Spontaneous Immunoglobulin-secreting Cells Research Articles

Close relationship between immunoglobulin secreting‐cells and Epstein–Barr virus reservoir in patients infected with HIV

Individuals infected with HIV have higher circulating Epstein-Barr virus (EBV) DNA load compared to healthy carriers. This study investigated whether level of spontaneous immunoglobulin secreting cells, one of the major hallmarks of HIV infection, is associated with an increase of EBV DNA load in PBMCs and the spontaneous EBV lytic cycle ex vivo in patients infected with HIV. Spontaneous virus production by cells infected with EBV and EBV DNA loads in PBMCs from which CD8(+) T-cells were removed were measured in 20 HIV-aviremic and 14 HIV-viremic patients. The number of circulating immunoglobulin-secreting cells (Ig-SCs) and CD8(+) T-lymphocyte activation were also investigated. Patients with detectable HIV RNA in plasma exhibited higher spontaneous ex vivo EBV secretion and higher levels of EBV DNA in PBMCs than their aviremic counterparts. In the two groups observed, a positive correlation was found between PBMCs EBV DNA viral load and Ig-SCs, CD38(bright) expression on CD8(+) T-cells and EBV DNA load in cell culture supernatants. These findings suggest that B-cell polyclonal activation and B-cell terminal differentiation into Ig-SCs may fuel EBV DNA reservoir and promote EBV production ex vivo in patients infected with HIV.

A fraction of murine placental extract enhances IgA production in cultured splenocytes

During the second half of murine pregnancy there is a characteristic increase in the number of spontaneous immunoglobulin-secreting cells in the maternal spleen (IgM, IgG and IgA), the uterus-draining lymph nodes (IgG) and Peyer's patches (IgA). There are indications that signals originating from the feto-placental unit are of importance for the generation of at least some of these changes. To evaluate further the role of placental factors as regulators of maternal Ig-secretion, placental extract (PE) was separated into eight fractions by gel-chromatography and each fraction was screened for its effect on splenic (1) background DNA-synthesis, (2) background Ig-secretion and (3) Ig-secretion in lipopolysaccharide (LPS) activated B-cells (anti-Thy1.2 + complement-treated splenocytes). Similarly prepared extracts of fetuses, maternal spleen and maternal liver served as controls. All tissue extracts and splenocytes were syngeneic to avoid reaction to allogeneic determinants. Placental extract fraction did not differ significantly from the other tissue extracts in affecting background lymphocyte activity. However, fraction number 3 from placental extract (PE3, corresponding to an approximate molecular weight of 18–70 kDa) was found strongly to increase the number of IgA-secreting cells ( P ⩽ 0.001) in LPS-activated B-cells. This effect was absent or much less pronounced in the corresponding controls. Blocking of PE3-activity by antibodies against rat prolactin indicated that the enhancing activity may be attributed to proteins of the prolactin/placental lactogen/growth hormone family. This assumption was further strengthened by experiments demonstrating that prolactin exerted preferential enhancement of IgA secretion when added to LPS-activated B-cell cultures in vitro. The possible role of placental prolactin-like molecules as regulators of maternal IgA secretion is discussed.

Spontaneous and cytokine‐inducible ‘natural’ immunoglobulin secreting cells in organized lymphoid tissues of mice

The number and frequency of spontaneous and cytokine-inducible 'natural' immunoglobulin-secreting cells (ISC) were determined in bone marrow (BM), spleen and Peyer's patch (PP), in vitro. Cells were cultured at limiting dilution in the presence or absence of exogenous recombinant cytokines and supernatants then assayed for total immunoglobulin (Ig) and Ig isotype using an ELISA. Most spontaneous ISC were found in the spleen and BM, with fewer in PP. The addition of recombinant interleukin 5 (rIL-5) promoted a marked increase in both the ISC frequency and the amount of Ig secreted/ISC whereas recombinant IL-6 (rIL-6) promoted only a marginal increase. Recombinant IL-4 (rIL-4) promoted a marginal increase in ISC frequency only. The isotype profile of ISC was in the order IgM greater than IgG2 greater than IgA greater than IgG3 greater than IgG1. The exposure of cells to 1200 rad of gamma-radiation resulted in decreased numbers of spontaneous ISC in all tissues, but the addition of rIL-5 or rIL-6 to the irradiated cells increased both the ISC frequency and Ig secreted. The Ig isotype profile was similar to that of non-irradiated ISC with a few minor exceptions. This large population of potential cytokine-inducible ISC could contribute to 'natural' Ig secretion in vivo.

