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Sample records for human lymphocyte subsets

  1. Restricted expression of LW antigen on subsets of human B and T lymphocytes.

    PubMed

    Oliveira, O L; Thomas, D B; Lomas, C G; Tippett, P

    1984-01-01

    NIM-M8 is a monoclonal IgM antibody, specific for the LWab antigen as shown by its reaction with red cells of all donors except those lacking LWa, LWb and LWab. Indirect immunofluorescent staining and cell sorter analyses have shown that LWab is present on a subpopulation of human lymphocytes. Cell fractionation studies indicate that subsets of both B and T cells express LWab and it may, therefore, provide a further marker for heterogeneity in these lymphocyte populations. PMID:6443217

  2. Interaction between nitric oxide and subsets of human T lymphocytes with differences in glutathione metabolism

    PubMed Central

    Roozendaal, Ramon; Kauffman, Henk F; Dijkhuis, Anne-Jan; Ommen, Elisabeth T V; Postma, Dirkje S; De Monchy, Jan G R; Vellenga, Edo

    2002-01-01

    Nitric oxide (NO) modulates human T-lymphocyte responses through several mechanisms. In the current study we show that interactions between NO and glutathione (GSH) metabolism are related to the selective persistent inhibition of interferon-γ (IFN-γ) production by NO, which we previously identified. T cells were exposed to NO using the NO-donor compound Spermine-nonoate (Sper) and activated using anti-CD3 plus anti-CD28 monoclonal antibodies. Persistent inhibition of IFN-γ by Sper was prevented by addition of the GSH precursor l-cysteine, which inhibits Sper induced GSH depletion. Subsets of cells were either susceptible (GSHlow) or resistant (GSHhigh) to NO-induced GSH depletion. The GSHlow subset was characterized by enhanced numbers of CD4+ cells, reduced numbers of activated cells as characterized by CD25 and CD69, and reduced numbers of memory (CD45RO+) cells relative to the GSHhigh population. Rather than directly affecting susceptibility to NO, these surface markers reflected different expression patterns. Particularly, the GSHlow subset was further characterized by decreased activity of the GSH synthesis related enzymes multi-drug resistance related protein (MRP)-1 and γ-glutamyltranspeptidase (γ-GT). Blocking γ-GT, using acivicin was shown to exacerbate NO-induced GSH depletion and NO-induced apoptosis. Since NO induced apoptosis selectively affects IFN-γ production these findings implicate GSH metabolism in the modulation and maintenance of the T helper (Th)1/Th2 balance. PMID:12423309

  3. Effects of human immunodeficiency virus type 1 on CD4 lymphocyte subset activation.

    PubMed

    Masci, A M; Paz, F L; Borriello, A; Cassano, S; Della Pietra, V; Stoiber, H; Matarese, G; Della Ragione, F; Zappacosta, S; Racioppi, L

    1999-06-01

    The pathogenesis of the decline of CD4 lymphocyte counts accompanying the typical course of HIV-1 infection is not completely defined and might be related to a differential susceptibility of naive and memory cells to HIV-1 exposure. Here, we examined the effects induced by heat-inactivated HIV-1 virions on these lymphocyte populations. Exposure of CD45RA naive T cells to inactivated viral particles induced a marked decrease of both mitogenic responses and activation-induced apoptosis. Conversely, the growth of CD45RO cells was less severely restrained. Analysis of intracellular levels of cell cycle regulatory proteins revealed an arrest at the G1/S restriction point of the naive but not memory subset. This effect was associated with alterations in phosphotyrosine profile and with a marked decrease of ERK and NJK kinase activation. Finally, up-regulation of the cAMP-dependent protein kinase A (PKA) activity induced by mitogens was not affected by virus. Altogether, these findings show that interaction of HIV-1 with the T cell surface is sufficient to inhibit the proliferative response of the CD4CD45RA subset by disturbing proximal TCR signaling. This mechanism would affect renewal of naive lymphocytes, contributing in such a way to the impairment of T cell turnover during the course of HIV-1 infection. PMID:10382750

  4. Locomotor responses of human CD45 lymphocyte subsets: preferential locomotion of CD45RO+ lymphocytes in response to attractants and mitogens.

    PubMed Central

    Newman, I; Wilkinson, P C

    1993-01-01

    The CD45RO+ population of lymphocytes from human blood contains a higher proportion of locomotor cells than the CD45RA+ population. Direct from blood there were few locomotor lymphocytes (< 15%), but, among these, a higher proportion of CD45RO+ than of CD45RA+ cells responded to the chemotactic stimuli, foetal calf serum (FCS) and interleukin-2 (IL-2) in polarization assays. Likewise, after overnight culture, a higher proportion of CD45RO+ cells responded to IL-8. Culture for 24-72 hr in activators such as anti-CD3, purified protein derivative (PPD), phytohaemagglutinin (PHA), concanavalin A (Con A), pokeweed mitogen (PWM) or in an allogeneic mixed leucocyte reaction (AMLR) increased the proportion of locomotor lymphocytes to 20-60%, and the CD45RO+ subset showed proportionately more polarized cells than the CD45RA+ subset after culture with all the above activators. Preferential migration of CD45RO+ cells into collagen gels was also seen after culture in antigenic stimuli (PPD or AMLR) but not with polyclonal activators (alpha CD3 or Con A). Double labelling showed that, within the CD4+ and CD8+ subsets, antigen-stimulated CD45RO+ T cells invaded collagen gels in higher proportions than CD45RA+ T cells. Clustering of lymphocytes with accessory cells is an essential prerequisite for locomotion and, after culture in alpha CD3, CD45RO+ lymphocytes were found preferentially in clusters with monocytes. In all of the above populations, CD45RO+ lymphocytes were larger in size. These findings suggest that, not only selective adhesion to vascular endothelium as reported earlier, but also selective locomotion recruits CD45RO+ lymphocytes into sites of inflammation. PMID:8436407

  5. Studies on human blood lymphocytes with iC3b (type 3) complement receptors. II. Characterization of subsets which regulate pokeweed mitogen-induced lymphocyte proliferation and immunoglobulin synthesis.

    PubMed Central

    Abo, W; Gray, J D; Bakke, A C; Horwitz, D A

    1987-01-01

    Human blood lymphocytes that express Type 3 complement receptors (CR3) can be divided into a major subset with high density Fc receptors for IgG (FcR) identified with the monoclonal antibody Leu 11 and two minor subsets which display either CD8 (Leu 2) or CD4 (Leu 3) markers. We isolated CR3+ lymphocyte subsets and examined them for regulatory effects on pokeweed mitogen (PWM) stimulated cells. The FCR CR3+ cell suppressed PWM-induced proliferation and Ig production. Pretreatment of these lymphocytes with immune complexes was required to suppress proliferation, but not IgG production. The CR3+ Leu 2+ FCR- subset also had suppressive activity, but this effect was not observed unless the CR3+ Leu 3+ enriched subset was removed. In fact, the CR3+ Leu 3+ enriched subset enhanced IgG synthesis. Brief exposure of CR3+ lymphocytes to recombinant interleukin 2, recombinant alpha-interferon, but not gamma-interferon, markedly enhanced the inhibitory effect. Time course studies and a comparison of inhibition of Ig synthesis with natural killer cell activity suggested that CR3+ lymphocytes act shortly after lymphocytes are exposed to PWM and that Ig production was regulated by suppression rather than cytotoxicity. These CR3+ lymphocyte subsets may have broad antigen non-specific effects on immunoglobulin synthesis. PMID:2955973

  6. Human spleen contains different subsets of dendritic cells and regulatory T lymphocytes

    PubMed Central

    Velásquez-Lopera, M M; Correa, L A; García, L F

    2008-01-01

    Most knowledge about dendritic cells (DCs) and regulatory T cells in humans has been gathered from circulating cells but little is known about their frequency and distribution in lymphoid organs. This report shows the frequency, phenotype and location of DCs and regulatory T cells in deceased organ donors' spleens. As determined by flow cytometry, conventional/myeloid DCs (cDCs) CD11chighHLA-DR+CD123−/low were 2·3 ± 0·9% and LIN- HLA-DR+CD11chigh 2·1 ± 0·3% of total spleen cells. Mature CD11chighHLA-DR+CD83+ were 1·5 ± 0·8% and 1·0 ± 1·6% immature CD11chighHLA-DR+CD83- cDC. There were 0·3 ± 0·3% plasmacytoid DCs (pDC) CD11c−/lowHLA-DR+CD123high and 0·3 ± 0·1% LIN-HLA-DR+CD123high. Cells expressing cDCs markers, BDCA-1 and BDCA-3, and pDCs markers BDCA-2 and BDCA-4 were observed in higher frequencies than DCs with other phenotypes evaluated. CD11c+, CD123+ and CD83+ cells were located in subcapsular zone, T cells areas and B-cell follicles. CD4+CD25high Tregs were 0·2 ± 0·2% and CD8+CD28- comprised 11·5 ± 8·1% of spleen lymphocytes. FOXP3+ cells were found in T- and B-cell areas. The improvement in cell separation, manipulation and expansion techniques, will facilitate the manipulation of donor spleen cells as a part of protocols for induction and maintenance of allograft tolerance or treatment of autoimmune diseases. PMID:18727627

  7. Relationships over 1 year between lymphocyte subsets and psychosocial variables among adults with infection by human immunodeficiency virus.

    PubMed

    Perry, S; Fishman, B; Jacobsberg, L; Frances, A

    1992-05-01

    To examine relationships between immune and psychosocial variables among adults infected with human immunodeficiency virus type 1, 221 subjects without acquired immunodeficiency syndrome were assessed for degree of depression, anxiety, psychiatric symptoms, social support, stressful life events, hardiness, hopelessness, bereavement, and intrusive and avoidant thoughts about acquired immunodeficiency syndrome. At entry, none of 22 psychosocial variables significantly correlated with lymphocyte subsets. Among subjects seen 6 and 12 months later, severity of physical symptoms was associated with greater emotional distress, but the CD4 cell count was predicted by neither clinical ratings of psychopathology and global functioning nor by standardized self-report measures of constructs used in psychoimmune research. We conclude that among our sample, physical symptoms contributed to emotional distress, but emotional distress did not contribute to the CD4 cell count, a marker of disease progression. PMID:1586275

  8. Cell surface Glut1 levels distinguish human CD4 and CD8 T lymphocyte subsets with distinct effector functions

    PubMed Central

    Cretenet, Gaspard; Clerc, Isabelle; Matias, Maria; Loisel, Severine; Craveiro, Marco; Oburoglu, Leal; Kinet, Sandrina; Mongellaz, Cédric; Dardalhon, Valérie; Taylor, Naomi

    2016-01-01

    CD4 and CD8 T lymphocyte activation requires the generation of sufficient energy to support new biosynthetic demands. Following T cell receptor (TCR) engagement, these requirements are met by an increased glycolysis, due, at least in part, to induction of the Glut1 glucose transporter. As Glut1 is upregulated on tumor cells in response to hypoxia, we assessed whether surface Glut1 levels regulate the antigen responsiveness of human T lymphocytes in both hypoxic and atmospheric oxygen conditions. Notably, Glut1 upregulation in response to TCR stimulation was significantly higher in T lymphocytes activated under hypoxic as compared to atmospheric oxygen conditions. Furthermore, TCR-stimulated human T lymphocytes sorted on the basis of Glut1-Lo and Glut1-Hi profiles maintained distinct characteristics, irrespective of the oxygen tension. While T cells activated in hypoxia divided less than those activated in atmospheric oxygen, Glut1-Hi lymphocytes exhibited increased effector phenotype acquisition, augmented proliferation, and an inverted CD4/CD8 ratio in both oxygen conditions. Moreover, Glut1-Hi T lymphocytes exhibited a significantly enhanced ability to produce IFN-γ and this secretion potential was completely dependent on continued glycolysis. Thus, Glut1 surface levels identify human T lymphocytes with distinct effector functions in both hypoxic and atmospheric oxygen tensions. PMID:27067254

  9. Lymphocytes subsets reference values in childhood.

    PubMed

    Tosato, F; Bucciol, G; Pantano, G; Putti, M C; Sanzari, M C; Basso, G; Plebani, M

    2015-01-01

    Immunophenotyping of blood lymphocyte subsets and activation markers is a basic tool in the diagnostic process of primary immunodeficiency diseases, its use becoming more and more widespread as the knowledge about these illnesses increases. However, the availability of reliable reference values, which need to be age-matched for the pediatric population, is a pre-requisite for the reliable interpretation of immunophenotyping data. Aim of this study is to analyze the lymphocyte subsets and activation markers distribution in children aged 0-18 years referring to the University Hospital of Padova and to create age-matched reference values expressed by percentiles, thus providing a valuable guideline for the interpretation of the immunophenotype. PMID:25132325

  10. Investigating chromosome damage and gammaH2AX response in human lymphocytes and lymphocyte subsets as potential biomarkers of radiation sensitivity

    NASA Astrophysics Data System (ADS)

    Beaton, Lindsay A.

