Developmental relationships between B-1 and B-2 progenitors

Comment on: Barber CL, et al. Proc Natl Acad Sci USA 2011; In press.     

Developmental stress can uncouple relationships between physiology and behaviour

Phenotypic correlations (r_P) have frequently been observed between physiological and behavioural traits, and the nature of these associations has been shown to be modulated by a range of environmental stressors. Studies to date have examined the effects of acute stressors on physiology–behaviour interrelations, but the potential for permanent changes induced by exposure to stress during development remains unexplored. We exposed female zebra finches to dietary restriction during the nestling stage and tested how this affected r_P among a variety of physiological traits (haematocrit, stress-induced corticosterone level and basal metabolic rate (BMR)) and behavioural traits (activity and feeding rates in novel and familiar environments). Developmental stress completely uncoupled the relationship between activity in a novel environment and two physiological traits: haematocrit and BMR. This suggests that nutritionally based developmental stress has provoked changes in the energy budget that alleviate the trade-off between maintenance (BMR) and locomotor activities.     

The H2 haplotype regulates the distribution of B cells into B-1a,B-1b and B-2 subsets

The size of B-cell subsets appears to be under genetic control, but the mechanism of this regulation is unknown. By analyzing five congenic strains of mice that differ only in their H2 haplotype, we addressed the issue of whether the MHC genes are involved in the relative proportions of B-1a, B-1b and B-2 cells. Not only were there considerable differences in the percentages of B-1 in B cells between H2s mice which were the highest [78.5+/-0.8% in the peritoneal cavity (PerC), and 26.3+/-0.5% in the spleen] and H2d mice, which were the lowest (15.2+/-0.6% in the PerC, and 10.9+/-0.6% in the spleen), but the percentages of B-1a cells varied inversely to those of B-1. Crosses between H2s and H2d strains showed that the highest B-1 frequencies occurred in F2 progeny expressing the homozygous H2s (70.8+/-2.1% in the PerC, and 30.0+/-0.5 in the spleen), and the lowest in that expressing the homozygous H2d haplotype (8.9+/-0.6% in the PerC, and 8.6+/-0.4% in the spleen). A dose effect of H2 was established in heterozygous F1 and F2 mice. As mice aged, there was a reduction of B-1 cells in the PerC, at the expense of B-1b in the H2s, but not in the H2d mice. Hence, the H2 genes appear to participate in regulating the proportions of B-1a, B-1b and B-2 cells.     

Restricted IgA repertoire in both B-1 and B-2 cell-derived gut plasmablasts

Mucosal IgA is the most abundantly produced Ig upon colonization of the intestinal tract with commensal organisms in the majority of mammals. The repertoire of these IgA molecules is still largely unknown; a large amount of the mucosal IgA cannot be shown to react with the inducing microorganisms. Analysis of the repertoire of used H chain Ig (V(H)) genes by H-CDR3 spectrotyping, cloning, and sequencing of V(H) genes from murine intestinal IgA-producing plasma cells reveals a very restricted usage of V(H) genes and multiple clonally related sequences. The restricted usage of V(H) genes is a very consistent observation, and is observed for IgA plasma cells derived from B-1 or conventional B-2 cells from different mouse strains. Clonal patterns from all analyzed V(H) gene sequences show mainly independently acquired somatic mutations in contrast to the clonal evolution patterns often observed as a consequence of affinity maturation in germinal center reactions in peripheral lymphoid organs and Peyer's patches. Our data suggest a model of clonal expansion in which many mucosal IgA-producing B cells develop in the absence of affinity maturation. The affinity of most produced IgA might not be the most critical factor for its possible function to control the commensal organisms, but simply the abundance of large amounts of IgA that can bind with relatively unselected affinity to redundant epitopes on such organisms.     

Stromal cell-dependent growth of B-1 B cell progenitors in the absence of direct contact

This protocol describes a transwell culture system in which stromal cells support the growth and differentiation of B cell progenitors in the absence of direct contact. In this system, a confluent layer of S17 stromal cells pre-established in 0.4 microm transwells is placed over wells seeded with purified B cell progenitors. The stromal cell-derived factors and additional cytokines added to the culture medium support the differentiation of the progenitors in the lower chamber. B-1 B cell progenitors seeded in the presence of thymic stromal lymphopoietin undergo significant expansion and differentiation in this culture system. Since the expanded B-1 B lineage cells are not contaminated with stromal cells, no additional purification steps are required before subsequent phenotypic, functional or genetic analyses of these lymphoid cells are performed. Once the transwell cultures and B cell progenitors are available, cultures can be initiated in less than an hour. The overall procedure, however, takes approximately 10 h when the initiation of the S17 transwell cultures and the isolation of the B cell progenitors steps are included.     