Spontaneous immunoglobulin secreting cells in idiopathic usual interstitial pneumonia

Abundant evidence suggests that activation of B lymphocytes and formation of immune complexes play important roles in the pathogenesis of idiopathic pulmonary fibrosis. We investigated the level of B lymphocyte activation in patients with idiopathic UIP (n = 10, all were biopsy proven cases), for the purpose of evaluating its role. In order to determine the level of B lymphocyte activation, the number of spontaneous immunoglobulin secreting cells in blood was counted by using reverse hemolytic plaque assay. The number of IgA and IgG secreting cells significantly increased in patients with idiopathic UIP when compared with those in healthy controls (n = 8). The number of IgG secreting cells also significantly increased in patients with IP-CVD, but no significant increase in the number of Ig secreting cells could be detected in patients with pulmonary emphysema, diffuse panbronchiolitis or sarcoidosis. The study of cell surface markers revealed significant increase of CD21 (B2 and OKB7) positive cells in patients with idiopathic UIP. CD21 positive cells were immature B lymphocytes. These results suggest that increase of resting B lymphocytes and immunoglobulin secreting cells occurred and that the differentiation of B lymphocytes may be accelerated in idiopathic UIP. Furthermore, there is the possibility that soluble factors produced by T lymphocytes influenced the function of B lymphocytes in our experiment.

Abnormalities of B cell Activation and Immunoregulation in Patients with Chronic Inflammatory Liver Diseases

The immunoglobulin production capacities of peripheral blood lymphocytes obtained from patients with various chronic inflammatory liver diseases and from normal individuals were compared. Using a reverse hemolytic plaque assay, immunoglobulin-secreting cells (ISC) were counted immediately after isolation (immediate ISC) and again after 6-day, in vitro cultivation without stimulant (spontaneous ISC) or in the presence of pokeweed mitogen, PWM (PWM-induced ISC). An increased number of immediate ISC were observed in patients with chronic active hepatitis (CAH) of the autoimmune type (n = 7) or with CAH type B (n = 32), probably reflecting a defect of the in vivo suppressor cell system as previously demonstrated. In vitro preincubation of cells with 5 × 10 −8 M prednisolone reduced the increase in the number of immediate ISC in patients with CAH of the autoimmune type. On the other hand, lymphocytes obtained from patients with CAH-type NANB (n = 9) and with primary biliary cirrhosis (PBC) (n = 12) showed an impaired capacity to generate ISC upon stimulation with PWM. Spontaneous ISC from patients' lymphocytes were not significantly different from those of normal individuals. Using allogeneic co-cultures with lymphocytes from normal individuals and from patients with CAH NANB hepatitis or primary biliary cirrhosis, we observed no increase in suppressor cell activity. Therefore, the diminished responses to PWM are probably attributable to an alteration in the peripheral helper T-cell compartment.

Selective decrease in Leu8-negative T cell subpopulations following treatment with recombinant interferon-α2a (rIFN-α2a)

The effects of in vivo treatment with recombinant human IFN-α2a (rIFN-α2a) on the distribution of T cell subpopulations were examined in 21 patients with renal cell adenocarcinoma, using two-color flow cytometry with anti-Leu8 in combination with anti-Leu2 and anti-Leu3. Other parameters indicative of immune status, such as the number and percentage of total (CD3) T cells, in vitro proliferation to mitogen, and spontaneous immunoglobulin secretion, were also measured, prior to the initiation of treatment with rIFN-α2a, and during treatment. Total T cell number decreased after treatment with rIFN-α2a, to a low of 54% of mean pretreatment values after 4 weeks. The CD4 CD8 ratio did not change appreciably following treatment with rIFN-α2a. However, the number of Leu8-negative T cells, within both the CD4 and the CD8 T cell populations, decreased more than the number of Leu8-positive T cells. An increase in spontaneous immunoglobulin-secreting cells followed treatment with rIFN-α2a.

Effects of L-canavanine on immune function in normal and autoimmune mice: disordered B-cell function by a dietary amino acid in the immunoregulation of autoimmune disease.

This study reports the effects in vitro and in vivo of L-canavanine (LCN), an amino acid found in commonly consumed legumes, on immune function in normal and autoimmune mice. L-Canavanine in high doses effectively blocks all DNA synthesis in vitro within 24 h. At lower doses, LCN affects B-cell function of autoimmune New Zealand Black/New Zealand White (NZB/NZW)F1 mice, inhibiting [3H]thymidine incorporation in response to B-cell mitogens, and pokeweed-induced intracytoplasmic immunoglobulin synthesis. LCN stimulates intracytoplasmic immunoglobulin (IgG greater than IgM). T-cell functions such as lymphoproliferation in response to concanavalin A or phytohemagglutinin and T-cell cytotoxicity are not affected. Suppression of the lipopolysaccharide response by LCN is removed by the addition of fresh B cells. Addition of the amino acid to mouse diet resulted in a decrease in the life-span of the autoimmune NZB and (NZB X NZW)F1 mice and abolished the protective effect of male sex on their survival. The decrease in survival in LCN-treated autoimmune mice correlated with an increase in spontaneous immunoglobulin-secreting cells (IgG greater than IgM) and antinuclear and double-stranded DNA antibodies. The histopathological analyses revealed increased glomerular damage and immunoglobulin deposition in the kidneys of the LCN-treated autoimmune and normal (DBA/2) mice. Ten percent of normal mice developed high titers of autoantibodies after 24 weeks of the diet. These data suggest a dietary amino acid, L-canavanine, affects B-cell function resulting in autoimmune phenomena and providing a new animal model of autoimmunity, a diet-induced systemic lupus erythematosus.