    This thesis examines in vitro irradiated blood samples from prostate cancer patients exhibiting late normal tissue damage after receiving radiotherapy, for lymphocyte response. Chromosomal aberrations, translocations and proliferation rate are measured, as well as gammaH2AX response in lymphocytes and lymphocyte subsets. The goal of this thesis is to determine whether the lymphocyte response to in vitro radiation could be used as a marker for radiosensitivity. Patients were selected from a randomized clinical trial evaluating the optimal timing of Dose Escalated Radiation and short course Androgen Deprivation Therapy. Of 438 patients, 3% developed Grade 3 late radiation proctitis and were considered to be radiosensitive. Blood was drawn from 10 of these patients along with 20 matched samples from patients with grade 0 proctitis. The samples were irradiated and were analyzed for dicentric chromosomes, excess fragments and proliferation rates (at 6 Gy), translocations, stable and unstable damage (at 4 Gy), and dose response (up to 10 Gy), along with time response after 2 Gy (0 -- 24 h). Chromosome aberrations, excess fragments per cell, translocations per cell and proliferation rates were analyzed by brightfield and fluorescent microscopy, while the gammaH2AX response in lymphocytes and lymphocyte subsets was analyzed by flow cytometry. Both groups were statistically similar for all endpoints at 0 Gy. At 6 Gy, there were statistically significant differences between the radiosensitive and control cohorts for three endpoints; the mean number of dicentric chromosomes per cell, the mean number of excess fragments per cell and the proportion of cells in second metaphase. At 4 Gy, there were statistically significant differences between the two cohorts for three endpoints; the mean number of translocations per cell, the mean number of dicentric chromosomes per cell and the mean number of deletions per cell. There were no significant differences between the gammaH2AX

  11. Diagnostic utility of lymphocyte subset analysis in AIDS case finding.

    PubMed Central

    Boyko, W J; Schechter, M T; MacLeod, A; Douglas, B; Maynard, M; Sharp, R; Wiggs, B

    1986-01-01

    Abnormalities of lymphocyte subsets, especially low absolute number of helper T cells, are characteristically present in acquired immune deficiency syndrome (AIDS). Similar abnormalities can be found in patients with persistent generalized lymphadenopathy (PGL) or AIDS-related complex (ARC) and, to a lesser degree, in asymptomatic people who have been exposed to human T-lymphotropic virus type III (HTLV-III). Nevertheless, there appears to be a widespread perception that lymphocyte subset analysis may be useful in AIDS case finding within high-risk groups. We evaluated the diagnostic utility of absolute number of helper T cells and ratio of helper to suppressor T cells in 33 patients with AIDS, 43 patients with PGL who had been referred for lymph node biopsy, 90 patients with PGL and 195 male homosexual controls. At conventional cutoff levels the tests did not appear to revise the probability of AIDS upward to any clinically significant degree when the pretest probability of AIDS was low. Lymphocyte subset analysis does not appear to be a cost-effective method of AIDS case finding in identified groups at risk in which the prevalence of AIDS is low. PMID:2938707

  12. Distinct microRNA signatures in human lymphocyte subsets and enforcement of the naive state in CD4+ T cells by the microRNA miR-125b.

    PubMed

    Rossi, Riccardo L; Rossetti, Grazisa; Wenandy, Lynn; Curti, Serena; Ripamonti, Anna; Bonnal, Raoul J P; Birolo, Roberto Sciarretta; Moro, Monica; Crosti, Maria C; Gruarin, Paola; Maglie, Stefano; Marabita, Francesco; Mascheroni, Debora; Parente, Valeria; Comelli, Mario; Trabucchi, Emilio; De Francesco, Raffaele; Geginat, Jens; Abrignani, Sergio; Pagani, Massimiliano

    2011-08-01

    MicroRNAs are small noncoding RNAs that regulate gene expression post-transcriptionally. Here we applied microRNA profiling to 17 human lymphocyte subsets to identify microRNA signatures that were distinct among various subsets and different from those of mouse lymphocytes. One of the signature microRNAs of naive CD4+ T cells, miR-125b, regulated the expression of genes encoding molecules involved in T cell differentiation, including IFNG, IL2RB, IL10RA and PRDM1. The expression of synthetic miR-125b and lentiviral vectors encoding the precursor to miR-125b in naive lymphocytes inhibited differentiation to effector cells. Our data provide an 'atlas' of microRNA expression in human lymphocytes, define subset-specific signatures and their target genes and indicate that the naive state of T cells is enforced by microRNA. PMID:21706005

  13. CD4CD8αα lymphocytes, a novel human regulatory T cell subset induced by colonic bacteria and deficient in patients with inflammatory bowel disease.

    PubMed

    Sarrabayrouse, Guillaume; Bossard, Céline; Chauvin, Joe-Marc; Jarry, Anne; Meurette, Guillaume; Quévrain, Elodie; Bridonneau, Chantal; Preisser, Laurence; Asehnoune, Karim; Labarrière, Nathalie; Altare, Frédéric; Sokol, Harry; Jotereau, Francine

    2014-04-01

    How the microbiota affects health and disease is a crucial question. In mice, gut Clostridium bacteria are potent inducers of colonic interleukin (IL)-10-producing Foxp3 regulatory T cells (Treg), which play key roles in the prevention of colitis and in systemic immunity. In humans, although gut microbiota dysbiosis is associated with immune disorders, the underlying mechanism remains unknown. In contrast with mice, the contribution of Foxp3 Treg in colitis prevention has been questioned, suggesting that other compensatory regulatory cells or mechanisms may exist. Here we addressed the regulatory role of the CD4CD8 T cells whose presence had been reported in the intestinal mucosa and blood. Using colonic lamina propria lymphocytes (LPL) and peripheral blood lymphocytes (PBL) from healthy individuals, and those with colon cancer and irritable bowel disease (IBD), we demonstrated that CD4CD8αα (DP8α) T lymphocytes expressed most of the regulatory markers and functions of Foxp3 Treg and secreted IL-10. Strikingly, DP8α LPL and PBL exhibited a highly skewed repertoire toward the recognition of Faecalibacterium prausnitzii, a major Clostridium species of the human gut microbiota, which is decreased in patients with IBD. Furthermore, the frequencies of DP8α PBL and colonic LPL were lower in patients with IBD than in healthy donors and in the healthy mucosa of patients with colon cancer, respectively. Moreover, PBL and LPL from most patients with active IBD failed to respond to F. prausnitzii in contrast to PBL and LPL from patients in remission and/or healthy donors. These data (i) uncover a Clostridium-specific IL-10-secreting Treg subset present in the human colonic LP and blood, (ii) identify F. prausnitzii as a major inducer of these Treg, (iii) argue that these cells contribute to the control or prevention of colitis, opening new diagnostic and therapeutic strategies for IBD, and (iv) provide new tools to address the systemic impact of both these Treg and the

  14. CD4CD8αα Lymphocytes, A Novel Human Regulatory T Cell Subset Induced by Colonic Bacteria and Deficient in Patients with Inflammatory Bowel Disease

    PubMed Central

    Sarrabayrouse, Guillaume; Bossard, Céline; Chauvin, Joe-Marc; Jarry, Anne; Meurette, Guillaume; Quévrain, Elodie; Bridonneau, Chantal; Preisser, Laurence; Asehnoune, Karim; Labarrière, Nathalie; Altare, Frédéric; Sokol, Harry; Jotereau, Francine

    2014-01-01

    How the microbiota affects health and disease is a crucial question. In mice, gut Clostridium bacteria are potent inducers of colonic interleukin (IL)-10-producing Foxp3 regulatory T cells (Treg), which play key roles in the prevention of colitis and in systemic immunity. In humans, although gut microbiota dysbiosis is associated with immune disorders, the underlying mechanism remains unknown. In contrast with mice, the contribution of Foxp3 Treg in colitis prevention has been questioned, suggesting that other compensatory regulatory cells or mechanisms may exist. Here we addressed the regulatory role of the CD4CD8 T cells whose presence had been reported in the intestinal mucosa and blood. Using colonic lamina propria lymphocytes (LPL) and peripheral blood lymphocytes (PBL) from healthy individuals, and those with colon cancer and irritable bowel disease (IBD), we demonstrated that CD4CD8αα (DP8α) T lymphocytes expressed most of the regulatory markers and functions of Foxp3 Treg and secreted IL-10. Strikingly, DP8α LPL and PBL exhibited a highly skewed repertoire toward the recognition of Faecalibacterium prausnitzii, a major Clostridium species of the human gut microbiota, which is decreased in patients with IBD. Furthermore, the frequencies of DP8α PBL and colonic LPL were lower in patients with IBD than in healthy donors and in the healthy mucosa of patients with colon cancer, respectively. Moreover, PBL and LPL from most patients with active IBD failed to respond to F. prausnitzii in contrast to PBL and LPL from patients in remission and/or healthy donors. These data (i) uncover a Clostridium-specific IL-10-secreting Treg subset present in the human colonic LP and blood, (ii) identify F. prausnitzii as a major inducer of these Treg, (iii) argue that these cells contribute to the control or prevention of colitis, opening new diagnostic and therapeutic strategies for IBD, and (iv) provide new tools to address the systemic impact of both these Treg and the

  15. Peripheral blood T-lymphocyte subsets in autoimmune thyroid disease.

    PubMed

    Covas, M I; Esquerda, A; García-Rico, A; Mahy, N

    1992-01-01

    Interest in T-lymphocyte subsets has arisen because of their involvement in the autoimmune process. Contradictory results have been published in the literature about the number of peripheral blood lymphocyte subsets in autoimmune diseases. In order to investigate the number and distribution of peripheral blood lymphocyte subsets in autoimmune thyroid disease, the levels of total T-lymphocytes (CD3), T-helper (CD4) and T-suppressor/cytotoxic (CD8) lymphocytes were determined in 44 patients with Graves' disease (1), multinodular goiter (2) and Hashimoto's thyroiditis (3). All patients had high levels of antithyroglobulin and thyroid antiperoxidase (antimicrosomal) antibodies. The T subset levels were related to the functional thyroid status, measured as serum free thyroxine (FT4) and thyrotropin (TSH). Our data show the existence of a strong influence of functional status on CD3, CD4 and CD8 levels, as reflected in the significant correlations obtained with FT4 (negative) and TSH (positive). A significant decrease in all populations was observed in Graves' disease hyperthyroid patients. A decrease in the CD4/CD8 ratio in Hashimoto's thyroiditis hypothyroid patients was observed, in contrast to an increase in the ratio in autoimmune hyperthyroid patients. This points to the CD4/CD8 ratio as a differential characteristic between the two autoimmune (hypothyroid and hyperthyroid) entities, independent of free thyroxine levels. No significant correlation was found between antithyroid antibody levels and peripheral blood T-lymphocyte subsets or serum levels of FT4 and TSH. PMID:1342892

  16. Effects of a Simulated Tennis Match on Lymphocyte Subset Measurements

    ERIC Educational Resources Information Center

    Schafer, Mark; Kell, Holly; Navalta, James; Tibana, Ramires; Lyons, Scott; Arnett, Scott

    2014-01-01

    Tennis is an activity requiring both endurance and anaerobic components, which could have immunosuppressive effects postexercise. Purpose: The purpose of this investigation was to determine the effect of a simulated tennis match on apoptotic and migratory markers on lymphocyte subsets. Method: Male high school (n = 5) and college (n = 3) tennis…

  17. Direct immunomagnetic quantification of lymphocyte subsets in blood.

    PubMed Central

    Brinchmann, J E; Vartdal, F; Gaudernack, G; Markussen, G; Funderud, S; Ugelstad, J; Thorsby, E

    1988-01-01

    A method is described where superparamagnetic polymer microspheres coated with monoclonal antibodies (MoAb) are used for the direct and fast quantification of the absolute number of cells of various lymphocyte subsets in blood. Blood samples were incubated with microspheres coated with a subset specific MoAb. Using a magnet the microsphere-rosetted cells were isolated and washed. Following lysis of the cell walls to detach the microspheres, the cell nuclei were stained with acridine orange and counted in a haemocytometer using an immunofluorescence microscope. With MoAb specific for CD2, CD4, CD8 and CD19, reproducible absolute counts of the corresponding lymphocyte subsets were obtained which correlated closely with those obtained by an indirect quantification method. PMID:3349645

