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PloS One 2013In Chronic Kidney Disease (CKD), immune cells are affected by uremic retention toxins. Given this effect, we analyzed lymphocyte proliferative response and immune...
INTRODUCTION
In Chronic Kidney Disease (CKD), immune cells are affected by uremic retention toxins. Given this effect, we analyzed lymphocyte proliferative response and immune modulators production following in vitro stimulation.
METHODS
Whole blood was drawn from healthy controls, patients with eGFR <20 ml/min/1.73 m(2) (Pre-dialysis, CKD stages 4 and 5) and hemodialysis patients (stage 5D). Peripheral cells were incubated for six days with pokeweed mitogen, concanavalin A, Staphylococcus enterotoxin A or influenza A vaccine. Peripheral lymphocyte proliferation was then analyzed by the "Flow-cytometric Assay of Specific Cell-mediated Immune response in Activated whole blood" (FASCIA) method, and cytokine profile in the cell supernatants was analyzed by the Milliplex multi-array method.
RESULTS
The absolute number of lymphoblasts in response to mitogenic stimulation and the number of cells in each CD4+ and CD8+ subpopulation were similar comparing the three groups, except for a single decline in number of lymphoblasts after stimulation with Staphylococcus enterotoxin A, comparing dialysis patients with healthy controls. Levels of interleukin (IL)-2 (p=0.026), -10 (p=0.019) and -15 (p=0.027) in the Staphylococcus enterotoxin A-stimulated supernatant were lower in hemodialysis patients compared to healthy controls. Levels of IL-15 (p=0.017) from pre-dialysis patients and levels of IL-5 (p=0.019) from hemodialysis patients in influenza A vaccine-stimulated supernatants were also lower compared to controls. In pokeweed mitogen-stimulated supernatant, IL-2 levels (p=0.013) were lower in hemodialysis patients compared to pre-dialysis patients. TNF-α, IL-10, IL-12, IL-15, IL-8, MCP-1, IP-10, IFN-α2, IL-1α and eotaxin levels were all significantly higher in plasma obtained from CKD patients.
CONCLUSION
Our results suggest that T-cells from CKD patients have similar proliferative response to stimulation compared with healthy individuals. Moreover, however the immune cells show inability to produce selected cytokines, most likely due to the uremic milieu or dialysis procedure.
Topics: Adult; Aged; CD4-Positive T-Lymphocytes; CD8-Positive T-Lymphocytes; Case-Control Studies; Cell Proliferation; Cells, Cultured; Concanavalin A; Cytokines; Enterotoxins; Female; Glomerular Filtration Rate; Humans; Influenza Vaccines; Male; Middle Aged; Mitogens; Pokeweed Mitogens; Renal Dialysis; Renal Insufficiency, Chronic
PubMed: 23951343
DOI: 10.1371/journal.pone.0073141 -
Immunology Feb 1992Monocyte subpopulations which differ in the expression of Fc receptor for human IgG (FcRI) differentially regulate the T-cell-dependent, pokeweed mitogen (PWM)-induced,...
Monocyte subpopulations which differ in the expression of Fc receptor for human IgG (FcRI) differentially regulate the T-cell-dependent, pokeweed mitogen (PWM)-induced, polyclonal B-cell response. We, thus, studied the cytokine production in human peripheral blood monocyte and T-lymphocyte cultures activated with this lectin. Monocytes or their FcR+ and FcR- subpopulations stimulated with PWM were cultured with or without T lymphocytes or their CD4+ and CD8+ subsets. Both monocyte subpopulations cultured alone produced similar amounts of tumour necrosis factor-alpha (TNF-alpha) and interleukin-6 (IL-6), but FcR- monocytes showed significantly enhanced ability to secrete interleukin-1 (IL-1). T cells, especially CD4+, added to monocyte cultures enhanced IL-1 production. This enhancement was presumably due to interferon-gamma (IFN-gamma) release by T lymphocytes, since this lymphokine enhanced IL-1 secretion when added to PWM-stimulated cultures of monocytes. Addition of monocytes, in particular the FcR+ subpopulation, greatly enhanced production of IFN-gamma by T lymphocytes. Although both T-cell subsets produced IFN-gamma, the CD4+ cells were more efficient. These results indicate that in PWM-stimulated cultures subpopulations of monocytes differ in secretion of cytokines, which might explain their differential effect on T-cell-dependent immune responses in vitro.
