ZAP-70 expression is low in normal precursor B cells or hematogones.
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BACKGROUND: Zeta-associated protein (ZAP-70) expression has been associated with a less favorable prognosis in chronic lymphocytic leukemia (CLL). The role of ZAP-70 in immature B-cells is not well understood. Immature or precursor B lymphocytes (hematogones) often occur in clinical samples and coexpress CD10, CD38, and CD19 lacking surface immunoglobulin expression. The present study was carried out in an attempt to further elucidate the role of ZAP-70 in the spectrum of B-cells seen in clinical specimens. METHODS: ZAP-70 expression was evaluated on 25 patient samples that expressed greater than 2% of CD10(+)/CD38(bright)/CD19(+) coexpression during routine evaluation. In our sample set, the hematogone expression ranged from 2 to 18% of total leukocytes and occurred in a variety of conditions, including CLL, NHL, AML, MDS, Hodgkins Disease, and Multiple Myeloma. The method of ZAP-70 detection was that routinely applied to our clinical testing of CLL samples supplemented with the determination of ZAP-70 levels in the CD38(bright)/CD19(+) coexpressing cells. RESULTS: In all cases ZAP-70 was not expressed on the CD5 negative, CD38/CD19 coexpressing precusor B cells. In all cases hematogones were found to not express significant levels of ZAP-70. CONCLUSION: The presence of hematogones in clinical samples should be recognized so as to not adversely influence prognostic studies of ZAP-70 or CD38 in CLL. (+info)
B cells and aging: balancing the homeostatic equation.
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The interplay of selective and homeostatic processes dominates the behavior of B lineage subsets following B cell antigen receptor (BCR) expression, and extends to determinants of immune response quality and the persistence of immunologic memory. A key concept emerging from these considerations is that primary events acting upstream of mature B lymphocyte pools can profoundly impact downstream populations as the system attempts homeostatic adjustments. Since, advancing age is accompanied by profound changes in B cell generation and homeostasis, establishing the relative contributions of primary lesions versus compensatory homeostatic processes is critical to understanding these perturbations. Exploration of this problem requires an understanding of: (1) the identity, dynamics, and progenitor/successor relationships of marrow and peripheral B cell subsets; (2) the nature and interactions of selective and homeostatic processes acting in these subsets; (3) how these change with age. Our data show that BLyS and its receptors mediate peripheral B cell homeostasis, and that the size, dynamics and behavior of all B cell subsets influenced by B Lymphocyte Stimulator change with age. These findings suggest that homeostatic processes mediated through B Lymphocyte Stimulator are altered with age, and that these perturbations may primarily reflect compensatory homeostatic adjustments to upstream reductions in B cell generation. (+info)
Allele-dependent variation in the relative cellular potency of distinct EGFR inhibitors.
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Targeted cancer therapies impede cancer cell growth by inhibiting the function of activated oncogene products. Patients with non-small cell lung cancer and somatic mutations of EGFR can have a dramatic response to treatment with erlotinib and gefitinib; different somatic mutations are associated with different times to progression and survival. In this study, the relative and absolute potencies of two distinct EGFR tyrosine kinase inhibitors, erlotinib and an investigational irreversible inhibitor, HKI-272, were found to vary significantly in a panel of Ba/F3 cells transformed by representative EGFR somatic mutations. HKI-272 more potently inhibited the primary exon 20 insertion mutants, the secondary erlotinib-resistance mutants including T790M and many erlotinib-sensitive mutants including L858R. In contrast, erlotinib is a more potent inhibitor of the major exon 19 deletion mutants than is HKI-272. Analyses of EGFR autophosphorylation patterns confirmed the mutation-specific variation in relative potency of these tyrosine kinase inhibitors. Our finding that distinct EGFR inhibitors are more effective in vitro for different mutant forms of the protein suggests that tyrosine kinase inhibitor treatment could be tailored to specific EGFR mutations. More broadly, these results imply that the development and deployment of targeted therapies should focus on inhibition of specific cancer-causing mutations, not only on the mutated target. (+info)
The novel adaptor protein Swiprosin-1 enhances BCR signals and contributes to BCR-induced apoptosis.
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B-cell receptor (BCR) signals are essential for B-cell differentiation, homeostasis and negative selection, which are regulated by the strength and quality of BCR signals. Recently, we identified a new adaptor protein, Swiprosin-1, in lipid rafts of B-cell lines that undergo apoptosis after BCR stimulation. During murine B-cell development, Swiprosin-1 exhibited highest expression in immature B cells of the bone marrow, but was also expressed in resting and activated splenic B cells and in non-lymphoid tissue, especially in the brain. Ectopic expression of Swiprosin-1 in the immature murine B-cell line WEHI231 enhanced spontaneous and BCR-induced apoptosis. In contrast, short hairpin RNA (shRNA)-mediated downregulation of Swiprosin-1 impaired specifically spontaneous and BCR-elicited apoptosis, but not BCR-induced G1 cell cycle arrest and upregulation of the cell cycle inhibitor p27(Kip1). In accordance, Swiprosin-1 abundance regulated net cell growth of WEHI231 cell populations through reciprocal regulation of Bcl-xL, but not Bim, thereby controlling spontaneous apoptosis. Swiprosin-1-enhanced apoptosis was blocked through nuclear factor kappaB-activating stimuli, namely B-cell-activating factor of the TNF family, anti-CD40 and lipopolysaccharide (LPS). This correlated with enhanced BCR-induced IkappaB-alpha phosphorylation and degradation in cells expressing a Swiprosin-1-specific shRNA. Finally, ectopic Swiprosin-1 expression enhanced BCR-induced cell death in primary, LPS-stimulated splenic B cells. Hence, Swiprosin-1 may regulate lifespan and BCR signaling thresholds in immature B cells. (+info)
Mycoplasma contaminants present in exosome preparations induce polyclonal B cell responses.