  18. Chemoattractant lymphokines specific for the helper/inducer T-lymphocyte subset.

    PubMed

    Berman, J S; Cruikshank, W W; Center, D M; Theodore, A C; Beer, D J

    1985-10-01

    The cellular content of T-lymphocyte-rich inflammatory sites is dependent in part on the in situ elaboration of chemoattractant factors. We have previously described three T-lymphocyte-specific chemoattractant lymphokines; a chemokinetic factor, lymphocyte chemoattractant factor (LCF, MW 56,000), and two distinct lymphocyte migration inhibitory factors (LyMIF75K, MW 75,000; and LyMIF35K, MW 35,000). These factors are produced by human T cells in response to antigen, concanavalin A, or histamine stimulation. In this communication, we report that LCF and LyMIF35K are produced by OKT8+ (suppressor/cytotoxic) and OKT4+ (helper/inducer) lymphocytes, respectively, and are selectively chemoattractant for the OKT4+ lymphocyte subset. LyMIF75K is produced by OKT4+ cells and inhibits both OKT4+ and OKT8+ lymphocyte migration. Production of LCF and LyMIF35K by infiltrating lymphocyte subsets may be one mechanism whereby unactivated helper/inducer T lymphocytes are selectively recruited to sites of inflammation. PMID:3161625

  19. Preparation of Internal Quality Control Material for Lymphocyte Subset Analysis.

    PubMed

    Roh, Eun Youn; Shin, Sue; Yoon, Jong Hyun; Oh, Sohee; Park, Kyoung Un; Lee, Nuri; Song, Eun Young

    2016-07-01

    Lymphocyte subset analysis is widely used in clinical laboratories, and more than two levels of daily QC materials are required for reliable results. Commercially available, expensive QC materials have short shelf lives and may not be suitable in resource-poor settings. We compared different methods for preparing homemade QC material, including fixation with 1%, 2%, or 4% paraformaldehyde (PFA); freezing with 10% dimethylsulfoxide (DMSO), 0.1% bovine serum albumin-phosphate buffered saline, or after ethanolic dehydration; and using cryopreservation temperatures of -20°C, -80°C, or -196°C. We found an optimal experimental condition, which is 'fixation with 4% PFA, freezing with 10% DMSO, and storage at 80°C'. To evaluate long-term stability of QC materials prepared in this optimal condition, two levels of QC materials (QM1 and QM2) were thawed after 30, 33, 35, 37, 60, 62, 64, and 67 days of cryopreservation. Lymphocyte subset was analyzed with BD Multitest IMK kit (BD Biosciences, USA). QM1 and QM2 were stable after 1-2 months of cryopreservation (CV <3% for CD3, CD4, and CD8 and 5-7% for CD16/56 and CD19). We propose this method as an alternative cost-effective protocol for preparing homemade internal QC materials for lymphocyte subset analysis in resource-poor settings. PMID:27139609

  20. Lymphocyte subset ratios and factor VIII usage in haemophilia.

    PubMed Central

    Beddall, A C; Al-Rubei, K; Williams, M D; Hill, F G

    1985-01-01

    Type and quantity of replacement treatment, together with haematological and immunological parameters were determined in 37 boys with severe haemophilia A and 41 children with other bleeding disorders. The quantity of factor VIII concentrate given to boys with severe haemophilia A (mean U/year) showed a significant inverse correlation with total white cell counts, lymphocyte counts, platelet counts, and the ratio of monoclonal antibody defined T lymphocyte subsets, T4 and T8 (T4:T8). Of the boys with severe haemophilia A, 49% had inversed T4:T8 ratios and 24% had thrombocytopenia. Treatment with high dose factor VIII concentrate (more than 25 000 U/year) was associated with low platelet counts, low lymphocyte counts, low T4:T8 ratios, and hypergammaglobulinaemia. In addition, six patients with severe haemophilia A and factor VIII inhibitors had inversed T4:T8 ratios. Patients treated exclusively with cryoprecipitate or prothrombin complex concentrates had normal T4:T8 ratios and platelet counts. The severity of the haematological and immunological abnormalities observed seems to be associated with high usage of factor VIII concentrates. Similar abnormalities have been described in patients with the acquired immune deficiency syndrome (AIDS). Prospective study of haemophiliacs is required to assess long term sequelae of factor concentrate usage, including the possible development of AIDS. PMID:3925894

  1. Cytokine profile and lymphocyte subsets in type 2 diabetes

    PubMed Central

    Francisco, C.O.; Catai, A.M.; Moura-Tonello, S.C.G.; Arruda, L.C.M.; Lopes, S.L.B.; Benze, B.G.; Del Vale, A.M.; Malmegrim, K.C.R.; Leal, A.M.O.

    2016-01-01

    Type 2 diabetes mellitus (T2D) is a metabolic disease with inflammation as an important pathogenic background. However, the pattern of immune cell subsets and the cytokine profile associated with development of T2D are unclear. The objective of this study was to evaluate different components of the immune system in T2D patients' peripheral blood by quantifying the frequency of lymphocyte subsets and intracellular pro- and anti-inflammatory cytokine production by T cells. Clinical data and blood samples were collected from 22 men (51.6±6.3 years old) with T2D and 20 nonsmoking men (49.4±7.6 years old) who were matched for age and sex as control subjects. Glycated hemoglobin, high-sensitivity C-reactive protein concentrations, and the lipid profile were measured by a commercially available automated system. Frequencies of lymphocyte subsets in peripheral blood and intracellular production of interleukin (IL)-4, IL-10, IL-17, tumor necrosis factor-α, and interferon-γ cytokines by CD3+ T cells were assessed by flow cytometry. No differences were observed in the frequency of CD19+ B cells, CD3+CD8+ and CD3+CD4+ T cells, CD16+56+ NK cells, and CD4+CD25+Foxp3+ T regulatory cells in patients with T2D compared with controls. The numbers of IL-10- and IL-17-producing CD3+ T cells were significantly higher in patients with T2D than in controls (P<0.05). The frequency of interferon-γ-producing CD3+ T cells was positively correlated with body mass index (r=0.59; P=0.01). In conclusion, this study shows increased numbers of circulating IL-10- and IL-17-producing CD3+ T cells in patients with T2D, suggesting that these cytokines are involved in the immune pathology of this disease. PMID:27007651

  2. Homeostasis of IL-15 dependent lymphocyte subsets in the liver.

    PubMed

    Cepero-Donates, Yuneivy; Rakotoarivelo, Volatiana; Mayhue, Marian; Ma, Averil; Chen, Yi-Guang; Ramanathan, Sheela

    2016-06-01

    IL-15 is a member of the gamma chain family of cytokines (γc - CD132). The IL-15 receptor (IL-15R) complex consists of 3 subunits: the ligand-binding IL-15Rα chain (CD215), the β chain (CD122; also used by IL-2), and the common γ chain. The biological activities of IL-15 are mostly mediated by the IL-15:IL-15Rα complex, produced by the same cell and 'trans-presented' to responder cells expressing the IL-15Rβγc. The peculiar and almost unique requirement for IL-15 to be trans-presented by IL-15Rα suggests that the biological effects of IL-15 signaling are tightly regulated even at the level of availability of IL-15. Tissue-specific deletion of IL-15Rα has shown macrophage-and dendritic cell-derived IL-15Rα mediate the homeostasis of different CD8(+) T cell subsets. Here we show that hepatocyte and macrophage- specific expression of IL-15Rα is required to maintain the homeostasis of NK and NKT cells in the liver. Thus, homeostasis of IL-15-dependent lymphocyte subsets is also regulated by trans-presentation of IL-15 by non-hematopoietic cells in the tissue environment. PMID:26778709

  3. [Changes in lymphocyte subsets in bronchoalveolar lavage fluid in patients with systemic sclerosis].

    PubMed

    Kopiński, P; Nalepa, P; Wojas-Pelc, A; Janowska, E; Gil, K

    2000-01-01

    The aim of this study was to estimate if alterations of lymphocyte subsets obtained by broncholaveolar lavage (BAL) were related to clinical data observed in nonsmoking patients with systemic sclerosis (SSc). Clinical examination included chest X-rays, spirometry and arterial blood gasometry. Patients were divided into group A (pulmonary changes present, n = 15) and B (without any changes, n = 7). Healthy subjects constituted the control group (n = 10). BAL lymphocytes were phenotyped using monoclonal antibodies coupling CD4, CD8 (both in coexpression with CD25), CD19 and HLA-DR human antigens and flow cytometer FACStar (Becton-Dickinson). Parallel staining was performed in peripheral blood. BAL lymphocyte typing was completed by BAL routine cytology. In SSc patients we found increased BAL total cell number, percentage of neutrophils, eosinophils and macrophage giant cells, as well as high percent of CD25+ and HLA-DR+ lymphocytes. In the group A neutrophilic alveolitis was observed in nearly half of cases: total lymphocyte number (per 1 ml of BAL fluid) and significantly reduced CD4/CD8 ratio were found. In the group B, as compared with controls, we found significantly elevated lymphocyte total cell number per 1 ml of BAL fluid (including particular subsets: CD3+, CD4+, CD8+). Also significantly high CD4+25+ lymphocyte percent was observed. Summing up, cytological and/or immunological alterations were observed in all examined SSc patients. The intensity of these alterations seems to be related to the clinical data. A decreased value of CD4/CD8 ratio may play a role in the local appearance of pulmonary changes in the course of systemic sclerosis. PMID:10765646

  4. Lymphocyte subsets, cardiovascular measures and anxiety state before and after a professional examination.

    PubMed

    Marazziti, Donatella; Ambrogi, Fabio; Abelli, Marianna; Di Nasso, Elena; Catena, Mario; Massimetti, Gabriele; Carlini, Marina; Dell'Osso, Liliana

    2007-03-01

    Controversies exist regarding the impact of psychological stress on the functioning of the immune system in humans. The aim of the present study, therefore, was to evaluate whether the condition of a pre-exam stress may or not modify resting lymphocyte subsets, as well as blood pressure and heart rate. About 22 medical residents of both sexes not suffering from any medical or psychiatric disorder were included in the study. Anxiety levels were measured by means of the Hamilton rating scale for anxiety (HRSA) and anxiety traits by means of the panic-agoraphobic spectrum self-report (PAS-SR) version and the obsessive-compulsive spectrum self-report (OBS-SR) version. The results showed that systolic blood pressure and heart rate increased significantly just before sitting an examination (t(1)) in all subjects, as compared with a calm situation (t(2)), in parallel with the increase in the HRSA total score, while no significant difference was observed in lymphocyte subsets at the two assessment times. However, men had a higher number of CD4+ cells than women at t(1) and t(2), while women showed a higher heart rate at t(1). In addition, significant correlations between CD4+ lymphocyte count and heart rate at t(1) or HRSA at t(2) were detected. These findings indicate that the acute stress determined by sitting for examination provokes changes in autonomic nervous system parameters, such as blood pressure and heart rate, as well as in the subjective feeling of anxiety, as shown by the increased HRSA total scores, which were not paralleled by modifications of lymphocyte subsets. However, individual differences, related to both sex and personality traits yet to be identified, seem to have an impact in shaping the stress response. PMID:17454970

  5. Multicentre evaluation of stable reference whole blood for enumeration of lymphocyte subsets by flow cytometry.

    PubMed

    Edwards, Cherry; Belgrave, Danielle; Janossy, George; Bradley, Nicholas J; Stebbings, Richard; Gaines-Das, Rose; Thorpe, Robin; Sawle, Alex; Arroz, Maria Jorge; Brando, Bruno; Gratama, Jan Willem; Orfao de Matos, Alberto; Papa, Stephano; Papamichail, Michael; Lenkei, Rodica; Rothe, Gregor; Barnett, David

    2005-06-22

    BACKGROUND: Clinical indications for lymphocyte subset enumeration by flow cytometry include monitoring of disease progression and timing of therapeutic intervention in infection with human immunodeficiency virus. Until recently international standardisation has not been possible due to a lack of suitable stable reference material. METHODS: This study consisted of two trials of a stabilised whole blood preparation. Eleven participants were sent two standard protocols for staining plus gating strategy and asked to report absolute counts for lymphocyte subsets. RESULTS: No significant difference was detected between the two methods when results from the two assays and all partners were pooled. Significant differences in results from the different partners were observed. However, representative mean counts were obtained for geometric means, geometric coefficient of variation, and 95% confidence interval for CD3 910 cells/mul, 9%, and 888 to 933, respectively), CD4 (495 cells/mul, 12%, and 483 to 507), and CD8 (408 cells/mul, 13%, and 393 to 422). CONCLUSION: We have introduced a stabilised blood preparation and a well-characterized biological standard. The availability of this reference material greatly simplifies the validation of new techniques for CD4(+) T-cell enumeration and the expansion of external quality assurance programmes for clinical laboratories, including those that operate in resource-restricted environments. (c) 2005 Wiley-Liss, Inc. PMID:15973699

  6. Characteristics of blood chemistry, hematology, and lymphocyte subsets in pregnant rhesus monkeys.

    PubMed

    Wu, Ming-Ling; Gong, Li; Qian, Can; Liang, Zhi-Gang; Zeng, Wen

    2015-06-01

    The present study was designed to characterize the blood chemistry, hematology, and lymphocyte subsets in pregnant rhesus monkeys and provide baseline parameters for future studies of reproductive and developmental toxicity and developmental immunotoxicity. Harem-mating was used in 96 female and 16 male rhesus monkeys. Pregnancy was confirmed on gestation day (GD)18 by ultrasound. The blood samples of rhesus monkeys were collected at various times (20 days before pregnancy and GD20, 100 and 150). The analyses of blood chemistry, hematology, and lymphocyte subsets were performed. Compared with 20 days before pregnancy, Significant decreases (P < 0.05) were observed in HCT and RBC on GD20, GD150 and in HGB on GD150, Significant increases in NEUT and decreases in LYMPH on GD20 were observed. Significant decreases in ALB from GD20 to GD150 were observed, significant decreases in TP was observed on GD100. Significant increases in mean GLU were observed on GD20 and GD150 during pregnancy. Significant decreases (P < 0.05) in CD20(+) subsets on GD100, GD150 and CD4(+)/CD8(+)ratio on GD150 were observed, The significant changes of MCV, MCHC, RDW-SD, MCV, MONO, ALT, AST, GLB, ALP, TBIL, DBIL, IBIL, GGT, CR-S, URIC, TC, TG and CK were observed during the pregnant period, but no biologic change were observed, There were no significant changes in MCH, RDW-CV, MPV, BUN, CD3(+), CD4(+) and CD8(+) during pregnancy. These data provide a database for preclinical study in rhesus monkeys. Physiological anemia, hyperglycemia, and immune suppression may occur in pregnant rhesus monkey which is similar to that found in human, and it is essential to distinguish the physiological changes from the pharmacological effects in reproductive and developmental toxicity and developmental immunotoxicity studies of pharmaceuticals. PMID:26073336

  7. T Helper Lymphocyte Subsets and Plasticity in Autoimmunity and Cancer: An Overview

    PubMed Central

    Ivanova, Ekaterina A.; Orekhov, Alexander N.

