B-cells In Lymph Nodes Research Articles

Establishment of long-term CD154-dependent porcine B-cell cultures.

Cells of the B-cell lineage play an essential part in the immune response, not only as the producers of antigen-specific antibodies, but also as antigen-presenting cells. Unlike T cells, however, the establishment of long-term normal B-cell lines has proved to be exceedingly difficult. In this paper we demonstrate that cell membrane-expressed CD154 (CD40 ligand) is able to support the continual growth of porcine mesenteric lymph node B-cell cultures for more than 4 months without the addition of exogenous cytokines, such as interleukin-4 (IL-4). Addition of IL-4, but not interferon-gamma (IFN-gamma) or IL-13, to these cultures enhanced proliferation, as, to a lesser extent, did addition of IL-2. Interestingly, however, whilst IFN-gamma-supplemented cultures largely consisted of immunoglobulin M (IgM)-positive cells, cultures with IL-13 or IL-4 contained a significantly increased proportion of IgG-positive cells.

Lymphocyte subsets and expression of differentiation markers in blood and lymphoid organs of rhesus monkeys

Rhesus macaques are invaluable experimental animals in biomedical research. Using three color flow cytometry, we screened anti-human antibodies for crossreactivity with macaque cells in order to determine the distribution of functionally important lymphocyte subsets in blood, lymph nodes (LN), and spleen. NK-cells are almost completely absent in LN. The percentage of B-cells expressing CD80, CD86, and the level of expression of CD20 is higher in blood than in LN. In contrast, a higher proportion of B-cells in LN stains positive for CD21 and CD35. Whereas the number of CD29hi expressing T-cells is lower, CD69 is expressed on more T-cells in LN than in blood. About one-third of CD8+ T-cells in blood are CD28-, a subset with a unique pattern of antigen expression which cannot be found in LN. In contrast to humans, a relatively high proportion of T-cells in blood also express the co-stimulatory molecules CD80 and CD86. With increasing age, the proportion of B-cells in blood declines, whereas the percentage of T-cells rises. In addition, the proportion of CD29hi expressing T-cells increases among both the CD4+ and CD8+ subsets.

Kinetics of Somatic Mutation in Lymph Node Germinal Centres

Somatic mutation activity in immunoglobulin V kappa genes during the response to the hapten 2-phenyl-5-oxazolone was measured in lymph node B-cell populations at various timepoints after footpad immunization. When the V kappa Ox1 genes rearranged to the J kappa 5 segment were amplified from genomic DNA using the polymerase chain reaction and sequenced, somatic mutations could be detected as early as day 4 after immunization. Somatic mutations were also detected after sequencing RNA from oxazolone-specific hybridomas derived from lymph node cells at day 4 after immunization. These early mutations were found mostly in cells with a germinal centre phenotype. No indication of selection at the population level by apoptosis was detected until day 7 after immunization. These results suggest somatic mutations can be induced very early during the immune response in lymph node cells, prior to the peak of clonal expansion and selection with regard to antigen binding.

Immunohistological analysis of macrophages, B‐cells, and T‐cells in the mouse lung

Numerous studies have described the anatomy of the large lymphoid aggregates of bronchus-associated lymphoid tissue (BALT) in rabbits and rats. Less work has been performed on other immunocompetent areas of the respiratory tract, and available data again mostly describe rabbit or rat tissues. Little is known therefore of the microanatomy of the mouse lung immune system. We report a study, devised in order to establish the immunohistological characteristics of normal healthy mice lungs, performed on whole lungs from 22 mice of various strains and/or ages. Snap frozen tissues were serially sectioned and analysed using histochemistry and immunohistological techniques. Scattered macrophages, IgA plasma cells, B and T cells were enumerated in each sample. The largest population was that of macrophages. B-cells were numerous in all mice but 3 adults. T-cells were always present, L3T4+ often more numerous than Lyt2+ cells. Small lymphoid aggregates, composed of B or T cells (L3T4+ and Lyt2+) were seen in all mice, in the vicinity of a bronchiole and a vein. In 12/22 mice, a peculiarly elongated para-esophageal lymph node with large peripheral B-cell nodules and medullary T-cells was observed. In the five strains of mice studied, large variations were noted, affecting all the cell types studied, and related either to age or strain. Besides providing a qualitative description and quantitative analysis of immunocompetent cells, this work reports age and strain-related variations in these cells' distribution. These data could be relevant for studies involving the analysis of mice respiratory immune responses to environmental antigens.

Evidence for B-lymphocyte mitogen activity in Borrelia burgdorferi-infected mice.

Borrelia burgdorferi produces a mitogen for murine B lymphocytes which can be measured in vitro by polyclonal stimulation of proliferation and immunoglobulin production (R. Schoenfeld, B. Araneo, Y. Ma, L. Yang, and J. J. Weis, Infect. Immun. 60:455-464, 1992). Sonicated B. burgdorferi cells also stimulated IL-6 production by splenocyte cultures. We have used the murine model for Lyme disease described by Barthold et al. (S. W. Barthold, D. S. Beck, G. M. Hansen, G. A. Terwilliger, and K. D. Moody, J. Infect. Dis. 162:133-138, 1990) to determine whether the B. burgdorferi B-cell mitogen is expressed during active infection. To correlate arthritic changes with immune events, we have studied two strains of mice injected with B. burgdorferi; one of them, C3H/HeJ, developed severe disease, and the other, BALB/c, developed only mild disease. C3H/HeJ mice displayed a persistent 10-fold increase in circulating immunoglobulin G (IgG) levels, a 2-fold increase in IgM levels, and a 15-fold increase in peripheral lymph node B-cell numbers, providing evidence of mitogenic activity. Infected BALB/c mice also had evidence for mitogen activity, since the IgG level in serum increased three- to fourfold. The bulk of the increase in circulating IgG levels was not directed against B. burgdorferi antigens, supporting the occurrence of polyclonal B-cell activation. Analysis of IgG isotypes pointed out a contrast between C3H/HeJ and BALB/c mice in that levels of all isotypes were elevated somewhat in both strains of infected mice but IgG2a levels were much more dramatically increased in the C3H/HeJ mice (28-fold) than in the BALB/c mice (4-fold). In this study, interleukin-6 levels were found to be persistently elevated in the serum of infected C3H/HeJ mice. Interestingly, interleukin-6 levels in serum were much lower in the infected BALB/c mice. These findings indicate that the B. burgdorferi mitogen is active in infected animals and may contribute to the inflammatory and immune response to infection.