Differentially regulated expression and function of CD22 in activated B-1 and B-2 lymphocytes

CD22 is a B cell-restricted transmembrane protein that apparently controls signal transduction thresholds initiated through the B cell Ag receptor (BCR) in response to Ag. However, it is still poorly understood how the expression of CD22 is regulated in B cells after their activation. Here we show that the expression levels of CD22 in conventional B-2 cells are markedly down-regulated after cross-linking of BCR with anti-IgM mAb but are up-regulated after stimulation with LPS, anti-CD40 mAb, or IL-4. In contrast, treatment with anti-IgM mAb barely modulated the expression levels of CD22 in CD5(+) B-1 cells, consistent with a weak Ca(2+) response in anti-IgM-treated CD5(+) B-1 cells. Moreover, in CD22-deficient mice, anti-IgM treatment did not trigger enhanced Ca(2+) influx in CD5(+) B-1 cells, unlike CD22-deficient splenic B-2 cells, suggesting a relatively limited role of CD22 in BCR signaling in B-1 cells. In contrast, CD22 levels were markedly down-regulated on wild-type B-1 cells in response to LPS or unmethylated CpG-containing oligodeoxynucleotides. These data indicate that the expression and function of CD22 are differentially regulated in B-1 and conventional B-2 cells, which are apparently implicated in innate and adaptive immunity, respectively.     

Developmental and immunological aspects of Drosophila-parasitoid relationships

The Drosophila-parasitic wasp (parasitoid) associations involve integrating adaptations of considerable complexity. This review focuses on some of the factors that influence these interactions including host immunity, nutrition and hormonal changes, and parasitoid virulence and mechanisms of immune suppression.     

Research resource: Comparative nuclear receptor atlas: basal and activated peritoneal B-1 and B-2 cells

Na?ve murine B cells are typically divided into three subsets based on functional and phenotypic characteristics: innate-like B-1 and marginal zone B cells vs. adaptive B-2 cells, also known as follicular or conventional B cells. B-1 cells, the innate-immune-like component of the B cell lineage are the primary source of natural antibodies and have been shown to modulate autoimmune diseases, human B-cell leukemias, and inflammatory disorders such as atherosclerosis. On the other hand, B-2 cells are the principal mediators of the adaptive humoral immune response and represent an important pharmacological target for various conditions including rheumatoid arthritis, lupus erythematosus, and lymphomas. Using the resources of the Nuclear Receptor Signaling Atlas program, we used quantitative real-time PCR to assess the complement of the 49 murine nuclear receptor superfamily expressed in quiescent and toll-like receptor (TLR)-stimulated peritoneal B-1 and B-2 cells. We report the expression of 24 nuclear receptors in basal B-1 cells and 25 nuclear receptors in basal B-2 cells, with, in some cases, dramatic changes in response to TLR 4 or TLR 2/1 stimulation. Comparative nuclear receptor profiling between B-1 and peritoneal B-2 cells reveals a highly concordant expression pattern, albeit at quantitatively dissimilar levels. We also found that splenic B cells express 23 nuclear receptors. This catalog of nuclear receptor expression in B-1 and B-2 cells provides data to be used to better understand the specific roles of nuclear receptors in B cell function, chronic inflammation, and autoimmune disease.     

B-1a, B-1b and B-2 B cells display unique VHDJH repertoires formed at different stages of ontogeny and under different selection pressures.

Analyses of VHDJH rearrangements isolated from murine peritoneal B-1a cells (CD5+, IgMhi, B220lo), peritoneal B-1b cells (CD5-, IgMhi, B220lo), and conventional splenic B cells provide evidence that a unique repertoire of VH regions is displayed by each of these B-cell subsets. The B-1a subset is characterized by a low N-region diversity, by a high frequency of sequence homologies in the VH-D and D-JH junctions, and by a limited exonuclease nibbling of the terminals of the joining gene segments. Through expansion in ageing mice, B-1a clones with these properties are favoured. B-1b cells are similar to conventional B-2 cells with respect to N-region diversity, but are unique in terms of D gene expression. Thus, while most murine pre-B and B cells preferentially use DSP and DFL gene segments in a given reading frame (RF1), B-1b cells frequently express D genes in another reading frame (RF2). Together, these findings provide structural evidence for a model where B-1a, B-1b and B-2 cells are produced by separate progenitors that are active at different stages of ontogeny.     

