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Clinical and Experimental Immunology Apr 1981Using a culture system that includes a B-enriched fraction of peripheral blood lymphocytes (PBL) mixed with non-proliferating "filler" cells (mitomycin-treated PBL or T...
Using a culture system that includes a B-enriched fraction of peripheral blood lymphocytes (PBL) mixed with non-proliferating "filler" cells (mitomycin-treated PBL or T cells), the kinetics of human B cell proliferation and differentiation into cells containing cytoplasmic immunoglobulin (cIg) have studied. Following stimulation with phytohaemagglutinin (PHA) or pokeweed mitogen (PWM), proliferative responses of enriched B cells were comparable to those for unseparated PBL, with a peak at 3 to 5 days and a steep decline thereafter; cIg-positive cells increased with both mitogens from day 3 until 7. Little proliferation or differentiation occurred in B cell cultures lacking T "filler" cells. The results indicate that human B cells can respond polyclonally to PHA in vitro, as well as to PWM, if non-proliferating T cell "help" is present. Further, these proliferative responses, but not subsequent differentiation, were inhibited by PHA-stimulated blasts that were treated with mitomycin and added on day 3 of culture.
Topics: Adult; B-Lymphocytes; Cell Differentiation; Cell Division; Cells, Cultured; Humans; Kinetics; Lymphocyte Activation; Phytohemagglutinins; Pokeweed Mitogens; Rosette Formation; T-Lymphocytes; Time Factors
PubMed: 6973432
DOI: No ID Found -
Immunology Oct 1996It has been reported that interleukin-6 (IL-6) is expressed in cells of acute inflammatory granulomas experimentally induced in mice by eggs of Schistosoma mansoni....
It has been reported that interleukin-6 (IL-6) is expressed in cells of acute inflammatory granulomas experimentally induced in mice by eggs of Schistosoma mansoni. Moreover, in vitro IL-6 was shown to enhance the cytotoxic activity of human platelets against larvae of S. mansoni. To elucidate further a proposed biological significance of this cytokine during the course of schistosomiasis, we studied the kinetics of IL-6 production and concomitantly performed a histopathological analysis of the livers in BALB/c mice subcutaneously infected with S. mansoni cercariae. Over a period of 24 weeks postinfection (p.i.) we monitored serum IL-6 levels, IL-6 production in vitro by pokeweed mitogen (PWM)-stimulated spleen cells as well as IL-6 mRNA expression in livers, spleens and kidneys. We found significantly elevated IL-6 levels in PWM-stimulated spleen cell-conditioned media (SCM) at weeks 6 to 20 p.i., peaking at week 10 p.i. In contrast, serum IL-6 concentrations started to rise not before week 8 but remained significantly elevated above normal control values until week 24 p.i. The time pattern of enhanced IL-6 mRNA expression detected in spleens and livers, but not in kidneys, as well as the rises of IL-6 in SCM and with a delay of 2 weeks in serum samples correlated with the onset of the egg-induced inflammatory reactions as well as the incidence and the number of the granulomas observed histopathologically in the livers of infected mice. Our data emphasize both a local and a systemic role of IL-6 in the host immune response following infection of mice with S. mansoni.
Topics: Animals; Blotting, Northern; Cells, Cultured; Female; Granuloma; Interleukin-6; Kidney; Liver; Mice; Mice, Inbred BALB C; Pokeweed Mitogens; RNA, Messenger; Schistosomiasis mansoni; Spleen; Stimulation, Chemical
PubMed: 8943723
DOI: 10.1046/j.1365-2567.1996.d01-737.x -
Blood Jul 2000Myeloma tumor cells, both freshly excised and cultured, are extremely resistant to cell-mediated cytolysis. As evidence suggests that B-cell susceptibility to lysis is...
Myeloma tumor cells, both freshly excised and cultured, are extremely resistant to cell-mediated cytolysis. As evidence suggests that B-cell susceptibility to lysis is dependent upon its state of differentiation and activation, we tested the ability of a variety of B-cell proliferation and differentiation agents, including pokeweed mitogen (PWM), to enhance the sensitivity of myeloma cells to cell-mediated lysis. PWM was found to significantly enhance the susceptibility of myeloma cell lines and freshly isolated myeloma cells to interleukin-2 (IL-2)-activated cell-mediated cytolysis. This effect was seen with the use of both IL-2-stimulated natural killer (NK) cells and T cells as effectors. The enhanced sensitivity of myeloma cells to cytolysis correlated with an increase in their cell surface expression of CD9, a pre-B cell marker and member of the transmembrane 4 superfamily. Incubation of PWM-stimulated myeloma cells with either monoclonal antibodies or antisense oligonucleotides directed against CD9 abrogated the effect of PWM. In order to determine whether there was a direct relationship between the expression of CD9 and enhanced sensitivity to cytolysis, myeloma cell lines that lacked CD9 expression were transfected with the CD9 gene. The level of cell surface CD9 expression correlates with enhanced susceptibility to lysis. Therefore, CD9 appears to be an important component in enhancing the sensitivity of myeloma cells to lysis mediated by IL-2-activated T cells and NK cells.