Effect of B lymphocytes from different organs on hematopoietic colony formation in the spleen by bone marrow cells

The influence of B-lymphocytes from various sources on splenic colony formation was studied in the syngeneic system. B-lymphocytes were obtained by panning with IgG-fraction of rabbit anti-mouse Ig, absorbed on Petri dishes. In addition, adherent cells, Thy-1+ and SC-1+ were eliminated from the fraction of Ig(+)-cells. SC-1- and SC-1+ fractions, containing, respectively, stem cells and T-lymphocyte precursors, were obtained by panning with IgG-fraction of rabbit anti-SC-1 serum. SC-1- cells transferred to irradiated syngeneic mice did not induce colony formation in the spleen. Introduction of SC-1- and SC-1+ cells induced formation of colonies. A similar helper effect occurred when SC-1(-)-cells were introduced with bone marrow or lymph node B-cells, but not with splenic B-cells. Splenic, but not bone marrow and lymph node B-cells inhibited colony formation by combination of SC-1- and SC-1+ cells. All effects of Ig+ cells were abolished by treatment of cells with rabbit anti-MBLA serum. Thus, B-cells of various origin can either enhance or inhibit colony formation. The enhancing of inhibitory effect after B (MBLA+)-cells elimination from suspension of bone marrow and lymph node (but not spleen) Ig(+)-cells resulted from the activity of B-contrasuppressors.

Fc epsilon receptors on rat B-lymphocytes: Specificity and binding kinetics

The binding kinetics of radiolabelled rat IgE to Fc epsilon receptors (R) of rat B-cells have already been studied in IgE-stimulated animals. The receptors expressed after Nippostrongylus brasiliensis infection or 2 injections of 5 mg IgE/100 g body wt were class-specific: IgE binding was not hindered by rat IgM, IgD, IgA, IgGI, IgG2a, IgG2b and IgG2c in immunocompetitive-binding assays. The rat B-cell Fc epsilon R were not species-specific, since mouse IgE competes with rat IgE for binding to these receptors. The apparent number of Fc epsilon R on rat mesenteric lymph node B-cells varied with temp and was 1.1–2.4 × 10 5 at 4°C and 5.9–7.7 × 10 5 at 37°C. The experimental K a was not influenced by temp and had an average value of 1.38 × 10 8 M −1. At 4°C the IgE binding to B-cell Fc epsilon R had an association rate of 4.9 × 10 3 M −1 sec −1 and a dissociation rate of 4.65 × 10 −5sec −1. After a very strong stimulation produced by injecting 5 mg IgE/100 g body wt every 24 hr for 5 days, the equilibrium binding curve became diphasic, indicating a probable heterogeneity of the B-cell Fc epsilon R.

Origin of permanently altered epithelial cells of the vagina in neonatally estrogen-treated mice.

In adult mice, neonatal injections of estrogen induce persistent, estrogen-independent cornification of the vaginal epithelium. Since the occurrence of nodules composed of affected epithelial cells (B cells) is a prodromic phenomenon of the irreversible vaginal alterations, the relationship between the occurrence of B-cell nodules and the time of DNA synthesis in the vaginal epithelial cells was examined by means of the techniques of serial mapping and autoradiography. B-cell nodules appeared in the epithelium of mullerian vagina 3-5 days after the initiation of neonatal estrogen treatment, without spatial continuity to cuboidal epithelial cells of the sinus vagina. The time of DNA synthesis was different in B cells from that in both columnar mullerian and cuboidal sinus cells, suggesting that the mother cells of B cells might be a population of still unidentified, undifferentiated cells.

Bronchogenic carcinoma: survival after surgical treatment according to stage, histologic type and immunomorphologic changes in regional lymph nodes.

The survival rates of 90 patients who underwent operation for bronchogenic carcinoma were analyzed statistically according to sex, age, tumor stage and histologic type, types of surgical procedures (lobectomy or pneumonectomy), and to immunomorphologic parameters of immunologic activity in peribronchial and hilar lymph nodes. The Stage I group survived significantly longer than did the Stage II and III groups, the lobectomized patients survived significantly longer than the pneumonectomized patients. The absence or presence of lymph node metastases was one of the major determinants of survival. The mean values (percent of total node cut surface) of lymph node sinus histiocytosis and of paracortical area could be correlated directly to survival in each of the histologic tumor-type groups, while the development of follicular cortex and germinal centers correlated inversely with survival. Increased survival might be associated with changes concomitant with immune reactivity in lymph node T-cell areas and with the sinus histiocytosis pattern, the latter representing probably a tumor--host reaction of the delayed hypersensitivity type. By contrast, increased activity of lymph node B-cell areas tended to correlate with poor prognosis of lung cancer patients. Moreover, patterns showing germinal center hyperplasia were statistically associated with lymph node neoplastic invasion, while the pattern with sinus histiocytosis and expanded paracortical areas was statistically associated with tumor-free lymph nodes.