Stimulation of B lymphocytes via CD72 (human Lyb-2).

A monoclonal antibody to CD72, the 45-kDa human homolog of murine Lyb-2, was found to augment selective activation pathways in tonsillar B lymphocytes. By itself, CD72 antibody provided a weak direct stimulus to resting B cells as assessed by [3H]dThd incorporation; this was modestly enhanced on the addition of interleukin 4 (IL4) or following ligation of surface CD40. CD72-delivered signals were more evident in co-stimulation assays with phorbol ester and with a synergistic combination of IL4 and CD40 antibody, but not with calcium ionophore or a CD23 antibody; rapidly cycling B cells were refractory to signaling via CD72 whether or not other co-stimuli were present. A unique feature of the CD72-delivered signal was its ability to enhance synergistically stimulations triggered with immobilized antibody to IgM, while failing to augment responses initiated by soluble anti-mu. On direct culture of resting B lymphocytes with CD72 antibody, an approximately twofold increase in the expression of major histocompatibility complex class II DQ antigen was observed, an augmentation similar to that achieved with IL 4; CD72 antibody also mimicked IL 4 in its ability to drive G0 cells into the early G1 phase of cell cycle. In contrast to IL 4-promoted stimulation, CD72 antibody failed to up-regulate the surface expression of either IgM or the CD23 antigen. CD72 expression itself was found to be weak on resting B lymphocytes and was modestly enhanced following culture with IL 4. The findings are discussed with reference to observations made on the triggering of murine B lymphocytes through Lyb-2 and within the context of previously defined human B cell activation pathways.     

Expression of the CD2 molecule on human B lymphoid progenitors.

CD2 expression on human B lymphoid progenitor cells was examined. By immunofluorescence analysis, a small fraction of bone marrow B cells was found to express CD2 on their surface. CD2 expression was not demonstrated on peripheral B cells. Epstein-Barr virus-transformed B cell lines derived from fetal liver at 8 weeks of gestation were analyzed to delineate the expression and function of CD2 at the early stage of human B cell development. Characterization of surface and genomic phenotypes of cell lines revealed that the established cell lines represent at least three different phenotypic characteristics of early B lineage cell: B progenitor, pre B, or early B cell. None of the 18 cell lines and 13 subclones with the phenotype of the early B lymphoid cells initially expressed CD2 antigen. However, CD2 expression was induced by the successive cultivation of some cloned B progenitor cell lines. In spite of the expression of CD2, these clones cell lines were unable to form rosettes with sheep red blood cells. By immunoprecipitation analysis, an identical 50 kDa protein was precipitated with anti-CD2 antibody from the lysates of the radioiodinated CD2+B progenitor cell line and peripheral blood T cells. Anti-CD2 antibody induced significant enhancement of proliferation of the CD2+B progenitor subline. These data indicate that human CD2 is expressed on a fraction of B lineage cells at a very early differentiation stage and may play a role in B lymphopoiesis.     

Expression and function of CD30 on T lymphocytes.

T cell receptor, accessory molecules, cytokines are important regulatory factors that determine the development and function of T lymphocytes. Among them are also molecules belonging to superfamily of tumor necrosis factor receptor (TNFR) which beside CD30 include CD27, CD40, TNFR-I and -II, Fas (CD95), OX40, 4-1BB (CDw137), nerve growth factor receptor, lymphotoxin-beta receptor, Apo3/DR3/Ws1-1/lymphocyte associated receptor of death, DR4, DR5/TNF-related apoptosis-inducing ligand, osteoprotegerin, and TNFR-related 2. CD30 recognized originally on Reed-Sternberg cells of Hodgkin's lymphoma became of interest in studies of Th1 and Th2 cell differentiation. This paper shows recent findings regarding CD30 expression and its pleiotropic role in T cell function. It provides information about controversial role of CD30 as Th2 cell differentiation marker and gives concise insight into the function of this receptor as a signal transducing molecule.     