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Exosome fractions of dendritic cells (DC) produced in long-term cultures (LTC) were found to contain Mycoplasma contaminants. In this study, Mycoplasma-infected, -uninfected, and -reinfected cultures of DC and control cell lines have been compared for their capacity to activate lymphocytes. Using differential centrifugation, size fractionation, and inhibition assays, it has been possible to map Mycoplasma to the exosome or vesicle fraction purified from culture supernatant (CSN). Mycoplasma fractions were shown to induce proliferation of B and not T cells. The B cell response was sensitive to mitomycin C and primaquine, both known antibiotics, but resistant to protease and DNase, suggesting a role for lipoproteins. Mycoplasma-contaminated exosome fractions of LTC-DC were potent mitogens for naive B cells and promoted Ig secretion. In contrast to the polyclonal B cell mitogen LPS, they were unable to promote Ig isotype switching. They induced polyclonal activation of all B cell subsets, including naive B cells, the T1 and T2 subsets of transitional B cells, marginal zone (MZ), and follicular (FO) B cells. The B cell proliferative response was not antigen-specific and occurred independently of T cell help. Implications for autoimmune sequelae associated with Mycoplasma infection are discussed along with the possibility that primaquine could be an effective treatment for Mycoplasma infection in humans. This study highlights the close association between exosomes and infectious agents like Mycoplasma and cautions about purification procedures for preparation of exosomes for studies on immunity. (+info)
Generation of functionally distinct B lymphocytes from common myeloid progenitors.
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Current models of adult haematopoiesis propose that haematopoietic stem cells (HSCs) differentiate into common lymphoid (CLP) and common myeloid (CMP) progenitors and establish an early separation between myeloid and lymphoid lineages. Nevertheless, the developmental potential of CMP-associated B cells suggests the existence of alternate pathways for B lymphopoesis. The aim of this study was to compare the developmental and functional properties of CMP- and CLP-derived B cells. While both populations matured through pro-B cell and transitional B cell intermediates in the bone marrow and spleen, respectively, following transfer into irradiated mice, mature CMP- and CLP-derived B cells exhibit distinct functional responses. Specifically, CMP-derived B cells did not respond to mitogenic stimulation to the same degree as their CLP-derived counterparts and secrete lower levels of IgM and the inflammatory cytokines such as interleukin (IL)-6 and IL-10. Together, these data suggest the existence of multiple pathways for generating functionally distinct B cells from bone marrow precursors. (+info)
Autoreactive B-1 B cells: constraints on natural autoantibody B cell antigen receptors.
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B-1 B-cells constitute a distinctive population of cells that are enriched for self-reactive B cell receptors (BCRs). These BCRs are encoded by a restricted set of heavy and light chains, including heavy chains that lack nontemplated nucleotide additions at the V-D and D-J joining regions. One prototype natural autoantibody produced by B-1 B cells binds to a cryptic determinant exposed on senescent red blood cells that includes the phosphatidylcholine (PtC) moiety. The V(H)11Vkappa9 BCR, which accounts for a large fraction of the anti-PtC specificity, is underrepresented in other B-cell populations, including newly formed B cells in bone marrow, and the transitional B cells, follicular B cells, and marginal zone B cells in spleen. Previous work has shown that V(H)11 heavy chains pair ineffectively with surrogate light chain (SLC) and so do not promote development in bone marrow, but instead allow fetal liver maturation because of a fetal preference for weaker pre-BCR signaling. Such inefficient SLC pairing constitutes one constraint on the maturation of B cells containing V(H)11 rearrangements that biases their generation to fetal development. Here, we examine another possible bottleneck to the B1 cell repertoire: light chain pairing with V(H)11 heavy chain, finding very significant preferences. (+info)
IL-15 regulates immature B-cell homing in an Ly49D-, IL-12 , and IL-18 dependent manner.
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To complete their maturation and participate in the humoral immune response, immature B cells that leave the bone marrow are targeted to specific areas in the spleen, where they differentiate into mature cells. Previously, we showed that immature B cells actively down-regulate their integrin-mediated migration to lymph nodes or to sites of inflammation, enabling their targeting to the spleen for final maturation. This inhibition is mediated by IFN-gamma, which is transcribed and secreted at low levels by these immature B cells; IFN-gamma expression is extinguished following B-cell maturation. Stimulation of the MHC class I receptor, Ly49D, triggers a signaling cascade that increases transcription of both IL-12 (p40) and IL-18; these, in turn, induce the secretion of IFN-gamma. In the present study, we demonstrate that Ly49D-dependent secretion of IL-12 and IL-18 induces IL-15 expression by immature B cells, and that these 3 factors together regulate IFN-gamma production that inhibits their ability to home to the lymph nodes or to sites of inflammation. Thus, IL-15 controls immature B-cell homing, resulting in shaping the B-cell repertoire to enable an efficient immune response. (+info)