Topics: Antigens, Differentiation; Cell Communication; Cells, Cultured; Humans; Interleukin-1; Interleukin-6; Monocytes; Monokines; Pokeweed Mitogens; Receptors, Fc; Receptors, IgG; T-Lymphocytes; Tumor Necrosis Factor-alpha
PubMed: 1532381
DOI: No ID Found -
Clinical and Experimental Immunology Aug 1989The relation between the percentage of circulating CD5+ CD20+ B cells and the ability to synthesize IgM and IgM rheumatoid factor (RF) in vitro in response to pokeweed...
The relation between the percentage of circulating CD5+ CD20+ B cells and the ability to synthesize IgM and IgM rheumatoid factor (RF) in vitro in response to pokeweed mitogen (PWM) and Staphylococcus aureus Cowan I (SAC) was assessed in 21 healthy controls. CD5+ CD20+ cells ranged from 7.3 to 19.9% of total CD20+ B cells. By Spearman's rank correlation, there was an inverse relation between the percentage of CD5+ CD20+ B cells and IgM production in response to PWM (rs = -0.452, p less than 0.05) and a direct correlation with RF production in response to SAC (rs = 0.450, P less than 0.05). The percentage of CD5+ CD20+ B cells was not related to any serologic HLA-A, B, C or D antigen. Healthy individuals may be predisposed to producing IgM or autoantibodies based on the percentage of circulating CD5+ Cd20+ B cells.
Topics: Antigens, CD20; Antigens, Differentiation; Antigens, Differentiation, B-Lymphocyte; B-Lymphocytes; CD5 Antigens; Humans; Immunoglobulin M; Pokeweed Mitogens; Rheumatoid Factor; Staphylococcus aureus
PubMed: 2476269
DOI: No ID Found -
The Journal of Experimental Medicine Mar 1982Monoclonal antibodies were used to examine the immunoglobulin isotypes expressed by B lymphocyte precursors of IgM, IgG, IgA, and IgA2 plasma cells. Plasma-cell...
Human b-cell differentiation. I. Analysis of immunoglobulin heavy chain switching using monoclonal anti-immunoglobulin M, G, and A antibodies and pokeweed mitogen-induced plasma cell differentiation.
Monoclonal antibodies were used to examine the immunoglobulin isotypes expressed by B lymphocyte precursors of IgM, IgG, IgA, and IgA2 plasma cells. Plasma-cell differentiation was induced by the addition of pokeweed mitogen to cultures of blood mononuclear cells. Anti-mu, -gamma, -alpha, and -alpha 1 antibodies were used in some experiments to inhibit differentiation of B lymphocytes bearing these heavy chain isotypes, and for selective removal of B lymphocyte precursors before culture with pokeweed mitogen in other experiments. Three major subpopulations of B lymphocyte precursors were identified: (a) a subpopulation of surface (s) IgM+ precursors of IgM plasma cells that did not express IgG or IgA isotypes, (b) a subpopulation of sIgG+ precursors of IgG plasma cells of which approximately one-half bore some IgM and none had detectable IgA receptors, and (c) a subpopulation of sIgA+ precursors of IgA plasma cells; one half of these precursors could be shown to express functional IgM receptors but none were found to express IgG receptors. The sIgA subpopulation could be further subdivided into sIgA1+ precursors of IgA1 plasma cells and IgA1-negative precursors of IgA2 plasma cells. These results suggest that normal human B cells can switch from mu directly to each of the other heavy chain isotypes, and that these represent the main switch pathways.