Characterization of porcine CD5 and CD5+ B cells

Evidence strongly supports a role for the lymphocyte transmembrane glycoprotein CD5 in intracellular signalling events, whereby antigen-dependent growth and differentiation signals are augmented. Apart from its role in activation-related signalling, CD5 has been regarded as a possible B cell lineage marker differentiating subsets, CD5⁺ B cells (also termed B1 cells) and conventional B cells (or B2 cells). To extend these investigations to the study of pigs, porcine B cells were examined for evidence of CD5 expression. The influence of cellular activation on CD5 expression and CD5's role in signal transduction events and lymphocyte proliferation were examined. Using an anti-porcine CD5 MoAb (b53b7), porcine B cells were shown to be heterogeneous for CD5 expression. As in other species, B lymphocyte CD5 expression is low (dull), while IgM is high (bright). Ten to 30% of pig blood B lymphocytes are CD5⁺, with the highest frequency in neonates. Anti-CD5 antibody treatment was sufficient to induce rapid but transient calcium ion flux in porcine peripheral blood lymphocytes (PBL). CD5 expression increased on PBL following treatment with phorbol myristate acetate (PMA), lipopolysaccharide (LPS), or immobilized anti-IgM. LPS, PMA, and concanavalin A (Con A) but not anti-CD5, anti-IgM, or combinations of these antibodies induced lymphocyte ³H-thymidine uptake. CD5⁺ B cells are a common constituent of porcine circulating lymphocytes and resemble B1 cells of mice, man and other species in CD5 expression, frequency and lymphoid organ distribution. Porcine CD5, like CD5 in other species, mediates signal transduction, leading to changes in intracellular calcium concentration, but this signal alone is insufficient to promote cell division. A subset of porcine B cells up-regulates CD5 expression following phorbol ester activation.     

Cytolytic function for pseudorabies virus-stimulated porcine CD4+ CD8dull+ lymphocytes

We previously observed that pseudorabies (PRV) virus-specific killing in vitro was mediated by CD6+ CD8+ lymphocytes. Also a high percentage of CD4+ lymphocytes, among these CD6+ CD8+ lymphocytes, was observed. The purpose of this study was, therefore, to further characterize the killing ability of PRV-stimulated CD4+ CD8+ lymphocytes. Peripheral blood mononuclear cells (PBMC) were isolated from blood of PRV-immune pigs and were stimulated in vitro with PRV. After 6 days, the frequency of CD4+ CD8+ lymphocytes in peripheral blood was determined by flow cytometry analyses. Lymphocytes were separated using a magnet-activated cell sorter or a FACSVantage SE, and the cytolytic activity of the isolated populations was determined. Flow cytometry analyses demonstrated that PRV stimulation of immune PBMC resulted in the occurrence of 26% +/- 4% CD4+ CD8dull+ lymphocytes. We further demonstrated that killing by PRV-stimulated PBMC was mediated by CD4+ CD8dull+ T lymphocytes and CD4- CD8+ T lymphocytes (classic cytolytic T lymphocytes and natural killer cells). The CD4+ CD8dull+ T lymphocytes showed major histocompatibility complex (MHC) II-restricted PRV-specific killing. The CD4- CD8+ T lymphocytes showed both PRV-specific and natural killing. The CD4+ CD8dull+ lymphocytes, which are unique in the pig, seemed to have a more heterogeneous function than was earlier demonstrated. In conclusion, we demonstrated that PRV-specific CD4+ CD8dull+ lymphocytes are able to kill PRV-infected target cells in a MHC II-restricted manner.     

CD28在多发性硬化患者CD8+淋巴细胞的表达

目的探讨CD28在多发性硬化(MS)患者CD8+淋巴细胞的表达水平.方法流式细胞仪测定16例复发期MS患者和20例对照组外周血淋巴细胞CD28+、CD8+CD28-和CD8+CD28+的百分率.结果复发期MS患者淋巴细胞CD8+CD28-百分率低于对照组,CD28+和CD8+CD28+的百分率与对照组无明显差异;甲基强的松龙治疗对CD28+、CD8+CD28-和CD8+CD28+的百分率无影响.结论参与MS的发病的CD8细胞是CD8+CD28-细胞.     

Allergen-induced CD23 on CD4+ T lymphocytes and CD21 on B lymphocytes in Patients with allergic asthma: Evidence and regulation