    2015-01-01

    In response to cytokine signalling and other factors, CD4-positive T lymphocytes differentiate into distinct populations that are characterized by the production of certain cytokines and are controlled by different master transcription factors. The spectrum of such populations, which was initially limited to Th1 and Th2 subsets, is currently broadened to include Th17 and Treg subsets, as well as a number of less studied subtypes, such as Tfh, Th9, and Th22. Although these subsets appear to be relatively stable, certain plasticity exists that allows for transition between the subsets and formation of hybrid transition forms. This provides the immune system flexibility needed for adequate response to pathogens but, at the same time, can play a role in the pathogenic processes in cases of deregulation. In this review, we will discuss the properties of T lymphocyte subsets and their plasticity, as well as its implications for cancer and autoimmune diseases. PMID:26583100

  8. The effect of 1 year of trilostane treatment on peripheral lymphocyte subsets in dogs with pituitary-dependent hyperadrenocorticism

    PubMed Central

    ODA, Hitomi; MORI, Akihiro; SHONO, Saori; ONOZAWA, Eri; SAKO, Toshinori

    2016-01-01

    This study investigated the changes in lymphocyte subsets during the trilostane medication of Pituitary-dependent hyperadrenocorticism (PDH) dogs. The cortisol level and lymphocyte subsets of eight dogs with PDH were monitored 0, 1, 3, 6, 9 and 12 months after the initiation of trilostane treatment. White blood cells (WBC), lymphocytes, CD3+ (T lymphocyte), CD4+ (helper T lymphocyte), CD8+ (cytotoxic T lymphocyte) and CD21+ (B lymphocyte) cells were measured. Although the post-ACTH stimulation test cortisol level was significantly lower during trilostane treatment, changes in the CD3+, CD4+, CD8+ and CD21+ counts were not observed. Meanwhile, significant decrease was observed in WBC counts during trilostane treatment. These indicate that long-term trilostane treatment has little effect on the lymphocyte subsets in PDH dogs. PMID:26782012

  9. Peripheral Blood Lymphocyte Subset Counts in Pre-menopausal Women with Iron-Deficiency Anaemia

    PubMed Central

    Reza Keramati, Mohammad; Sadeghian, Mohammad Hadi; Ayatollahi, Hossein; Mahmoudi, Mahmoud; Khajedaluea, Mohammad; Tavasolian, Houman; Borzouei, Anahita

    2011-01-01

    Background: Iron-deficiency anaemia (IDA) is a major worldwide public health problem. Children and women of reproductive age are especially vulnerable to IDA, and it has been reported that these patients are more prone to infection. This study was done to evaluate alteration of lymphocyte subgroups in IDA. Methods: In this prospective study, we investigated lymphocyte subsets in pre-menopausal women with iron-deficiency anaemia; 50 normal subjects and 50 IDA (hypochromic microcytic) cases were enrolled. Experimental and control anticoagulated blood samples were evaluated using flow cytometry to determine the absolute and relative numbers of various lymphocyte subgroups. Finally, the results of the patient and control groups were compared. Results: Mean (SD) absolute counts of lymphocytes, CD3+ cells, CD3+/CD4+ subsets (T helper) and CD3+/CD8+ subsets (T cytotoxic) in the patient group were 2.08 (0.65) x 109/L, 1.53 (0.53) x 109/L, 0.87 (0.28) x 109/L, and 0.51 (0.24) x 109/L, respectively. The results showed significant differences between case and control groups in mean absolute counts of lymphocytes (P = 0.014), T lymphocytes (P = 0.009), helper T cells (P = 0.004), and cytotoxic T cells (P = 0.043). Conclusion: This study showed that absolute counts of peripheral blood T lymphocytes as a marker of cell-mediated immunity may be decreased in pre-menopausal women with iron-deficiency anaemia, and that these patients may be more prone to infection. PMID:22135572

  10. The effect of 2-acetyl-4-tetrahydroxybutylimidazole on lymphocyte subsets in peripheral blood of the rat.

    PubMed

    Gobin, S J; Legg, R F; Paine, A J; Phillips, J A

    1989-01-01

    A constituent of Ammonia Caramel, 2-acetyl-4-tetrahydroxybutylimidazole (THI), is known to cause a reduction in the number of circulating lymphocytes when fed to rats. In the present study the effect of giving THI 1 mg/kg by gavage daily for 7 days on the numbers of lymphocytes in subsets has been monitored in peripheral blood. Both immunoglobulin light chain-bearing B-cells (MARK-1+) and CD5 marker-bearing T-cells (OX-19+) were reduced in number within 1 day of treatment. Within the pan-T-cell population, Class II MHC reactive helper T-lymphocytes (W3/25-) were more reduced than the Class I MHC reactive cytotoxic/suppressor T cells (OX-8+). The number of null cells (MARK-1-, OX-19-) was not affected; the majority of these cells appeared to be large granular lymphocytes. PMID:2575604

  11. Reference ranges of hematology and lymphocyte subsets in healthy Korean native cattle (Hanwoo) and Holstein dairy cattle.

    PubMed

    Kim, Yun-Mi; Lee, Jin-A; Jung, Bock-Gie; Kim, Tae-Hoon; Lee, Bong-Joo; Suh, Guk-Hyun

    2016-06-01

    There are no accurate reference ranges for hematology parameters and lymphocyte subsets in Korean native beef cattle (Hanwoo). This study was performed to establish reliable reference ranges of hematology and lymphocyte subsets using a large number of Hanwoo cattle (n = 350) and to compare differences between Hanwoo and Holstein dairy cattle (n = 334). Additionally, age-related changes in lymphocyte subsets were studied. Bovine leukocyte subpopulation analysis was performed using mono or dual color flow cytometry. The leukocyte subpopulations investigated in healthy cattle included: CD2(+) cells, sIgM(+) cells, MHC class II(+) cells, CD3(+) CD4(+) cells, CD3(+) CD8(+) cells, and WC1(+) cells. Although Hanwoo and Holstein cattle are the same species, results showed several differences in hematology and lymphocyte subsets between Hanwoo and Holstein cattle. This study is the first report to establish reference ranges of hematology and lymphocyte subsets in adult Hanwoo cattle. PMID:26419947

  12. Neurobehavioral, autonomic nervous function and lymphocyte subsets among aluminum electrolytic workers.

    PubMed

    He, S C; Qiao, N; Sheng, W

    2003-01-01

    The purpose of our study is to determine the alteration of neurobehavioral parameters, autonomic nervous function and lymphocyte subsets in aluminum electrolytic workers of long-term aluminum exposure. 33 men who were 35.16 +/- 2.95 (mean +/- S.D) years old occupationally exposed to aluminum for 14.91 +/- 6.31 (mean +/- S.D) years. Air Al level and urinary aluminum concentration was measured by means of graphite furnace atomic absorption spectrophotometer. Normal reference group were selected from a flour plant. Neurobehavioral core test battery (NCTB) recommended by WHO was utilized. Autonomic nervous function test battery recommended by Ewing DJ was conducted on subjects. FAC SCAN was used to measure the lymphocyte subsets of peripheral blood. The mean air aluminum level in the workshop was 6.36 mg/m3, ranged from 2.90 to 11.38 mg/m3. Urinary aluminum of the Al electrolytic workers (40.08 +/- 9.36 microgram/mg.cre) was obviously higher than that of control group (26.84 +/- 8.93 m/mg.cre). Neurobehavioral results showed that the scores of DSY, PAC and PA in Al electrolytic workers were significantly lower than those of control group, The score of POMSC, POMSF and SRT among Al exposed workers were significantly augmented in relation to those of control group. Autonomic nervous function test results showed that R-R interval variability of maximum ratio of immediately standing up in Al electrolytic workers were decreased compare with the control group, while the BP-IS, HR-V, HR-DB, R30:15 had no significant change. Peripheral blood lymphocyte subsets test showed that CD4-CD8+ T lymphocyte in Al electrolytic workers increased. This study suggests that Al exposure exerts adverse effects on neurobehavioral performance, especially movement coordination and negative mood, and parasympathetic nervous function; moreover it increase CD4-CD8+ T lymphocyte subsets. PMID:12797904

  13. Subsets of T lymphocytes in relation to T lymphocyte function in multiple sclerosis.

    PubMed Central

    Craig, J C; Hawkins, S A; Swallow, M W; Lyttle, J A; Patterson, V H; Merrett, J D; Haire, M

    1985-01-01

    T lymphocyte control of Epstein-Barr virus (EBV) infection of autologous B lymphocytes was examined in parallel to the enumeration of subpopulations of mononuclear cells in 22 multiple sclerosis (MS) patients and in 22 healthy individuals. All were seropositive for EBV. The incidence of lack of T cell control was significantly higher in patients than in controls, confirming previous published work. In the present study, we have shown in addition a significantly reduced proportion of OKT8+ cells and a significantly increased ratio of OKT4/OKT8 cells in the group of patients with lack of control. The findings point to abnormal immunoregulation in MS. PMID:3000660

  14. Flow-cytometric analysis of T-lymphocyte subsets in sinistral and dextral patients with gingivitis.

    PubMed

    Orbak, Recep; Canakçi, Varol; Erciyas, Kamile; Kaya, Hasan

    2003-01-01

    The aim of this study was to determine whether there was any change in T-lymphocyte subsets in sinistral and dextral patients with gingivitis. The study was carried out on 36 patients (16 males and 20 females) with gingivitis. The age of the patients ranged from 16 to 25 (mean age = 18.50 +/- 3.85). Patients were divided into two equal groups according to their right or left hand use. Being right- or left-handed was determined with Edinburgh Handedness Inventory (Oldfield). At the beginning of the study, gingival index (GI-Löe-Silness) and plaque index (PI-Silness-Löe) scores were recorded in order to assess the gingival tissue health in patients. At the same time, the biopsy samples were taken from the gingival pocket wall tissues at sites of gingivitis. Then, CD4+ and CD8+ lymphocyte and CD4/CD8 ratio values were determined using flow-cytometry in the biopsy samples. The two groups were compared by using Student's t-test. The normal value in peripheral blood of CD4+ lymphocyte and that of CD8+ lymphocyte are 25-29% and 19-48%, respectively. According to flow cytometry findings, in both sinistrality and dexterity with gingivitis, CD+ and CD8+ lymphocyte values were under the normal value while the CD4/CD8 rate was within normal distribution interval. CD4+ lymphocyte values observed in the sinistral patients were found to be lower than those in the dextral patients. The difference between the CD8+ lymphocyte values in left-handed patients and that obtained in right-handed patients was not found to be statistically significant while the difference between the CD4+ lymphocyte values in left-handed patients and that obtained in right-handed patients was found to be statistically significant (p < .05). In addition, the difference between the CD4/CD8 rate obtained in left-handed patients and that obtained in right-handed patients was found to be statistically very significant (p < .001). Consequently, these findings suggested CD4+ lymphocyte value and CD4/CD8 rate was

  15. Human uterine lymphocytes.

    PubMed

    King, A; Burrows, T; Verma, S; Hiby, S; Loke, Y W

    1998-01-01

    During the luteal phase and the early months of pregnancy, there is a dense mucosal infiltration of CD56+ natural killer (NK) cells. These uterine NK cells have a phenotype (CD56bright, CD16-, mCD3-) which distinguishes them from peripheral blood NK cells (CD56dim, CD16bright, mCD3-). The uterine NK cells are in close association with extravillous trophoblast (EVT) cells which infiltrate into the decidua and maternal spiral arteries. This subpopulation of trophoblast expresses two human leukocyte antigen (HLA) class I molecules, HLA-G and HLA-C. Circulating NK cells express receptors for HLA class I molecules. We have recently found evidence that similar receptors are present on decidual NK cells belonging to both the Killer Inhibitory Receptor (KIR) and CD94 families. The repertoire of NK receptors expressed varies between different women. The findings indicate that decidual NK cells do have receptors for trophoblast HLA class I molecules. Experiments are underway to determine the effects of this interaction on NK cell function. PMID:10027599

  16. Lymphocyte cAMP and ageing: significance of subset composition, plasma noradrenaline, regular physical training and long-term smoking.

