs and autoantibodies in Lyn^-/- mice. Approximately, 50% of splenic PCs in Lyn^-/- mice originated from T-bet⁺ cells, a significant increase compared to WT mice. In vitro, splenic PCs derived from T-bet⁺ B cells secreted both IgM and IgG anti-dsDNA antibodies. To determine the role of these cells in autoantibody production in vivo, we prevented T-bet⁺ B cells from differentiating into PCs or class switching in Lyn^-/- mice. This resulted in a partial reduction in splenic PCs and anti-dsDNA IgM and complete abrogation of anti-dsDNA IgG. Thus, T-bet⁺ B cells make an important contribution to the autoreactive PC pool in Lyn^-/- mice.