Interaction of CD4⁺ T cells and B cells is necessary for IgE production. It has been recently demonstrated that cell surface antigen CD21 is a ligand for CD23 (Fc?RII) and that the pairing of these molecules may participate in the control of IgE production. In this study we investigated the effect of the Dermatophagoides pteronyssinus (Dpt) allergen and recombinant interleukin(rIL)-4 on the expression of CD21 and CD23 on T and B cells of asthmatic patients allergic to Dpt and of healthy controls. Peripheral blood mononuclear cells (PBMC) were incubated alone or with Dpt allergen (100 biological units/ml) and/or rIL-4 (100 U/ml) for up to 7 days. The flow-cytometric analysis of double-fluorescence staining revealed that Dpt allergen and/or rIL-4 induced CD23 on CD4⁺ T lymphocytes only in allergic patients. The allergen-induced CD23 on T cells is de novo synthesized antigen since no induction of CD23 on T cells was observed in cultures with 0.4 μg/ml actinomycin D. Moreover, 100 U/ml of interferon-γ inhibited the induction of CD23 on CD4⁺ T cells. T cells obtained from healthy donors did not express CD23 or CD21 antigen upon incubation with allergen and/or rIL-4. Although rIL-4 also induced CD23 in controls, the expression was only observed on CD20⁺ cells. The allergen alone induced a significant elevation of the mean fluorescence intensity of both CD21 and CD23 only in allergic in dividuals. When the cell proliferation was analyzed, a slightly increased stimulation index upon cultivation of PBMC was obtained from non-allergic donors as well, but less than in allergic patients. The co-expression of major histocompability complex class II molecules and CD23 on CD4⁺ T lymphocytes in allergic patients, as assessed by the three-color immunofluorescence analysis, indicates that these cells were activated. We conclude that CD4⁺ T lymphocytes possess a unique capability to express CD23 upon exposure to allergen. Moreover, the allergen-mediated induction of CD23 on T cells observed only in allergic patients may be the reason for the increase of IgE production. This would not occur in non-allergic individuals as there is no CD23 expression on T cells.     

Distribution of porcine CD4/CD8 double-positive T lymphocytes in mucosa-associated lymphoid tissues.

The present study describes the distribution of CD4/CD8 double-positive (DP) T cells in lymph nodes and mucosa-associated lymphoid tissues of pigs at 6-7 months of age. Proportions of lymphocytes isolated from peripheral, bronchial and mesenteric lymph nodes expressing CD4 and/or CD8 molecules were similar and ranged from 10-13% CD4/CD8 DP cells, 25-27% CD4 single-positive (SP) cells, and 27-32% CD8 SP cells. Mucosa-associated lymphoid tissues had significantly smaller proportions of CD4+ and/or CD8+ T cells than lymph nodes and CD4/CD8 DP cells accounted for a larger proportion of the total CD4+ lymphocytes than in lymph nodes. Compared to the lymph nodes, the predominant CD4+ and/or CD8+ T-cell subset in tonsils was the CD4/CD8 DP population (18%), because of both a higher proportion of these cells and a lower proportion of CD4 SP (12%) and CD8 SP (14%) lymphocyte subsets. Jejunal Peyer's patches were comprised of 12% CD4 SP, 28% CD8 SP and 10% CD4/CD8 DP lymphocytes. In contrast, the mid-section of the continuous Peyer's patch in the ileum contained 7% CD4 SP, 8% CD8 SP and 4% CD4/CD8 DP lymphocytes. The distribution of T lymphocytes in porcine mucosal lymphoid aggregates agrees with the reported correlation between high and low rates of lymphocyte entry into these tissues and the abundance or scarcity of T cells, respectively. Defining the role of CD4/CD8 DP lymphocytes and their unique distribution in mucosa-associated lymphoid tissues in the pig may reveal novel T-cell-mediated regulatory pathways.     

Expression of aberrant forms of CD22 on B lymphocytes in Cd22a lupus-prone mice affects ligand binding

CD22 functions primarily as a negative regulator of B-cell receptor signaling. The Cd22a allele has been proposed as a candidate allele for murine systemic lupus erythematosus. In this study, we explored the possible expression of aberrant forms of CD22, which differ in the N-terminal sequences constituting the ligand-binding site due to synthesis of abnormally processed Cd22 mRNA, in several Cd22a mouse strains, including C57BL/6 Cd22 congenic mice. The staining pattern of splenic B cells obtained with CY34 anti-CD22 mAb, which was expected to bind poorly to the aberrant CD22, was more heterogeneous in Cd22(a) mice than in Cd22b mice. Moreover, CD22 detected on B cells of Cd22a mice was expressed more weakly and as a smaller-sized protein, compared with Cd22b mice. Significantly, analysis with a synthetic CD22 ligand demonstrated that Cd22a mice carried a larger proportion of CD22 that was not bound by cis ligands on the B-cell surface than Cd22b mice. Finally, the study of C57BL/6 Cd22 congenic mice revealed that Cd22a B cells displayed a phenotype reminiscent of constitutively activated B cells (reduced surface IgM expression and augmented MHC class II expression), as reported for B cells expressing a mutant CD22 lacking the ligand-binding domain. Our demonstration that Cd22a B cells express aberrant forms of CD22, which can potentially deregulate B-cell signaling because of their decreased ligand-binding capacity, provides further support for Cd22a as a potential candidate allele for murine systemic lupus erythematosus.     