    PubMed

    Knudsen, J H; Gustafsson, F; Toft, J; Christensen, N J

    1996-11-01

    1. We studied 37 healthy men at rest in the supine. position to examine the effect of ageing, smoking and physical training of beta 2-adrenoceptor function, plasma catecholamines and the proportions of various lymphocyte subsets. 2. In 14 young subjects the proportion of natural killer cells was correlated with cAMP production in lymphocytes and inversely correlated with plasma noradrenaline level. 3. In 16 elderly non-smokers plasma noradrenaline was negatively correlated with the natural killer cell subset CD3-CD16+. Lymphocyte cAMP responses did not differ between young and elderly non-smokers, whereas plasma noradrenaline increased slightly but significantly with age. Physical training did not influence either plasma noradrenaline or adrenaline at rest or cAMP in lymphocytes. 4. In seven elderly long-term smokers cAMP production and the viability of lymphocytes were reduced. Plasma noradrenaline attained its highest values in long-term smokers. 5. It is concluded that cAMP production and plasma noradrenaline are related to lymphocyte subset composition. The greater the proportion of natural killer cells and related subsets, the higher is cAMP production and the lower is plasma noradrenaline. Thus, the inverse correlation between lymphocyte cAMP and plasma noradrenaline is indirect and most likely mediated by variability in lymphocyte subset composition. In elderly subjects, reduced cAMP production was observed in long-term smokers, and this abnormality was probably due to a reduced viability of lymphocytes and especially of natural killer cells. The negative correlation between the proportion of natural killer cells and plasma noradrenaline at rest contracts with a well-known mobilizing effect of adrenaline on natural killer cells. PMID:8942401

  17. Myeloperoxidase in human peripheral blood lymphocytes: Production and subcellular localization.

    PubMed

    Okada, Sabrina Sayori; de Oliveira, Edson Mendes; de Araújo, Tomaz Henrique; Rodrigues, Maria Rita; Albuquerque, Renata Chaves; Mortara, Renato Arruda; Taniwaki, Noemi Nosomi; Nakaya, Helder Imoto; Campa, Ana; Moreno, Ana Carolina Ramos

    2016-02-01

    Myeloperoxidase (MPO) is an important enzyme in the front-line protection against microorganisms. In peripheral blood, it is accepted that MPO is only produced by myeloid-lineage cells. Thus, MPO presence is unexpected in lymphocytes. We showed recently that B1-lymphocytes from mice have MPO. Here, we showed that subsets of human peripheral B, CD4(+) and CD8(+) T lymphocytes express MPO. The content of MPO in lymphocytes was very low compared to neutrophils/monocytes with a preferential distribution in the nucleus and perinuclear region. Also, we performed a MPO mRNA expression analysis from human blood cells derived from microarray raw data publicly available, showing that MPO is modulated in infectious disease. MPO was increased in CD4(+) T lymphocytes from HIV chronic infection and in CD8(+) T lymphocytes from HCV-positive patients. Our study points out MPO as a multifunctional protein due to its subcellular localization and expression modulation in lymphocytes indicating alternative unknown functions for MPO in lymphocytes. PMID:26632272

  18. Immunologic Monitoring of T-Lymphocyte Subsets and Hla-Dr-Positive Monocytes in Kidney Transplant Recipients

    PubMed Central

    Cho, Jang-Hee; Yoon, Young-Deuk; Jang, Hye Min; Kwon, Eugene; Jung, Hee-Yeon; Choi, Ji-Young; Park, Sun-Hee; Kim, Yong-Lim; Kim, Hyung-Kee; Huh, Seung; Won, Dong-Il; Kim, Chan-Duck

    2015-01-01

    Abstract The clinical significance of circulating T-lymphocyte subsets and human leukocyte antigen (HLA)-DR-positive monocytes in the peripheral blood of kidney transplant recipients (KTRs) remains unclear. We examined the efficacy of enumerating these cells for the immunologic monitoring of KTRs. Blood samples were obtained before transplantation, 2 weeks after transplantation and at diagnosis, and 2 weeks after treating biopsy-proven acute cellular rejection and cytomegalovirus (CMV) infection. Serial flow cytometric analysis was performed using peripheral blood obtained from 123 patients to identify the frequencies of HLA-DR+, CD3+, CD4+, CD8+, and CD25+ T-lymphocytes and HLA-DR-positive monocytes. Frequencies of CD4+CD25+/CD4+ T cells, CD8+CD25+/CD8+ T cells, and HLA-DR-positive monocytes were significantly lower at 2 weeks after transplantation than before transplantation (all P < 0.001). This decrease was not correlated with clinical parameters. The frequency of CD4+CD25+/CD4+ T cells was significantly higher in KTRs with acute rejection than in KTRs at 2 weeks after transplantation (9.10% [range 4.30–25.6%] vs 5.10% [range 0.10–33.3%]; P = 0.024). However, no significant differences were observed between stable KTRs and KTRs with CMV infection. Analysis of the receiver operating characteristic curve adjusted by covariates showed that acute rejection could be predicted with 75.0% sensitivity and 68.4% specificity by setting the cutoff value of CD4+CD25+/CD4+ T cell frequency as 5.8%. Circulating T-lymphocyte and monocyte subsets showed significant and consistent changes in their frequencies after immunosuppression. Of the various immune cells examined, circulating levels of CD4+CD25+ T cells might be a useful noninvasive immunologic indicator for detecting acute rejection. PMID:26554788

  19. Influence of fingolimod on basic lymphocyte subsets frequencies in the peripheral blood of multiple sclerosis patients – preliminary study

    PubMed Central

    Rudnicka, Julia; Czerwiec, Michał; Siwicka-Gieroba, Dorota; Walankiewicz, Monika; Grafka, Agnieszka; Zgurski, Michał; Surdacka, Agata; Bartosik-Psujek, Halina; Roliński, Jacek

    2015-01-01

    Background Fingolimod is a drug administered orally to adult patients treated for relapsing remitting course of multiple sclerosis (MS). Mode of action of fingolimod is based on intense S1P1 receptor stimulation and “arresting” lymphocytes in lymphatic organs. Objective of the research was to assess changes in the frequencies of basic lymphocyte subsets in patients treated for multiple sclerosis with the use of fingolimod. Material and methods Study group comprised of 25 previously untreated adult patients with MS. Venous blood samples were collected from each patient before and one month, three months and six months after treatment initiation. Peripheral blood lymphocyte immunophenotype was assessed with a set of monoclonal antibodies bounded to appropriate fluorochromes and flow cytometer FACSC alibur. Statistical analysis of the results was conducted using Statistica 9.0 software. Results Before fingolimod administration median of lymphocyte subsets percentage in each patient was in reference range. After 1 month of treatment we noticed significant changes in frequencies of following lymphocyte subsets: NK cells – 51.22% (p = 0.016), T CD4+ cells – 11.58% (p = 0.01), T CD4+:T CD8+ cells ratio – 0.61 (p = 0.005). After 3 and 6 months of treatment there was further increase of deviation from normal state. Conclusions The use of fingolimod is associated with profound changes in lymphocyte subsets distribution, which might bear a risk of the development of cellular immune deficiency symptoms. PMID:26648781

  20. Identification of distinct lymphocyte subsets responding to subcellular fractions of Mycobacterium bovis bacille Calmette–Guérin (BCG)

    PubMed Central

    Batoni, G; Esin, S; Pardini, M; Bottai, D; Senesi, S; Wigzell, H; Campa, M

    2000-01-01

    In order to investigate the ability of Mycobacterium bovis BCG vaccination to induce immune responses toward different classes of mycobacterial antigens and the cell populations involved in such responses, proliferation of distinct human lymphocyte subsets from BCG-vaccinated donors in response to different subcellular fractions of BCG was analysed and compared with that of not sensitized subjects. Proliferation of different cell subsets was evaluated by flow cytometric determination of bromodeoxyuridine incorporated into DNA of dividing cells and simultaneous identification of cell surface markers. Although a certain degree of variability was observed among different donors, after 6 days of in vitro stimulation BCG-vaccinated subjects displayed, as a mean, a stronger blastogenic response to all the classes of antigens compared with non-sensitized ones. PPD, culture filtrates and membrane antigens induced a predominant proliferation of CD4+ T cells. In contrast, preparations enriched in cytosolic antigens elicited strong proliferation of γδ+ T cells which, as a mean, represented 55% of the proliferating cells. Although to a lesser extent, proliferation of γδ+ T cells was also elicited by preparations enriched in membrane and cell wall antigens. In response to the latter preparation proliferation of CD4+ T cells and CD16+/CD3− (natural killer (NK)) cells was observed, as well. In particular, cell wall antigens were found to induce significantly higher levels of proliferation of NK cells compared with all the other classes of antigens. PMID:10632662

  1. Different spectra of recurrent gene mutations in subsets of chronic lymphocytic leukemia harboring stereotyped B-cell receptors

    PubMed Central

    Sutton, Lesley-Ann; Young, Emma; Baliakas, Panagiotis; Hadzidimitriou, Anastasia; Moysiadis, Theodoros; Plevova, Karla; Rossi, Davide; Kminkova, Jana; Stalika, Evangelia; Pedersen, Lone Bredo; Malcikova, Jitka; Agathangelidis, Andreas; Davis, Zadie; Mansouri, Larry; Scarfò, Lydia; Boudjoghra, Myriam; Navarro, Alba; Muggen, Alice F.; Yan, Xiao-Jie; Nguyen-Khac, Florence; Larrayoz, Marta; Panagiotidis, Panagiotis; Chiorazzi, Nicholas; Niemann, Carsten Utoft; Belessi, Chrysoula; Campo, Elias; Strefford, Jonathan C.; Langerak, Anton W.; Oscier, David; Gaidano, Gianluca; Pospisilova, Sarka; Davi, Frederic; Ghia, Paolo; Stamatopoulos, Kostas; Rosenquist, Richard

    2016-01-01

    We report on markedly different frequencies of genetic lesions within subsets of chronic lymphocytic leukemia patients carrying mutated or unmutated stereotyped B-cell receptor immunoglobulins in the largest cohort (n=565) studied for this purpose. By combining data on recurrent gene mutations (BIRC3, MYD88, NOTCH1, SF3B1 and TP53) and cytogenetic aberrations, we reveal a subset-biased acquisition of gene mutations. More specifically, the frequency of NOTCH1 mutations was found to be enriched in subsets expressing unmutated immunoglobulin genes, i.e. #1, #6, #8 and #59 (22–34%), often in association with trisomy 12, and was significantly different (P<0.001) to the frequency observed in subset #2 (4%, aggressive disease, variable somatic hypermutation status) and subset #4 (1%, indolent disease, mutated immunoglobulin genes). Interestingly, subsets harboring a high frequency of NOTCH1 mutations were found to carry few (if any) SF3B1 mutations. This starkly contrasts with subsets #2 and #3 where, despite their immunogenetic differences, SF3B1 mutations occurred in 45% and 46% of cases, respectively. In addition, mutations within TP53, whilst enriched in subset #1 (16%), were rare in subsets #2 and #8 (both 2%), despite all being clinically aggressive. All subsets were negative for MYD88 mutations, whereas BIRC3 mutations were infrequent. Collectively, this striking bias and skewed distribution of mutations and cytogenetic aberrations within specific chronic lymphocytic leukemia subsets implies that the mechanisms underlying clinical aggressiveness are not uniform, but rather support the existence of distinct genetic pathways of clonal evolution governed by a particular stereotyped B-cell receptor selecting a certain molecular lesion(s). PMID:27198719

  2. Different deoxyribonucleases in human lymphocytes

    PubMed Central

    Zöllner, E.Jürgen; Helm, Wolfgang; Zahn, Rudolf K.; Beck, Jörn; Reltz, Manfred

    1974-01-01

    The distribution pattern of deoxyribonuclease activities in human lymphocytes has been examined by micro-disc-electrophoresis. Four groups of deoxyribonuclease activities, differing in their electrophoretic mobility, in the nature of their optimal substrate and in their optimal incubation conditions, are characterized. There are two alkaline DNase-activities. One corresponds to DNase I (EC 3.1.4.5), the other having pH optimum of about pH 9.0, prefers denatured DNA as substrate and is not dependent on divalent cations. The fractions with an acid pH optimum can be subdivided into two groups, which differ in their activity towards native DNA, towards denatured DNA, in their activity when succinate is present and in their pH optimum. PMID:10793736

  3. Stimulation of human tonsillar lymphocytes in vitro

    PubMed Central

    Oettgen, H. F.; Silber, R.; Miescher, P. A.; Hirschhorn, K.