Topics: Antigens, CD; B-Lymphocytes; Calcium; Cytotoxicity, Immunologic; Humans; Immunophenotyping; Interleukin-2; Killer Cells, Natural; Lymphocyte Activation; Membrane Glycoproteins; Multiple Myeloma; Oligodeoxyribonucleotides, Antisense; Pokeweed Mitogens; Recombinant Proteins; T-Lymphocytes; Tetraspanin 29; Transfection; Tumor Cells, Cultured
PubMed: 10891455
DOI: No ID Found -
Journal of Dairy Science Jul 2003The composition and functional capacity of peripheral blood mononuclear leukocyte populations from dairy cows are altered substantially during the peripartal period....
The composition and functional capacity of peripheral blood mononuclear leukocyte populations from dairy cows are altered substantially during the peripartal period. These changes are associated with a heightened susceptibility of the mammary gland to infection. It has been postulated that the metabolic demands associated with lactogenesis may impact negatively leukocyte function during the periparturient period. In the present study, serum immunoglobulin G1 concentration and functional capacities of peripheral blood mononuclear leukocytes from intact (n = 6) and mastectomized (n = 6) periparturient Jersey cows were evaluated and compared. Cell function assessments included lymphocyte proliferation, immunoglobulin M secretion, and interferon-gamma secretion by unstimulated and pokeweed mitogen stimulated mononuclear leukocytes. Data were summarized as mean responses for 5-d periods beginning 21 d prepartum and concluding at 19 d postpartum. The progressive decrease in serum immunoglobulin G in intact but not mastectomized cows before parturition likely was attributable to the selective uptake of this isotype by the mammary gland. Lymphocyte proliferation and secretion of interferon-gamma and polyclonal IgM by mitogen-stimulated leukocytes from intact cows decreased during the 15-d period before calving, reaching a nadir at 0 to 4 d postpartum. From 5 to 19 d postpartum, these functions often were comparable to those observed 2 to 3 wk prepartum. Functions of leukocytes from mastectomized cows did not change during the study period, although they often were of lower magnitude than those of cells from nonlactating cows. These results reconfirm the occurrence of a generalized reduction in blood mononuclear leukocyte function during the periparturient period. They also suggest that the reduction in leukocyte function during the period may be, in part, due to the physiologic demands imposed on the dairy cow by the lactating mammary gland.
Topics: Animals; Calcitriol; Calcium; Cattle; DNA; Female; Immunity; Immunoglobulin G; Immunoglobulin M; Interferon-gamma; Kinetics; Leukocytes, Mononuclear; Lymphocyte Activation; Lymphocyte Subsets; Mammary Glands, Animal; Mastectomy; Parturient Paresis; Parturition; Pokeweed Mitogens; Pregnancy
PubMed: 12906053
DOI: 10.3168/jds.S0022-0302(03)73829-6 -
Proceedings of the National Academy of... May 1990Identification of human immunodeficiency virus type 1 (HIV-1)-infected individuals is of paramount importance for the control of the spread of AIDS worldwide. Currently,...
Identification of human immunodeficiency virus type 1 (HIV-1)-infected individuals is of paramount importance for the control of the spread of AIDS worldwide. Currently, the vast majority of screening centers throughout the world rely on serological techniques. As such, clinically asymptomatic but HIV-infected, seronegative individuals are rarely identified. In this report we show that 18% (30/165) of seronegative individuals who were considered to be a unique cohort of patients at high risk for HIV infection had circulating B cells that, upon in vitro polyclonal activation with pokeweed mitogen, produced antibodies reactive with HIV. Furthermore, polymerase chain reaction analysis of DNA obtained from aliquots of the peripheral blood mononuclear cells from these seronegative but pokeweed mitogen assay-positive individuals tested revealed the presence of HIV-specific sequences in a significant number of samples. In addition, depletion of CD8+ T cells from peripheral blood mononuclear cells of HIV-1-seronegative individuals prior to in vitro culture with pokeweed mitogen resulted in increased sensitivity for detecting HIV-reactive antibodies. This assay has obvious epidemiological implications, especially in the case of high-risk groups, and also provides a simple technique to enhance detection of HIV-infected individuals. Of further interest is the determination of the mechanisms related to the lack of HIV-specific antibodies in the serum of these infected individuals.