Identical forms of the CD2 antigen expressed by mouse T and B lymphocytes

A monoclonal antibody (12-15) reactive with the mouse CD2 was used to study the expression of the antigen in different lymphoid cell subsets. By two-color immunofluorescence using B or T cell-specific reagents and cell sorting in combination with biochemical analysis we provide evidence that the CD2 antigen is present on mouse B and T cells. The antigen is expressed by both subsets at similar density and appears to be biochemically indistinguishable.     

CD2-CD48 interaction prevents apoptosis in murine B lymphocytes by up-regulating bcl-2 expression

Antigen receptor engagement initiates clonal expansion and antibody secretion in B lymphocytes in response to foreign antigens. However, binding of self antigen to antigen receptors targets self-reactive B cell clones for elimination or inactivation. The antigen-triggered biochemical events and the eventual response of the cells are dependent on the simultaneous occupancy of co-stimulatory receptors. CD2 is an intercellular adhesion molecule implicated in cell activation and expressed in human T and natural killer cells as well as in mouse B lymphocytes. Mouse B cells specific for allogeneic major histocompatibility complex (MHC) class I initiate a suicide program that leads to DNA fragmentation and cell death when confronted with soluble MHC class I while undergoing clonal expansion when the antigen is present on mitomycin C-treated cells. Here we show that occupancy of CD2 in mouse B cells by the presence of either monoclonal antibody (mAb) specific for CD2, or soluble recombinant mouse CD48, its natural ligand in mouse, prevents the induction of apoptosis. Furthermore, the in vitro activation by mitomycin C-treated allogeneic cells, is abrogated in the presence of anti-CD48 mAb (OX78). These results indicate that a CD2-CD48 interaction is involved in the control of B cell activation.     

CD8 T cell priming by B lymphocytes is CD4 help dependent

While it is generally accepted that B lymphocytes can present antigen and activate CD4 T cells, priming of CD8 T cells by B lymphocytes remains controversial. Recently, we showed that mice injected with genetically programmed B lymphocytes generate antigen specific CD4 and CD8 T cell responses in vivo that could also be induced in mice lacking functional dendritic cells. To gain further insights into the requirements for T cell priming by antigen-presenting B lymphocytes, in vitro experiments were performed using ovalbumin (OVA) and OVA-specific TCR-transgenic CD4 and CD8 T cells. We found that while B lymphocytes can directly prime CD4 T cells, the activation of CD8 T cells requires T cell help. Transfer experiments show that help can either be contact dependent or be mediated by soluble factors in the supernatants of activated OVA-specific CD4 T cells. Furthermore, the effect of activated CD4 T cells can be replaced by soluble recombinant IL-4. Collectively, the data show the existence of different requirements for priming of CD4 and CD8 T cells and point to the previously unappreciated fact that the induction of CD8 T cell responses by B lymphocytes requires T cell help.     

Simultaneous expression of CD4 and CD8 antigens by a substantial proportion of resting porcine T lymphocytes

The existence of four subpopulations in resting porcine T lymphocytes is documented. In addition to the two known subpopulations which are typified by a mutually exclusive expression of either the CD8 or the CD4 differentiation antigen, CD4-CD8+ and CD4+CD8- T lymphocytes, respectively, two unusual subpopulations were prominent not only in peripheral blood, but also in lymphoid tissues: CD4-CD8- T lymphocytes expressing neither of these antigens and CD4+CD8+ T lymphocytes coexpressing both antigens. While CD4+CD8+ lymphoblasts have been found also in other species, resting T lymphocytes with that phenotype are without precedent among all species analyzed to date. This unique composition of the porcine T lymphocyte population has to be taken into consideration when the swine is used as a large animal model in experimental medicine.     

反义CD40RNA对EB病毒转化的B细胞CD40分子表达和增殖能力的影响

目的 :探讨瞬时表达的反义CD4 0RNA ,对EB病毒转化的健康人B细胞膜表面CD4 0分子表达和增殖能力的影响。方法 :应用T A克隆技术和亚克隆技术 ,构建人反义CD4 0RNA的真核表达载体pcDNA3/CD4 0 ,并以其转染本室建立的EB病毒转化的健康人B细胞。应用流式细胞仪 (FACS) ,检测B细胞膜上CD4 0分子表达的变化。应用MTT比色法检测反义CD4 0RNA对B细胞增殖能力的影响。结果 :与转染空载体pcDNA3组相比 ,转染pcDNA3/CD4 0细胞上CD4 0分子的表达降低 (P<0 .0 1) ,其增殖能力明显降低 (P <0 .0 1)。结论 :反义CD4 0RNA技术 ,可作为有效的免疫调控手段。CD4 0基因本身在细胞的生长代谢中也起着重要作用     

Expression of CD34 on human B cell precursors.