    1966-01-01

    We have studied the in vitro behaviour of cultured human tonsillar lymphocytes. In comparison with peripheral blood lymphocytes these cells show a higher degree of formation of large cells and mitoses in control cultures without any additive. They behave in a manner similar to peripheral blood lymphocytes when cultured with phytohaemagglutinin (PHA), streptolysin S (SLS) and specific antigens. The only exception is a lack of response to streptolysin O (SLO). PMID:5916348

  4. Lymphocyte subset distribution and natural killer activity in growth hormone deficiency before and during short-term treatment with growth hormone releasing hormone.

    PubMed

    Kiess, W; Malozowski, S; Gelato, M; Butenand, O; Doerr, H; Crisp, B; Eisl, E; Maluish, A; Belohradsky, B H

    1988-07-01

    Natural killer (NK) cell activity was assessed in the peripheral blood of 20 patients with growth hormone (GH) deficiency due to a hypothalamic deficit of GH-releasing hormone (GHRH). All patients failed to respond to at least two provocative tests of GH secretion (GH below 7 ng/ml) but responded to a single GHRH iv bolus injection (1 microgram/kg body wt). In 14 of the 20 patients (20 determinations), lymphocyte subsets were also measured; in all patients the distribution of lymphocyte subsets was within the normal range. More importantly, NK cell activity in the 20 patients was significantly lower than in controls (P less than 0.01). To assess the in vivo effect of GH and GHRH on NK activity and lymphocyte subset distribution, immunologic tests were performed (i) before and after a single iv bolus injection of GHRH (1 microgram/kg body wt) in six patients; (ii) before and after 3 weeks of GHRH treatment (3-9 micrograms/kg body wt, one to four times daily) in five patients; and (iii) after 6 weeks of GH treatment (5 IU sc every alternate day) in one patient. Neither NK activity nor the distribution of lymphocyte subsets was altered during short-term GHRH administration. In conclusion, low NK activity is found in GH-deficient patients, and short-term administration of GH or GHRH fails to restore this immunological abnormality. This result suggests that the hypothalamus may be a regulator of NK activity in the human and that patients with hypothalamic deficiencies should be monitored for the development of discrete immunodeficiencies. PMID:3133146

  5. Sources of heterogeneity in human monocyte subsets

    PubMed Central

    Appleby, Laura J.; Nausch, Norman; Midzi, Nicholas; Mduluza, Takafira; Allen, Judith E.; Mutapi, Francisca

    2013-01-01

    Human monocytes are commonly defined and discriminated by the extent of their cell surface expression of CD14 and CD16, with associated differences in function and phenotype related to the intensity of expression of these markers. With increasing interest into the function and behaviour of monocytes, it is important to have a clear understanding of how differing strategies of analysis can affect results and how different protocols and population backgrounds can affect this highly morphogenic cell type. Using PBMCs from populations with differing ethnicities and histories of parasite exposure we have characterized monocyte phenotype based on intensity of CD14 and CD16 expression. Using the surface markers HLA-DR, CCR2 and CX3CR1, we compared monocyte phenotype between populations and further assessed changes in monocytes with freezing and thawing of PBMCs. Our results reveal that there is a progression of surface marker expression based on intensity of CD14 or CD16 expression, stressing the importance of careful gating of monocyte subtypes. Freezing and thawing of the PBMCs has no effect generally on the monocytes, although it does lead to a decrease in CD16 and CX3CR1 expression. We show that there are differences in the monocyte populations based on ethnicity and history of exposure to the common parasites Plasmodium falciparum and Schistosoma haematobium. This study highlights that blood monocytes consist of a continuous population of cells, within which the dominant phenotype may vary dependent on the background of the study population. Comparing results from monocyte studies therefore needs to be done with great care, as ethnic background of donor population, gating strategy and processing of PBMCs may all have an effect on outcome of monocyte phenotype. PMID:23557598

  6. Characterization of a novel subset of T cells from human spleen that lacks L-selectin.

    PubMed Central

    Wallace, D L; Beverley, P C

    1993-01-01

    Human L-selection (LAM-1, Leu-8, TQ1, DREG 56) is a member of the 'selection' family of adhesion molecules. Antibodies to L-selectin have been shown to block the binding of T cells to peripheral lymph node high endothelial venules (HEV). Most unstimulated peripheral blood T cells express high levels of L-selectin whilst it is only weakly expressed on the majority of T cells from secondary lymphoid organs. We show here (a) that T cells from tonsil and lymph node up-regulate L-selectin when released from their microenvironment, (b) that in contrast, spleen contains a stable L-selectin negative subset, (c) that this subset remains surface L-selection negative after stimulation even though the T cells can respond by proliferation, (d) that this subset expresses minimal levels of LAM-1 mRNA and (e) that mucosal lymphocyte antigen (MLA) positive and T-cell receptor (TcR) gamma delta positive T cells found within the L-selectin negative population are similar to subsets of T cells found amongst lamina propria (LP) and intraepithelial lymphocytes (IEL) of the gut. Images Figure 3 PMID:7684357

  7. Lymphocyte subset analysis by Boolean algebra: a phenotypic approach using a cocktail of 5 antibodies and 3 color immunofluorescence.

    PubMed

    Hunter, S D; Peters, L E; Wotherspoon, J S; Crowe, S M

    1994-03-01

    Commercial reagent kits for the evaluation of leukocyte subsets involve the staining of a panel of up to six tubes using combinations of pre-mixed fluorescein isothiocyanate (FITC) and R-phycoerythrin (PE) conjugated monoclonal antibodies. We describe a rapid method whereby total CD3+ T-cells, CD4+ T-cells (CD3+ CD4+), CD8+ T-cells (CD3+ CD8+), putative gamma delta-receptor-T-cells (CD3+ CD4- CD8-), and T-cells that are CD3+ CD4+ CD8+ as well as B-lymphocytes and NK-cells can be enumerated after staining in a single tube. Whole blood specimens are labelled with a mixture of antibodies: FITC-conjugated antibodies to CD4 and CD19, PE-conjugated antibodies to CD8 and CD16, and either peridinin chlorophyll protein (PerCP) or allophycocyanin (APC) labelling for antibodies to CD3. After recording 20,000 events the data were analysed on the Consort 32 computer system and LYSYS-II (Becton Dickinson, San Jose, CA) and all of the lymphocyte subset values were determined by Boolean algebra using a technique we refer to as Boolean gate analysis (BGA). Our study has shown that BGA is statistically equivalent to SimulSET lymphocyte subset analysis. Furthermore, the procedure reduces the number of tubes required to two with consequential saving in reagents, consumables, and time. PMID:7514523

  8. Immunohistochemical localization of T-lymphocyte subsets in the developing lymphoid tissues of the tammar wallaby (Macropus eugenii).

    PubMed

    Duncan, Louise G; Nair, Sham V; Deane, Elizabeth M

    2012-12-01

    Research into marsupial adaptive immunity during ontogeny has been hampered by the lack of antibodies that react to marsupial immunological cell populations. In this study, newly synthesised polyclonal antibodies to the T cell marker, CD8, have been developed and used to investigate the ontogeny and distribution of this T cell population in the tammar wallaby. Immunohistochemical analysis indicated that the distribution of the CD8 lymphocytes in the lymphoid tissues of tammar neonates during the first 144 days of pouch life was similar to that of the eutherian mammals. However, CD8α(+) lymphocytes were observed in the intestines of tammar neonates prior to their first appearance in the cervical thymus, an observation that has not been found in eutherians. A dual labelling immunohistochemical approach was used for the indirect demonstration of CD4 and enabled the simultaneous detection in the tammar wallaby tissues of the two major T-lymphocyte populations, CD4 and CD8 that are associated with adaptive immunity. As in eutherian mammals, CD4(+) cells were the predominant T cell lymphocyte subset observed in the spleen while in the nodal tissues, an age-related decrease in the CD4(+)/CD8(+) ratio was noted. These antibodies provide a new immunological tool to study the role of T cell subsets in marsupial immunity and disease pathogenesis studies. PMID:22929957

  9. An analysis of T lymphocyte subsets in tumour-transplanted mice on the basis of Lyt antigenic markers and functions

    PubMed Central

    Lala, P. K.; McKenzie, I. F. C.

    1982-01-01

    Small lymphocyte subsets were characterized radioautographically on the basis of several surface markers, viz. surface Ig (S-Ig), Thy-1 and Lyt (Ly-1, Ly-2 and 3) antigens in host lymphoid organs (thymus, spleen and blood) as well as at the tumour site at various stages of subcutaneous growth of two different syngeneic tumours—MPC-11 plasmacytoma and WEHI-164 fibrosarcoma in BALB/c mice. In both tumour-host combinations there was a rise in the levels of null (S-Ig-, Thy-1-) small lymphocytes as well as the Ly-23+ subset of T small lymphocytes at all the sites examined. The absolute number of these two subsets also increased excepting the case of null cell rise in the thymus which was relative. The functional potentials of Lyt subsets were explored by employing in vitro and in vivo assays. While no appreciable levels of anti-tumour cytotoxic T cells (Tc) were detectable by a 51Cr release assay in the host spleen or the tumour-draining lymph nodes at any stage of growth of MPC-11 tumour, such Tc was generated in vitro by a co-cultivation of unprimed spleen cells with irradiated MPC-11 cells. These Tc were Thy-1+ and Ly-12+, as noted from antibody+C′ mediated abrogation of cytotoxicity. These results suggested that the generation of anti-tumour Tc in vivo was suppressed in tumour-bearing hosts. The possibility of a cell-mediated suppression was tested by an adoptive transfer of thymocytes or splenocytes from tumour-bearing mice into naive or pre-immunized recipients which then received fresh tumour transplants. This procedure caused a specific enhancement of tumour growth in three tumour-host combinations: MPC-11 or WEHI-164 tumour in BALB/c mice and W-1 fibrosarcoma in CBA mice. The suppressor lineage lymphocytes appearing in vivo were found to be Thy-1+ and Ly-1-, 2+, as noted from antibody +C′ mediated abrogation of their tumour-growth promoting ability. They appeared earlier (7 days) in the thymus and later (>2 weeks) in the spleen and then persisted during

  10. Selective Recruitment of T-Cell Subsets to the Udder during Staphylococcal and Streptococcal Mastitis: Analysis of Lymphocyte Subsets and Adhesion Molecule Expression

    PubMed Central

    Soltys, Jindrich; Quinn, Mark T.

    1999-01-01

    During bacterial infection of the bovine mammary gland, large numbers of leukocytes migrate into the udder, resulting in the establishment of a host response against the pathogen. Currently, the specific leukocyte populations mediating this immune response are not well defined. In the studies described here, we analyzed blood and milk from healthy cows and cows with naturally occurring mastitis to determine if distinct αβ and γδ T-lymphocyte subsets were involved in the response of the udder to a mastitis pathogen and if the type of mastitis pathogen influenced the subset composition of these responding leukocytes. Although blood samples from cows with confirmed staphylococcal and streptococcal mastitis were characterized by increased numbers of γδ T cells, the most dramatic changes in leukocyte distributions occurred in milk samples from these cows, with a 75% increase in αβ T-cell levels and a 100% increase in γδ T-cell levels relative to the levels in milk samples from healthy animals. Interestingly, the increase in αβ T-cell numbers observed in milk from cows with staphylococcal mastitis was primarily due to increased numbers of CD4+ T cells, while the increase in αβ T-cell numbers observed in cows with streptococcal mastitis was due to a parallel increase in both CD4+ and CD8+ T-cell numbers. The increased numbers of γδ T cells in milk from cows with staphylococcal and streptococcal mastitis were due to a selective recruitment of a distinct γδ T-cell subset (GD3.1+), while no change in the numbers of GD197+ γδ T cells was observed. We also analyzed adhesion protein expression on blood and milk leukocytes and found that, in comparison to the situation for healthy cows, L-selectin was down-regulated and CD18 was up-regulated on leukocytes from cows with mastitis. Thus, shedding of L-selectin and up-regulation of CD18 by neutrophils may provide a sensitive indicator of early inflammatory responses during bovine mastitis. Overall, these studies