Topics: Antigens, Differentiation, T-Lymphocyte; B-Lymphocytes; Blotting, Western; CD8 Antigens; Cells, Cultured; DNA; Enzyme-Linked Immunosorbent Assay; HIV Antibodies; HIV Seropositivity; HIV-1; Humans; Lymphocyte Activation; Pokeweed Mitogens; Polymerase Chain Reaction
PubMed: 2111024
DOI: 10.1073/pnas.87.10.3972 -
Clinical and Experimental Immunology Dec 1980A simultaneous analysis was made of numbers and proportions of T cell subsets (T mu and T gamma cells), lymphocyte responsiveness to non-specific mitogens in vitro and...
A simultaneous analysis was made of numbers and proportions of T cell subsets (T mu and T gamma cells), lymphocyte responsiveness to non-specific mitogens in vitro and 'short-lived suppressor cell activity' in peripheral blood mononuclear cells (PBMC) of normal individuals. No correlation was found between either T gamma or T mu cells and the 'short-lived suppressor cell activity', suggesting that suppression in this system is not a reflection of quantitative alteration in these subsets. However, a highly significant positive correlation was found between numbers of T mu cells and PBMC responses to the mitogens phytohaemagglutinin, concanavalin A and pokeweek mitogen. This may reflect either a helper effect of T mu cells on lymphocyte proliferation in response to mitogens or the presence of the majority of mitogen-responsive cells within this subpopulation. As in normal individuals lymphocyte responsiveness correlates with the number of circulating T mu cells, it is possible that a reduction in these cells in disease states may contribute to defects in cell-mediated immunity.
Topics: Adolescent; Adult; Aged; Concanavalin A; Humans; Lectins; Leukocyte Count; Lymphocyte Activation; Middle Aged; Phytohemagglutinins; Pokeweed Mitogens; T-Lymphocytes; T-Lymphocytes, Regulatory; Time Factors
PubMed: 6452237
DOI: No ID Found -
Molecular and Cellular Biology Feb 1994A novel gene coding for the pre-B-cell colony-enhancing factor (PBEF) has been isolated from a human peripheral blood lymphocyte cDNA library. The expression of this...
A novel gene coding for the pre-B-cell colony-enhancing factor (PBEF) has been isolated from a human peripheral blood lymphocyte cDNA library. The expression of this gene is induced by pokeweed mitogen and superinduced by cycloheximide. It is also induced in the T-lymphoblastoid cell line HUT 78 after phorbol ester (phorbol myristate acetate) treatment. The predominant mRNA for PBEF is approximately 2.4 kb long and codes for a 52-kDa secreted protein. The 3' untranslated region of the mRNA has multiple TATT motifs, usually found in cytokine and oncogene messages. The PBEF gene is mainly transcribed in human bone marrow, liver tissue, and muscle. We have expressed PBEF in COS 7 and PA317 cells and have tested the biological activities of the conditioned medium as well as the antibody-purified protein in different in vitro assays. PBEF itself had no activity but synergized the pre-B-cell colony formation activity of stem cell factor and interleukin 7. In the presence of PBEF, the number of pre-B-cell colonies was increased by at least 70% above the amount stimulated by stem cell factor plus interleukin 7. No effect of PBEF was found with cells of myeloid or erythroid lineages. These data define PBEF as a novel cytokine which acts on early B-lineage precursor cells.
Topics: Amino Acid Sequence; Animals; B-Lymphocytes; Base Sequence; Cell Line; Cells, Cultured; Cloning, Molecular; Cycloheximide; Cytokines; DNA, Complementary; Gene Expression; Gene Library; Humans; Interleukin-7; Kinetics; Molecular Sequence Data; Nicotinamide Phosphoribosyltransferase; Oligonucleotide Probes; Pokeweed Mitogens; T-Lymphocytes; Tetradecanoylphorbol Acetate; Transfection
PubMed: 8289818
DOI: 10.1128/mcb.14.2.1431-1437.1994 -
Clinical and Experimental Immunology Apr 1982Monoclonal antibodies of the OKT series were used to identify T lymphocytes (OKT3+) and their inducer (OKT4+) and suppressor-cytotoxic (OKT8+) subsets in the peripheral...