CD34 is a 110-kD glycoprotein previously shown by a variety of monoclonal antibodies (MoAbs) to be expressed selectively on immature hematopoietic cells. However, more detailed characterization of CD34+ cells has been hampered by lack of anti-CD34 MoAbs that can be labelled directly with fluorochromes to facilitate subpopulation analysis by multi-parameter flow cytometry. We have recently isolated a murine anti-CD34 MoAb, designated as 8G12, that can be directly labelled with fluorochromes such as FITC. In this study, we have exploited this property of 8G12 to compare the reactivity of 8G12 and My10 with normal and leukaemic human marrow cells and to characterize normal early human B cell precursors by two- and three-colour immunofluorescence analysis. Comparison of three-colour staining profiles of normal bone marrow cells incubated with both 8G12 and MY10, and either anti-CD10 or anti-CD19 MoAb revealed the reactivity patterns of 8G12 and MY10 to be indistinguishable. This conclusion was confirmed by a similar comparative analysis of 8G12 and MY10 staining of blood and bone marrow cells from 4 patients with B lineage acute lymphoblastic leukaemia (ALL). Of interest, both 8G12 and MY10 detected a CD34+CD10+CD19- population in normal adult bone marrow. To determine whether a CD34+CD10+CD19- precursor population previously reported by others to exist in fetal liver could also be identified, CD10+CD16- marrow cells were first isolated by FACS and the sorted cells then re-analysed for expression of CD19 and CD34. These studies showed that all of the sorted CD10+ cells that expressed CD34 appeared to coexpress CD19. No CD34+CD10+CD19- cells were detected (at a sensitivity of less than or equal to 0.1%). Further studies will be required to determine whether a very minor population of CD34+CD10+CD19- cells may still be generated in the normal development of B cells in adult human marrow.     

Characterization of a porcine CD1-specific mAb that distinguishes CD4/CD8 double-positive thymic from peripheral T lymphocytes

Porcine peripheral T-cells bear CD4 and CD8 cell surface antigens that distinguish helper from cytotoxic T-cells. In distinction from what has been found in other species, a large percentage of peripheral T-cells simultaneously express both CD4 and CD8. Monoclonal antibody 76-7-4 was found to stain all cortical and 7 +/- 3% of medullary thymocytes, Ia+ epidermal cells (i.e. Langerhans cells), no peripheral T-cells and 50% of peripheral B-cells. The antigen detected appears analogous to human CD1. All cortical thymocytes were also stained with CD4 and CD8 mAb. Since 76-7-4 did not stain peripheral T-cells, we conclude that CD4/CD8 dual-expressing peripheral T-cells are not simply immature thymic emigrants.     

Expression of functional interleukin 2 receptors on chronic lymphocytic leukaemia B lymphocytes is modulated by recombinant interleukin 2.

In 16 of 18 chronic lymphocytic leukaemia (CLL) patients examined, a significant proportion of B cells in the leukaemic clone bound monoclonal antibodies specific for the interleukin 2 (IL-2) receptor site (CD25). B lymphocytes from patients tested showed a direct response to recombinant interleukin (rIL-2) during culture in vitro as shown by: (a) a ligand-mediated upregulation in the level of IL-2 receptor (IL-2R) expression (12 of 12 patients), (b) an increase in cell size (eight of nine patients), (c) an increase in 3H-thymidine uptake (four of six patients). Taken together, this evidence suggests that the majority of leukaemic B cells from all the CLL patients examined expressed functional IL-2 receptors in vitro. Intriguingly, maximal receptor upregulation or increase in cell size was achieved at a lower concentration (50 u/ml) of rIL-2 than was required to achieve maximal 3H-thymidine incorporation.     

CD20: a regulator of cell-cycle progression of B lymphocytes

CD20 is a B-cell-specific cell-surface molecule with four membrane-spanning domains, as well as cytoplasmic N- and C-terminal domains. Here, Thomas Tedder and Pablo Engel discuss the suggestion that CD20 is a regulator of transmembrane Ca²⁺ onductance and plays an important functional role in B-cell activation, proliferation and differentiation.