  11. The effect of dietary lipid manipulation on rat lymphocyte subsets and proliferation.

    PubMed Central

    Yaqoob, P; Newsholme, E A; Calder, P C

    1994-01-01

    Polyunsaturated fatty acids (PUFA) have been shown to suppress immune cell functions in vitro. Dietary studies investigating the effects of PUFA-containing oils on lymphocyte functions have yielded contradictory findings: such studies are difficult to compare as there are many variations in protocols. The present study investigated the effects of diets containing oils rich in saturated fatty acids, monounsaturated fatty acids, n-6 PUFA or n-3 PUFA on rat lymphocyte proliferation and on receptor and surface marker expression. Rats were fed for 10 weeks on a low-fat (LF) diet (approximately 2% fat by weight) or on one of five high-fat diets, which contained 20% (by weight) hydrogenated coconut oil (HCO), olive oil (OO), safflower oil (SO), evening primrose oil (EPO) or menhaden (fish) oil (MO). Compared with feeding the LF diet, all of the high-fat diets suppressed the proliferation of lymphocytes from the spleen: although there was no significant effect of diet on the proliferation of lymphocytes from the thymus, there was a trend towards decreased proliferation with high-fat feeding. Feeding the OO, EPO or MO diets significantly suppressed proliferation of mesenteric lymph node lymphocytes compared with feeding the LF, HCO or SO diets. Dietary lipid manipulation had no effect on the proportion of T cells, B cells or monocytes/macrophages in the spleen, thymus or lymph nodes. Dietary lipid manipulation also had no significant effect on the proportions of CD4+ or CD8+ lymphocytes in spleen, thymus or lymph nodes, either in freshly prepared cells or in cells cultured in the presence of mitogen. There were no significant effects of dietary lipid manipulation on the expression of IL-2 receptors or transferrin receptors by concanavalin A (Con A)-stimulated lymphocytes. However, there was a trend towards a decrease in transferrin receptor expression by Con A-stimulated lymphocytes from the thymus and lymph nodes of the MO-fed rats and towards a decrease in the expression

  12. Changes in lymphocyte and macrophage subsets due to morphine and ethanol treatment during a retrovirus infection causing murine AIDS

    SciTech Connect

    Watson, R.R.; Prabhala, R.H.; Darban, H.R.; Yahya, M.D.; Smith, T.L.

    1988-01-01

    The number of lymphocytes of various subsets were not significantly changed by the ethanol exposure except those showing activation markers which were reduced. The percentage of peripheral blood cells showing markers for macrophage functions and their activation were significantly reduced after binge use of ethanol. Ethanol retarded suppression of cells by retroviral infection. However by 25 weeks of infection there was a 8.6% survival in the ethanol fed mice infected with retrovirus which was much less than virally infected controls. Morphine treatment also increased the percentage of cells with markers for macrophages and activated macrophages in virally infected mice, while suppressing them in uninfected mice. The second and third morphine injection series suppressed lymphocyte T-helper and T-suppressor cells, but not total T cells. However, suppression by morphine was significantly less during retroviral disease than suppression caused by the virus only. At 25 weeks of infection 44.8% of morphine treated, infected mice survived.

  13. [The effect of various occupational exposures to microwave radiation on the concentrations of immunoglobulins and T lymphocyte subsets].

    PubMed

    Moszczyński, P; Lisiewicz, J; Dmoch, A; Zabiński, Z; Bergier, L; Rucińska, M; Sasiadek, U

    1999-01-01

    The immunoglobulins' concentrations and T lymphocyte subsets during occupational exposures to microwave radiation were assessed. In the workers of retransmission TV center and center of satellite communications on increased IgG and IgA concentration and decreased count of lymphocytes and T8 cells was found. However, in the radar operators IgM concentration was elevated and a decrease in the total T8 cell count was observed. The different behaviour of examined immunological parameters indicate that the effect of microwave radiation on immune system depends on character of an exposure. Disorders in the immunoglobulins' concentrations and in the T8 cell count did not cause any clinical consequences. PMID:10335122

  14. Plasticity of human CD4 T cell subsets.

    PubMed

    Geginat, Jens; Paroni, Moira; Maglie, Stefano; Alfen, Johanna Sophie; Kastirr, Ilko; Gruarin, Paola; De Simone, Marco; Pagani, Massimiliano; Abrignani, Sergio

    2014-01-01

    Human beings are exposed to a variety of different pathogens, which induce tailored immune responses and consequently generate highly diverse populations of pathogen-specific T cells. CD4(+) T cells have a central role in adaptive immunity, since they provide essential help for both cytotoxic T cell- and antibody-mediated responses. In addition, CD4(+) regulatory T cells are required to maintain self-tolerance and to inhibit immune responses that could damage the host. Initially, two subsets of CD4(+) helper T cells were identified that secrete characteristic effector cytokines and mediate responses against different types of pathogens, i.e., IFN-γ secreting Th1 cells that fight intracellular pathogens, and IL-4 producing Th2 cells that target extracellular parasites. It is now well established that this dichotomy is insufficient to describe the complexity of CD4(+) T cell differentiation, and in particular the human CD4 compartment contains a myriad of T cell subsets with characteristic capacities to produce cytokines and to home to involved tissues. Moreover, it has become increasingly clear that these T cell subsets are not all terminally differentiated cells, but that the majority is plastic and that in particular central memory T cells can acquire different properties and functions in secondary immune responses. In addition, there is compelling evidence that helper T cells can acquire regulatory functions upon chronic stimulation in inflamed tissues. The plasticity of antigen-experienced human T cell subsets is highly relevant for translational medicine, since it opens new perspectives for immune-modulatory therapies for chronic infections, autoimmune diseases, and cancer. PMID:25566245

  15. Plasticity of Human CD4 T Cell Subsets

    PubMed Central

    Geginat, Jens; Paroni, Moira; Maglie, Stefano; Alfen, Johanna Sophie; Kastirr, Ilko; Gruarin, Paola; De Simone, Marco; Pagani, Massimiliano; Abrignani, Sergio

    2014-01-01

    Human beings are exposed to a variety of different pathogens, which induce tailored immune responses and consequently generate highly diverse populations of pathogen-specific T cells. CD4+ T cells have a central role in adaptive immunity, since they provide essential help for both cytotoxic T cell- and antibody-mediated responses. In addition, CD4+ regulatory T cells are required to maintain self-tolerance and to inhibit immune responses that could damage the host. Initially, two subsets of CD4+ helper T cells were identified that secrete characteristic effector cytokines and mediate responses against different types of pathogens, i.e., IFN-γ secreting Th1 cells that fight intracellular pathogens, and IL-4 producing Th2 cells that target extracellular parasites. It is now well established that this dichotomy is insufficient to describe the complexity of CD4+ T cell differentiation, and in particular the human CD4 compartment contains a myriad of T cell subsets with characteristic capacities to produce cytokines and to home to involved tissues. Moreover, it has become increasingly clear that these T cell subsets are not all terminally differentiated cells, but that the majority is plastic and that in particular central memory T cells can acquire different properties and functions in secondary immune responses. In addition, there is compelling evidence that helper T cells can acquire regulatory functions upon chronic stimulation in inflamed tissues. The plasticity of antigen-experienced human T cell subsets is highly relevant for translational medicine, since it opens new perspectives for immune-modulatory therapies for chronic infections, autoimmune diseases, and cancer. PMID:25566245

  16. Virus-lymphocyte interaction: T cells of the helper subset are infected with lymphocytic choriomeningitis virus during persistent infection in vivo.

    PubMed Central

    Ahmed, R; King, C C; Oldstone, M B

    1987-01-01

    The lifelong persistence of lymphocytic choriomeningitis virus (LCMV) in neonatally or congenitally infected mice is accompanied by a suppression of virus-specific T-cell responses. In this study, we identified the subset of T cells infected with LCMV during persistent infection in vivo. Using specific monoclonal antibodies to separate the different lymphocyte cell populations and employing both an infectious center assay and immunofluorescence to detect the virus, we found that infection is confined primarily to T cells of the helper subset (L3T4+ Lyt2-), with minimal involvement of cytotoxic T cells (Lyt2+ L3T4-) and mature B cells. About 0.54 to 1.1% of L3T4+ T cells were producing the virus, as determined by the infectious center assay. In contrast, 9.1 to 12.2% of these L3T4+ T cells contained viral antigen, as shown by immunofluorescence studies. This finding suggested that, at any given time, a substantial number of infected T cells were not producing infectious virus. This infection of T helper cells may be involved in the suppression of LCMV-specific T-cell responses observed in persistently infected mice. Images PMID:2952807

  17. The Difference of Lymphocyte Subsets Including Regulatory T-Cells in Umbilical Cord Blood between AGA Neonates and SGA Neonates

    PubMed Central

    Yoon, Sang Hee; Hur, Mina; Hwang, Han Sung; Kwon, Han Sung

    2015-01-01

    Purpose This study aimed to compare the regulatory T cells in cord blood of appropriate for gestational age (AGA) neonates with those of small for gestational age (SGA) neonates. Materials and Methods Umbilical cord blood was collected upon labor in 108 healthy full-term (between 37 and 41 gestational weeks) neonates, who were born between November 2010 and April 2012. Among them, 77 samples were obtained from AGA neonates, and 31 samples were obtained from SGA neonates. Regulatory T cells and lymphocyte subsets were determined using a flow cytometer. Student's t-test for independent samples was used to compare differences between AGA and SGA neonates. Results Regulatory T cells in cord blood were increased in the SGA group compared with normal controls (p=0.041). However, cytotoxic T cells in cord blood were significantly decreased in the SGA group compared with normal controls (p=0.007). Conclusion This is the first study to compare the distribution of lymphocyte subsets including regulatory T cells in cord blood between AGA neonates and SGA neonates. PMID:25837188

  18. Effects of Anti-CD45RB Monoclonal Antibody for T Lymphocyte Subsets in Mice Heart Transplantation Model.

    PubMed

    Deng, C-Y; Wang, X-F; Qi, H; Li, F-R

    2016-08-01

    Anti-CD45RB monoclonal antibody (anti-CD45RBmAb), as a new immune tolerance inducer, may inhibit T cell proliferation and induce immune tolerance through competitive combination with CD45RB on the T cell surface, which blocks the conduction of activation signals. However, how anti-CD45RBmAb plays its role on T lymphocyte subsets during immunosuppression remains unclear. In this work, we investigate the effects of anti-CD45RBmAb on CD3(+) T lymphocyte both in vitro and in allogeneic heart transplant model in vivo. Interestingly, anti-CD45RBmAb could inhibit the proliferation of T cells, promote the transformation of T lymphocyte to Treg and Th2 cells, suppress the transformation to Th17 and Th1 cells, increase the number of Ts cells, decrease the number of Tm cells and thus play a role in immune inhibition and induction of immune tolerance. PMID:27146476

  19. A reassessment of IgM memory subsets in humans

    PubMed Central

    Bagnara, Davide; Squillario, Margherita; Kipling, David; Mora, Thierry; Walczak, Aleksandra M.; Da Silva, Lucie; Weller, Sandra; Dunn-Walters, Deborah K.; Weill, Jean-Claude; Reynaud, Claude-Agnès

    2015-01-01

    From paired blood and spleen samples from three adult donors we performed high-throughput V-h sequencing of human B-cell subsets defined by IgD and CD27 expression: IgD+CD27+ (“MZ”), IgD−CD27+(“memory”, including IgM (“IgM-only”), IgG and IgA) and IgD−CD27− cells (“double-negative”, including IgM, IgG and IgA). 91,294 unique sequences clustered in 42,670 clones, revealing major clonal expansions in each of these subsets. Among these clones, we further analyzed those shared sequences from different subsets or tissues for Vh-gene mutation, H-CDR3-length, and Vh/Jh usage, comparing these different characteristics with all sequences from their subset of origin, for which these parameters constitute a distinct signature. The IgM-only repertoire profile differed notably from that of MZ B cells by a higher mutation frequency, and lower Vh4 and higher Jh6 gene usage. Strikingly, IgM sequences from clones shared between the MZ and the memory IgG/IgA compartments showed a mutation and repertoire profile of IgM-only and not of MZ B cells. Similarly, all IgM clonal relationships (between MZ, IgM-only, and double-negative compartments) involved sequences with the characteristics of IgM-only B cells. Finally, clonal relationships between tissues suggested distinct recirculation characteristics between MZ and switched B cells. The “IgM-only” subset (including cells with its repertoire signature but higher IgD or lower CD27 expression levels) thus appear as the only subset showing precursor-product relationships with CD27+ switched memory B cells, indicating that they represent germinal center-derived IgM memory B cells, and that IgM memory and MZ B cells constitute two distinct entities. PMID:26355154

  20. A Reassessment of IgM Memory Subsets in Humans.

    PubMed

    Bagnara, Davide; Squillario, Margherita; Kipling, David; Mora, Thierry; Walczak, Aleksandra M; Da Silva, Lucie; Weller, Sandra; Dunn-Walters, Deborah K; Weill, Jean-Claude; Reynaud, Claude-Agnès

    2015-10-15

    From paired blood and spleen samples from three adult donors, we performed high-throughput VH sequencing of human B cell subsets defined by IgD and CD27 expression: IgD(+)CD27(+) ("marginal zone [MZ]"), IgD(-)CD27(+) ("memory," including IgM ["IgM-only"], IgG and IgA) and IgD(-)CD27(-) cells ("double-negative," including IgM, IgG, and IgA). A total of 91,294 unique sequences clustered in 42,670 clones, revealing major clonal expansions in each of these subsets. Among these clones, we further analyzed those shared sequences from different subsets or tissues for VH gene mutation, H-CDR3-length, and VH/JH usage, comparing these different characteristics with all sequences from their subset of origin for which these parameters constitute a distinct signature. The IgM-only repertoire profile differed notably from that of MZ B cells by a higher mutation frequency and lower VH4 and higher JH6 gene usage. Strikingly, IgM sequences from clones shared between the MZ and the memory IgG/IgA compartments showed a mutation and repertoire profile of IgM-only and not of MZ B cells. Similarly, all IgM clonal relationships (among MZ, IgM-only, and double-negative compartments) involved sequences with the characteristics of IgM-only B cells. Finally, clonal relationships between tissues suggested distinct recirculation characteristics between MZ and switched B cells. The "IgM-only" subset (including cells with its repertoire signature but higher IgD or lower CD27 expression levels) thus appear as the only subset showing precursor-product relationships with CD27(+) switched memory B cells, indicating that they represent germinal center-derived IgM memory B cells and that IgM memory and MZ B cells constitute two distinct entities. PMID:26355154

  1. Morphological heterogeneity of Leu7, Leu11 and OKM1 positive lymphocyte subsets: an ultrastructural study with the immunogold method.