Topics: Animals; Antibodies, Anti-Idiotypic; Antibodies, Monoclonal; B-Lymphocytes; Cell Differentiation; Humans; Immunoglobulin A; Immunoglobulin G; Immunoglobulin Heavy Chains; Immunoglobulin M; Immunoglobulin alpha-Chains; Immunoglobulin gamma-Chains; Immunoglobulin mu-Chains; Immunoglobulins; Lymphocyte Activation; Mice; Mice, Inbred Strains; Plasma Cells; Pokeweed Mitogens; Receptors, Antigen, B-Cell
PubMed: 6801181
DOI: 10.1084/jem.155.3.839 -
Clinical and Vaccine Immunology : CVI Mar 2006Commercially available pokeweed mitogen (PWM) has been reported to activate macrophages, leading to production of proinflammatory cytokines and nitric oxide (NO).... (Comparative Study)
Comparative Study
Commercially available pokeweed mitogen (PWM) has been reported to activate macrophages, leading to production of proinflammatory cytokines and nitric oxide (NO). However, we found that polymyxin B (PMB), a specific inhibitor of endotoxin activity, inhibited the PWM-induced expression of proinflammatory cytokines and NO and the activation of Toll-like receptor 4 (TLR4). A kinetic-turbidimetric Limulus amebocyte lysate assay demonstrated that commercial PWM contained substantial endotoxin, over 10(4) endotoxin units/mg of the PWM. A PWM repurified by PMB-coupled beads no longer induced the expression of proinflammatory cytokines, TLR4 activation, or dendritic cell maturation. However, the repurified PWM remained able to induce proliferation of human lymphocytes, which is a representative characteristic of PWM. These results suggest that commercial PWM might be contaminated with a large amount of endotoxin, resulting in the attribution of misleading immunological properties to PWM.
Topics: Animals; Cell Differentiation; Cell Line; Cell Proliferation; Cells, Cultured; Dendritic Cells; Dose-Response Relationship, Immunologic; Drug Contamination; Endotoxins; Humans; Macrophage Activation; Macrophages; Mice; Phytolacca americana; Pokeweed Mitogens
PubMed: 16522770
DOI: 10.1128/CVI.13.3.309-313.2006 -
Immunology Aug 1979Saline extracts from the roots of the pokeweed species. Phytolacca octandra were separated by ion-exchange chromatography into three fractions, Po-1, Po-2 and Po-3. Po-1...
Saline extracts from the roots of the pokeweed species. Phytolacca octandra were separated by ion-exchange chromatography into three fractions, Po-1, Po-2 and Po-3. Po-1 contained two monomeric proteins with molecular weights of 36,000 and 29,000 and these were partially purified by gel filtration. Po-2 was purified as a single polymeric protein composed of approximately ten 14,000 mol. wt polypeptides and is a new pokeweed mitogen. Po-3 was purified as a single polymeric protein composed of approximately four 31,000 mol. wt subunits, and apart from its polymeric structure closely resembles commercial pokeweed mitogen (PWM). Po-2 and Po-3 were mitogenic for unseparated human peripheral blood lymphocytes but the degree of mitogenic activity in Po-2 preparations was dependent on storage following purification. Purified B cells were not stimulated by either mitogen. Po-3 was a potent mitogen for T cells but preparations of Po-2 required storage before they stimulated T cells. Higher responses were observed in co-cultures of B and T cells than in separated B and T cell cultures. It is suggested that human B and T lymphocytes show synergy in their responses to Po-2 and Po-3.
Topics: Amino Acids; B-Lymphocytes; Cells, Cultured; Dose-Response Relationship, Immunologic; Electrophoresis, Polyacrylamide Gel; Humans; Mitosis; Molecular Weight; Plant Lectins; Plants; Pokeweed Mitogens; T-Lymphocytes
PubMed: 315368
DOI: No ID Found -
Clinical and Experimental Immunology Jul 1984A system has been established to produce in vitro IgG specific for cell wall determinants of S. aureus by human peripheral blood mononuclear cells (PBMC). PBMC of...