Monoclonal antibodies of the OKT series were used to identify T lymphocytes (OKT3+) and their inducer (OKT4+) and suppressor-cytotoxic (OKT8+) subsets in the peripheral blood mononuclear cells (PBMC) of 32 healthy old-aged people more than 70 years old (16 men and 16 women) compared to 47 adults (29 men, 18 women) less than 40 years old. The absolute lymphocyte count in the peripheral blood was not significantly influenced by age or sex. Both the proportions and the absolute numbers of T3+ and T4+ cells were significantly lower in aged than in young participants. The proportions but not the absolute counts of OKT8+ cells were higher in the elderly. Most interesting is the influence of sex and these parameters. Old women have normal numbers and proportions of T3+, T4+ and T8+ cells when compared to young women. The latter have a significantly higher proportion of T8+ cells than young adult males. Old men have a striking reduction of both the numbers and proportions of OKT3+ and OKT4+ cells when compared with young men and with women. In addition, old men have an elevated proportion, but a normal absolute number, of OKT8+ cells. The responses of PBMC to phytohaemagglutinin extent (PHA), concanavalin A (Con A) and pokeweed mitogen (PWM) are reduced to the same extent in ageing male and female subjects when compared to young adults. In the older group, the magnitude of the lymphocyte response to PHA and Con A but not to PWM is negatively correlated with the proportions of OKT8+ cells. Surprisingly, these correlations are observed only in old women but not in old men. The latter finding excludes the possibility that the age-associated decline of the lymphocyte response to T cell mitogens is secondary to an imbalance between T4+ and T8+ lymphocytes.
Topics: Adult; Aged; Aging; Antibodies, Monoclonal; Concanavalin A; Female; Humans; Leukocyte Count; Lymphocyte Activation; Lymphocytes; Male; Phytohemagglutinins; Pokeweed Mitogens; Sex Factors; T-Lymphocytes; T-Lymphocytes, Regulatory
PubMed: 6211314
DOI: No ID Found -
Clinical and Experimental Immunology Feb 1982Recently we have described the existence of high levels of polymeric IgA, partially as immune complexes, in the serum and kidney from patients with IgA mesangial...
Recently we have described the existence of high levels of polymeric IgA, partially as immune complexes, in the serum and kidney from patients with IgA mesangial glomerulonephritis. As these patients often have macroscopic haematuria, following upper respiratory tract infections, our working hypothesis in this paper was that circulating lymphocytes from secretory tissues after viral stimulus could produce in these patients a large amount of polymeric IgA. To test it, peripheral blood lymphocytes (PBL) from patients and controls were cultured for seven days in the presence or absence of pokeweed mitogen (PWM). In cell culture supernatants immunoglobulin synthesis was measured by RIA and the proportion of polymeric and monomeric IgA was determined on Ultrogel Ac A22 column. There was no difference in spontaneous production of immunoglobulins between patients and controls. On the contrary, the IgA synthetized by PWM-stimulated PBL was significantly higher in patients than in controls. The percentages of IgA with molecular weight between 600,000 and 250,000 after supernate fractionation were significantly higher in patients than in controls. The true nature of polymeric IgA was confirmed by their ability to bind secretory component, the existence of covalent structures, and the decrease of the larger forms of IgA after reduction and alkylation. The percentage of IgA producing cells binding secretory component was significantly higher in patients than in controls (69 +/- 21 versus 44 +/- 27) after seven days of culture. IgM and IgG produced in patient culture were similar to controls. These results show that mitogen stimulated PBL from patients with Berger's disease synthetized a large amount of true polymeric IgA. It is suggested that a similar situation could occur in vivo after viral of other stimuli.
Topics: Cells, Cultured; Glomerulonephritis; Humans; Immunoglobulin A; Lymphocyte Activation; Lymphocytes; Pokeweed Mitogens; Receptors, Immunologic; Secretory Component
PubMed: 7075026
DOI: No ID Found -
Blood Sep 1981We studies the surface phenotype and the functional activities of leukemic cells from three patients with Japanese adult T-cell leukemic (ATL) using the panel of OK and...
We studies the surface phenotype and the functional activities of leukemic cells from three patients with Japanese adult T-cell leukemic (ATL) using the panel of OK and anti-Tac monoclonal antibodies, which react with differentiation antigens and define functionally distinct T-cell subsets or activated and terminally differentiated T cells. The phenotype of ATL cells were determined to be OKT1+T3+T4+T10+T5-T8-Okla1-, although cells from two patients suppressed pokeweed mitogen (PWM) induced normal B-cell differentiation, and cells from all patients lacked helper activity in this system. In addition, after cultivation with PWM, ATL cells from all patients were reactive with anti-Tac monoclonal antibody, and cells from one patient were reactive with OKlal. These findings suggest that ATL cells arise from peripheral mature T-cell subsets and also suggest that the transition of surface phenotype of ATL cells to functionally mature and activated T cells occurs in culture.
Topics: Adult; Antibodies; Antibodies, Monoclonal; Cells, Cultured; Humans; Japan; Leukemia; Phenotype; Pokeweed Mitogens; Receptors, Antigen, B-Cell; T-Lymphocytes; T-Lymphocytes, Regulatory
PubMed: 6455129
DOI: No ID Found