Abnormalities of B-cell activation and immunoregulation in patients with Crohn's disease.

We have studied B-lymphocyte function in 39 patients with Crohn's disease and 35 normal individuals using a reverse haemolytic plaque assay as the effector system. Ten patients had active Crohn's disease, the others being in an inactive state of the disease. Compared with normal individuals, the Crohn's disease patients - especially those in the active state of the disease - had markedly raised numbers of spontaneous immunoglobulin secreting cells and severely decreased responses to the polyclonal activator pokeweed mitogen. The differences between the reactivity of patients with active disease and those with inactive disease were statistically significant. These findings indicate an in vivo polyclonal B-cell activation in Crohn's disease patients, possibly due to antigen(s) or infectious agent(s). In vitro experiments were performed with separated lymphocytes in order to characterise the mechanism responsible for the altered immune reactivity in Crohn's disease. These revealed an intrinsic B-cell defect as well as an impaired T-helper cell capacity in patients with Crohn's disease. Findings supporting the hypothesis of an increased suppressor activity in Crohn's disease patients could not be observed, and marker analyses revealed normal proportions with the exception of raised Leu 7 positive cells that mediate 'natural killer' and 'killer' cytolysis. We conclude that immune dysfunction in peripheral blood lymphocytes of Crohn's disease patients involves B-cells as well as T-helper cells.

Disorders of B cell activation in splenectomized patients

Using a reverse hemolytic plaque assay as the effector system, we studied in vitro B-lymphocyte function in 12 patients after posttraumatic splenectomy as well as in 25 normal individuals. The time interval between the splenectomy and the immunological studies varied between 2 days and 7 years. Compared to normal individuals, the splenectomized patients had markedly elevated numbers of spontaneous immunoglobulin-secreting cells and severely decreased responses to the polyclonal activator pokeweed mitogen, indicating an in vivo polyclonal B-cell activation. A tendency towards normalization of these abnormalities, especially the high spontaneous ISC levels, could be observed during the time interval extending up to 7 years after splenectomy. In order to characterize the mechanism responsible for the altered immune response in splenectomized patients, in vitro experiments were performed with separated lymphocytes. These revealed an impaired T-helper cell capacity as well as an intrinsic B-cell defect. Marker analysis with monoclonal antibodies revealed normal proportions with the exception of OKT 4 positive and B 1 positive cells that identify T-helper/inducer and peripheral B-cells, respectively. We conclude that immune dysfunction in peripheral blood lymphocytes of splenectomized patients involves mainly the OKT 4 positive cell population.

Spontaneous in vitro immunoglobulin secretion at the diagnosis of insulin-dependent diabetes.

Blood mononuclear cells obtained from 17 newly diagnosed insulin-dependent diabetic (IDDM) patients treated with insulin for 5-7 days were assessed for the number of spontaneous and pokeweed mitogen (PWM)-stimulated immunoglobulin-secreting cells in a reverse haemolytic plaque assay. The spontaneous in vitro immunoglobulin secretion was evanescent and decreased in individual patients within 1-4 months of insulin treatment. Compared to matched controls, 53% (9/17) of the IDDM patients had an elevated spontaneous secretion of immunoglobulin, 41% (7/17) for IgG, 35% (6/17) for IgM, and 35% (6/17) for IgA. The quantities of PWM-stimulated IgG, IgM, or IgA secreting cells in IDDM were comparable to the controls. The IDDM patients with spontaneous immunoglobulin secreting cells had higher fasting C-peptide levels compared to the patients with immunoglobulin-producing cells within the normal range (P less than 0.05). The average titre of islet cell cytoplasmic antibodies was 1:26 in (9 out of 9 were positive) patients with, compared to 1:1 in patients (4 out of 8 were positive) without spontaneous secretion (P = 0.025). These results suggest that the clinical onset of IDDM is associated with a polyclonal B lymphocyte activation and that higher levels of fasting C-peptide islet cell antibodies are associated with this immunoregulatory abnormality.