A system has been established to produce in vitro IgG specific for cell wall determinants of S. aureus by human peripheral blood mononuclear cells (PBMC). PBMC of healthy individuals, of patients with S. aureus infections and of patients with other bacterial infections were cultured for 12 days. In the culture supernatants (SN) total IgG was determined by a competitive RIA, and IgG to purified cell walls (PCW) of S. aureus strain H by a two step ELISA. PBMC of 11 healthy persons produced anti-PCW IgG upon stimulation by pokeweed mitogen (PWM). This indicates the presence in some healthy persons of circulating B cells which can be induced in vitro to synthesize PCW specific IgG. PBMC of S. aureus infected patients, however, synthesized anti-PCW IgG in culture medium alone. This is the first description of spontaneous in vitro production of specific IgG during a bacterial infection and may be analogous to short phases of spontaneous specific IgG production described after immunizations and viral infections. Finally, compared to healthy individuals, an increased total IgG synthesis in vitro by PBMC obtained from patients with S. aureus and from patients with other severe bacterial infections was found. It is concluded that this polyclonal B cell activation has been initiated in vivo. Its biological significance is unknown.
Topics: Adult; Antibodies, Bacterial; Bacterial Infections; Cell Wall; Cells, Cultured; Epitopes; Female; Humans; Immunoglobulin G; Lymphocytes; Male; Middle Aged; Pokeweed Mitogens; Staphylococcal Infections; Staphylococcus aureus; Time Factors
PubMed: 6204800
DOI: No ID Found -
Clinical and Experimental Immunology Aug 1989A characteristic of active cytomegalovirus (CMV) infection is its suppressive effect on in vitro assays of immune function. The expression of CD11b by the Cd4+ and Cd8+...
Phenotypic study of CD4+ and CD8+ lymphocyte subsets in relation to cytomegalovirus carrier status and its correlate with pokeweed mitogen-induced B lymphocyte differentiation.
A characteristic of active cytomegalovirus (CMV) infection is its suppressive effect on in vitro assays of immune function. The expression of CD11b by the Cd4+ and Cd8+ lymphocytes allows the identification of subsets with distinct regulatory functions of pokeweed mitogen (PWM) induced B cell differentiation. In order to relate that result with our previous observation that CMV carriers have significantly increased numbers of CD4+, HNK1+ and CD8+, HNK1+ lymphocytes in their peripheral blood compared with non-carriers, we performed a three-colour flow cytometric analysis of the co-expression of Cd11b and HNK1 by CD4+ and CD8+ lymphocytes obtained from 27 CMV carriers and 42 non-carriers. The differences between CMV carriers and non-carriers were significant for the CD4+, HNK1+ lymphocytes (median [5th and 95th percentiles], 59 [18 and 123 versus 24/7 and 73 per mm3, respectively; P less than 0.001) and CD8+, HNK1+ lymphocytes (59 [18 259] versus 52 [23 and 139] per mm3; P less than 0.001), but not for the CD4+, CD11b+ lymphocytes (59 [18 and 135] versus 52 [17 and 104] per mm3) and the CD8+, CD11b+ lymphocytes (85 [34 and 293] versus 82 [21 and 248] per mm3). The CD4+, HNK1+ and CD8+, HNK1+ lymphocytes that were increased in CMV carriers compared with non-carriers included mostly CD11b-, but also CD11b+ lymphocytes. After sorting CD4+ and CD8+ lymphocytes for four CMV carriers into HNK1+ and HNK1- fractions, we analyzed their regulatory functions on PWM-driven B cell Helper function to PWM-driven B cell differentiation was exclusively associated with the CD4+, HNK1- lymphocytes; the CD4+, HNK1+ generally did not show helper or suppressor activity in this assay. Both CD8+, HNK1+ and CD8+, HNK1- lymphocytes showed suppressor activity. Thus, the NHK1 marker does not constitute a phenotypical correlate for suppressor cells of PWM-driven B-cell differentiation.
Topics: Adolescent; Adult; Antigens, Differentiation; Antigens, Differentiation, T-Lymphocyte; B-Lymphocytes; CD57 Antigens; CD8 Antigens; Carrier State; Cytomegalovirus Infections; Female; Humans; Lymphocyte Activation; Macrophage-1 Antigen; Male; Membrane Glycoproteins; Pokeweed Mitogens; T-Lymphocytes
PubMed: 2476271
DOI: No ID Found -
Clinical and Experimental Immunology Sep 1978Lymphocytes from sheep in late pregnancy and at parturition showed markedly impaired proliferative responses to phytohaemagglutinin (PHA) in vitro when cultures were...