Developmental kinetics of hemopoietic progenitors in the avian embryo spleen

By means of plasma clot clonal cultures, the content of the avian spleen in granulomonocytic progenitors was studied during ontogeny. Serum-free media were used that were supplemented with growth activities produced either by embryonic fibroblasts or adult spleen cells. These two conditioned media not only permitted the growth of M-CFC, G-CFC, and GM-CFC but also F-CFU (fibroblast colony-forming units) from quail or chick embryonic spleen cells. The presence of spleen cell-conditioned medium promoted the development of large colonies of immature granulocytes. In the chick the first hemopoietic progenitors appeared at E9 and their number displayed two peaks, one at E15 and a smaller one at E18. In the quail the first progenitors were detected as early as E7 and their number peaked at E10. In this species, hemopoietic progenitors disappeared definitively before hatching while in the chick some were still present at P3. The progenitor content of the chick embryo spleen was compared to that of the bone marrow. This content remained stable during all of embryonic life, while the bone marrow exhibited a very different profile, where a sharp peak at E16 was followed by an acute decline and a stabilization at a rather low level. The particular profile in the spleen speaks in favor of a special role of this organ in the development of the hemopoietic system.     

Glial progenitors in the CNS and possible lineage relationships among them

Glial cells are derived from stem cells that mature through specific stages of development to generate fully differentiated astrocytes and oligodendrocytes. Several types of intermediate precursors have been described and in some cases lineage relationships identified although this remains a subject of controversy. We review recent findings and discuss some possibilities. Motoneuron-oligodendrocyte precursors (MNOPs), white matter progenitor cells (WMPCs), polydendrocytes, glial restricted precursors (GRPs), astrocyte precursor cells (APCs), and oligodendroblasts are likely all derived from earlier appearing stem cells but segregate at different stages in development. Some of these precursors persist in the adult, and it is these glial progenitors rather than stem cells that respond after injury and participate in the repair process. Although which specific glial progenitor responds remains unclear, the availability of new markers will likely resolve this issue. We believe that the development of consensus sets of markers and an improvement in our ability to define stages of glial maturation will lead to a clearer appreciation of the importance of glia in the etiopathology of disease.     

A comparative NMR and molecular dynamics study of the conformations of bradykinin B1 and B2, B2, and B1-specific receptor antagonists B-9430, B-9436, and B-9858

Extensive proton magnetic resonance experiments were carried out on three bradykinin peptide antagonists B-9430, B-9436, and B-9858 in aqueous solutions as well as in sodium dodecylsulphate micelles (B-9430 and B-9436) and CD₃OH/H₂O (60%/40%) mixtures for B-9858. All three peptides showed no observable secondary structure in aqueous solution. However, in their respective structure-inducing solvents, B-9430 (B₁ and B₂ receptor antagonist) and B-9436 (a B₂ receptor antagonist) exhibit a type II β-turn involving residues 2–5, and B-9430 also exhibits a type II′ β-turn involving residues 6–9 (in sodium dodecylsulfate micellar solutions), whereas B-9858, a B₁-specific receptor antagonist, exhibits only a type II β-turn involving residues 2–5 (in CD₃OH/H₂O solutions). Simulated annealing calculations on B-9858 confirm the experimental conclusions based on the nmr data. In addition, simulated annealing of the (2S, 3aS, 7aS)-octahydroindole-2-carboxylic acid (Oic residue), which is present in two of the three decapeptides studied, show that the one-chair conformation of the six-membered ring predominates, in agreement with the experimental data. The activities of these peptides are compared with their secondary structures and the specific receptor activity appears to depend on the presence of specific amino acid residues, such as N-(2-indanyl)glycine (Nig) and D[α-(2-indanyl)glycine] (D-Igl) as well as on elements of secondary structure. © 1997 John Wiley & Sons, Inc. Biopoly 42: 521–535, 1997     

Rapid,economical qualitative method for separation of aflatoxins B-1, B-2 & G-1, G-2 by dry column chromatography

A good correlation of four components of aflatoxins was accomplished by using the dry column chromatography method. The decolorization process of interfering substances, by 0.01 N KOH and defatting the extract with petroleum ether yields a clean residue for DCC separation. It is clear that the dry column chromatography is a very simple and time-saving procedure for separation of aflatoxins. DCC columns are more economical than precoated 'thick layer' preparative plates and, in DCC, no large developing tanks need to be used. Hazards associated with the use of large volumes of flammable solvents are greatly reduced.