Lymphocytes from sheep in late pregnancy and at parturition showed markedly impaired proliferative responses to phytohaemagglutinin (PHA) in vitro when cultures were supplemented with foetal bovine serum (FBS), as compared to the responses of lymphocytes from non-pregnant sheep, sheep at 40 days of gestation and sheep at 80 days of gestation. Similar responses to PHA were observed when the medium was supplemented with autologous plasma (AP), although the responses were of a lower order. In both cases elevated responses to PHA were apparent at 10 days post-parturition. The response with FBS was more marked than with AP. Progressive reduction of lymphocyte responses to pokeweed mitogen (PWM) in the presence of FBS and AP were less obvious, although it was still apparent that responses to PWM were depressed at 120 days of gestation and at parturition, when compared with lymphocyte responses during early pregnancy (at 40 days and 80 days of gestation). The difference was much more apparent with AP than with FBS and responses during early pregnancy were markedly higher than those with FBS. An increase in lymphocyte responsiveness to PWM 10 days post-parturition was evident whether FBS or AP was incorporated in the cultures. The response with FBS was again more marked than with AP.
Topics: Animals; Female; Lymphocyte Activation; Lymphocytes; Phytohemagglutinins; Pokeweed Mitogens; Postpartum Period; Pregnancy; Pregnancy, Animal; Sheep
PubMed: 737895
DOI: No ID Found -
Journal of Animal Science Aug 2021Disease resilience refers to the productivity of an animal under disease. Given the high biosecurity of pig nucleus herds, traits that can be measured on healthy pigs...
Disease resilience refers to the productivity of an animal under disease. Given the high biosecurity of pig nucleus herds, traits that can be measured on healthy pigs and that are genetically correlated with disease resilience, that is, genetic indicator traits, offer a strategy to select for disease resilience. Our objective was to evaluate mitogen stimulation assays (MSAs) on peripheral blood mononuclear cells (PBMCs) from young healthy pigs as genetic indicators for disease resilience. Data were from a natural disease challenge in which batches of 60 or 75 naïve Yorkshire × Landrace piglets were introduced every 3 wk into a continuous flow barn that was seeded with multiple diseases. In this environment, disease resilience traits, including growth, treatment, and mortality rates, were recorded on 3,136 pigs that were genotyped with a high-density marker panel. PBMCs from 882 of these pigs from 19 batches were isolated from whole blood collected prior to the disease challenge and stimulated with five mitogens: concanavalin A (ConA), phytohemagglutinin (PHA), pokeweed mitogen (PWM), lipopolysaccharide (LPS), and phorbol myristate acetate (PMA). The proliferation of cells was evaluated at 48, 72, and 96 h and compared with unstimulated samples (rest count). Heritabilities of cell proliferation were estimated using a model with batch as a fixed effect and covariates of entry age; rest count; complete blood count proportions of lymphocytes, monocytes, eosinophils, and basophils; and pen, litter, and animal genetics as random effects. Heritability estimates were highest for response to ConA (0.30 ± 0.09, 0.28 ± 0.10, 0.17 ± 0.10, and 0.25 ±0.10 at 48, 72, and 96 h after stimulation and for area under the curve across the three time points, respectively). Estimates were in a similar range for response to PHA and PMA but low for PWM and LPS. Responses to ConA, PHA, and PMA were moderately genetically correlated with several disease resilience traits and in the expected direction, but individual estimates were not significantly different from zero due to large SEs. In conclusion, although validation is needed, MSAss, in particular based on ConA, show promise as genetic indicator traits for disease resilience.
Topics: Animals; Cell Proliferation; Leukocytes, Mononuclear; Lymphocyte Activation; Mitogens; Phytohemagglutinins; Pokeweed Mitogens; Swine
PubMed: 33944943
DOI: 10.1093/jas/skab084