    PubMed Central

    Polli, N; Matutes, E; Robinson, D; Catovsky, D

    1987-01-01

    The morphological features of normal peripheral blood lymphocytes reactive with three monoclonal antibodies (MoAb) against natural killer (NK) cells, Leu7, OKM1 (CD11b) and Leu11 (CD16) and with two anti-T cell MoAb, CD4 and CD8, have been analysed at ultrastructural level by an indirect immunogold method. Cells having the features of large granular lymphocytes (LGL) but also lymphocytes displaying different morphological characteristics (non LGL; e.g. high nucleo-cytoplasmic ratio and few cytoplasmic organelles) were seen reactive with each of the MoAb investigated. Leu7 identified a higher proportion of LGL (60-80%) than OKM1 (10-95%) and Leu11 (20-48%), and with a stronger binding. A distinct granular structure, recognized as parallel tubular arrays, was more characteristic of the Leu7+, CD8+ LGL and was less frequently seen in the OKM1 and Leu11 positive LGL subpopulation in four out of the five donors investigated. It is of interest that the Leu11 and OKM1 positive subsets, which correspond functionally to cells with greater NK function, had relatively less LGL than the Leu7 positive subsets, raising the issue of the true morphology of NK cells in man. The existence of a minority of CD4 positive LGL was confirmed. Our findings demonstrate that there is a degree of morphological heterogeneity within the normal NK lymphoid population as defined by the membrane phenotype and that certain variability among normal individuals regarding the proportion and structural features of the NK subpopulations may be present. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 Fig. 5 Fig. 6 PMID:3498572

  2. Immunity to Pathogens Taught by Specialized Human Dendritic Cell Subsets

    PubMed Central

    Geginat, Jens; Nizzoli, Giulia; Paroni, Moira; Maglie, Stefano; Larghi, Paola; Pascolo, Steve; Abrignani, Sergio

    2015-01-01

    Dendritic cells (DCs) are specialized antigen-presenting cells (APCs) that have a key role in immune responses because they bridge the innate and adaptive arms of the immune system. They mature upon recognition of pathogens and upregulate MHC molecules and costimulatory receptors to activate antigen-specific CD4+ and CD8+ T cells. It is now well established that DCs are not a homogeneous population but are composed of different subsets with specialized functions in immune responses to specific pathogens. Upon viral infections, plasmacytoid DCs (pDCs) rapidly produce large amounts of IFN-α, which has potent antiviral functions and activates several other immune cells. However, pDCs are not particularly potent APCs and induce the tolerogenic cytokine IL-10 in CD4+ T cells. In contrast, myeloid DCs (mDCs) are very potent APCs and possess the unique capacity to prime naive T cells and consequently to initiate a primary adaptive immune response. Different subsets of mDCs with specialized functions have been identified. In mice, CD8α+ mDCs capture antigenic material from necrotic cells, secrete high levels of IL-12, and prime Th1 and cytotoxic T-cell responses to control intracellular pathogens. Conversely, CD8α− mDCs preferentially prime CD4+ T cells and promote Th2 or Th17 differentiation. BDCA-3+ mDC2 are the human homologue of CD8α+ mDCs, since they share the expression of several key molecules, the capacity to cross-present antigens to CD8+ T-cells and to produce IFN-λ. However, although several features of the DC network are conserved between humans and mice, the expression of several toll-like receptors as well as the production of cytokines that regulate T-cell differentiation are different. Intriguingly, recent data suggest specific roles for human DC subsets in immune responses against individual pathogens. The biology of human DC subsets holds the promise to be exploitable in translational medicine, in particular for the development of vaccines against

  3. Immunity to Pathogens Taught by Specialized Human Dendritic Cell Subsets.

    PubMed

    Geginat, Jens; Nizzoli, Giulia; Paroni, Moira; Maglie, Stefano; Larghi, Paola; Pascolo, Steve; Abrignani, Sergio

    2015-01-01

    Dendritic cells (DCs) are specialized antigen-presenting cells (APCs) that have a key role in immune responses because they bridge the innate and adaptive arms of the immune system. They mature upon recognition of pathogens and upregulate MHC molecules and costimulatory receptors to activate antigen-specific CD4(+) and CD8(+) T cells. It is now well established that DCs are not a homogeneous population but are composed of different subsets with specialized functions in immune responses to specific pathogens. Upon viral infections, plasmacytoid DCs (pDCs) rapidly produce large amounts of IFN-α, which has potent antiviral functions and activates several other immune cells. However, pDCs are not particularly potent APCs and induce the tolerogenic cytokine IL-10 in CD4(+) T cells. In contrast, myeloid DCs (mDCs) are very potent APCs and possess the unique capacity to prime naive T cells and consequently to initiate a primary adaptive immune response. Different subsets of mDCs with specialized functions have been identified. In mice, CD8α(+) mDCs capture antigenic material from necrotic cells, secrete high levels of IL-12, and prime Th1 and cytotoxic T-cell responses to control intracellular pathogens. Conversely, CD8α(-) mDCs preferentially prime CD4(+) T cells and promote Th2 or Th17 differentiation. BDCA-3(+) mDC2 are the human homologue of CD8α(+) mDCs, since they share the expression of several key molecules, the capacity to cross-present antigens to CD8(+) T-cells and to produce IFN-λ. However, although several features of the DC network are conserved between humans and mice, the expression of several toll-like receptors as well as the production of cytokines that regulate T-cell differentiation are different. Intriguingly, recent data suggest specific roles for human DC subsets in immune responses against individual pathogens. The biology of human DC subsets holds the promise to be exploitable in translational medicine, in particular for the development of

  4. Peripheral T lymphocyte subset imbalances in children with enterovirus 71-induced hand, foot and mouth disease.

    PubMed

    Li, Shuxian; Cai, Chunyan; Feng, Jinyan; Li, Xuejing; Wang, Yingshuo; Yang, Jun; Chen, Zhimin

    2014-02-13

    Inflammatory mediators (i.e. cytokines) play a pivotal role in the regulation of pathophysiological processes during EV71-induced hand, foot and mouth disease (HFMD). Different T cell subsets have distinct cytokine secretion profiles, and alteration in the T cell subsets frequency (imbalance) during infection leads to changed cytokine patterns. However, the effects of EV71 infection on T cell subsets were not clear. The objective of this study was to determine whether EV71-induced HFMD can be explained by the emergence of particular T-cell subsets (Th1, Th2, Tc1, Tc2, Th17, Tc17 and Treg cells) and the cytokine they produced (IFN-γ, IL-4, IL-17A and TGF-β1), as well as distinct responses to EV71 infection. We found that when compared to the control group, the percentage of Th1 and Tc1 cells was significantly higher in mild and severe HFMD group. Similar results were found in the Th1/Th2 ratio and IFN-γ levels. On the other hand, the percentage of Th17 cells and IL-17A levels were the highest in severe HFMD cases, and lowest in controls. Similar trend was also found for the Th17/Treg cell ratio. An optimal cutoff value of 2.15% for Th17 cell and 6.72 pg/ml for IL-17A provided a discriminatory value for differentiating the severity of HFMD cases by receiver operating characteristic curve analyses. These findings reveal that the Th1/Th2 and Th17/Treg imbalance exist in HFMD patients, suggesting their involvement in the pathogenesis of EV71 infection, which may have potential value as biomarkers. PMID:24316007

  5. Immunoregulatory CD4+ CD45R+ suppressor/inducer T lymphocyte subsets and impaired cell-mediated immunity in patients with Down's syndrome.

    PubMed Central

    Raziuddin, S; Elawad, M E

    1990-01-01

    The monoclonal antibodies 2H4 and 4B4 allow CD4+ and CD8+ T lymphocytes to be subdivided into CD45R+ and CDW29+ functional subpopulations. The CD4+ CD45R+ lymphocytes are designated as suppressor/inducer and CD4+ CDW29+ as helper/inducer subsets. Peripheral blood lymphocytes from 19 patients with Down's syndrome and 19 age- and sex-matched normal controls were analysed for the CD45R+ and CDW29+ subsets from the CD4+ and CD8+ T lymphocytes. The percentage of CD4+ CD45R+ cells (suppressor inducer) was markedly increased and of CD4+ CDW29+ cells (helper/inducer) decreased in all patients with Down's syndrome. In contract, the percentage of CD8+ CD45R+ and CD8+ CDW29+ subsets showed no major differences between patients with Down's syndrome and normal controls. Moreover, an alteration in the CD4+ and CD45R+ and CD4+ CDW29+ T cell subsets was accompanied by a markedly reduced proliferative response to phytohaemagglutinin and concanavalin A stimulation of the CD4+ T lymphocytes. Thus, a deficiency exists in patients with Down's syndrome in the CD4+ CDW29+ helper/inducer T cell subset which may contribute to their impaired cell-mediated immunity. PMID:1967994

  6. Reconstitution of maturating and regulatory lymphocyte subsets after cord blood and BMT in children.

    PubMed

    Charrier, E; Cordeiro, P; Brito, R-M; Mezziani, S; Herblot, S; Le Deist, F; Duval, M

    2013-03-01

    Some clinical characteristics of cord blood transplantation (CBT) might be explained by specificities in the reconstitution of immune subsets differing by their maturation stage or their implication in GVHD, tolerance or immune responses against tumor or infectious agents. Here, we compare the immune reconstitution of several of these subsets after CBT and BMT. B-cell count recovery was faster after CBT. There was no difference in the recovery of CD4(+) and CD8(+) cell counts. There was no difference either in the frequency of several subsets: CD45RO(+) memory, and CD45RA(+) naïve cells within the CD4(+) T-cell compartment, CD27(+) among B cells, CD56(bright), NKG2A(+), and KIR(+) cells among natural killer (NK) cells, CD25(+)FOXP3(+) regulatory T cells and invariant NKT cells. The proportion of the thymic naïve CD31(+)CD45RA(+)CD4(+) T cells was lower after CBT at 6 months post-transplant, and was still below normal at 1 year in both groups. NK-cell expansion was more sustained after CBT, with fewer double-negative NKG2A(-)KIR(-) hyporesponsive cells and more double-positive NKG2A(+)KIR(+) hyper-responsive NK cells. These results, therefore, indicate that further research to improve CBT outcome should try to improve thymopoieisis and take advantage of the sustained NK-cell reconstitution. PMID:23064038

  7. Methionine dependency of cultured human lymphocytes.

    PubMed

    Hall, C A; Begley, J A; Chu, R C

    1986-06-01

    Human peripheral blood lymphocytes stimulated with phytohemagglutinin and a lymphocyte model consisting of the RPMI 6410 cell, a human virus-transformed B cell, required added methionine (Met) for growth of the cultures. This failure to meet all needs for Met via endogenous synthesis, which is characteristic of oncogenic transformation, occurred even in the presence of adequate homocysteine, methylfolate (5-CH3-H4PteGlu) and cobalamin (Cbl)-dependent methionine synthetase activity. Folinic acid (5-CHO-H4PteGlu), which provides available folate independently of Cbl, improved growth only slightly in the absence of Met. Free Cbl at 222 nM, an amount great enough to alter other intracellular events, failed to increase growth in the absence of Met, but 0.22 nM Cbl bound to transcobalamin II did, however, enhance growth. PMID:3703873

  8. Immunoregulation in onchocerciasis. Functional and phenotypic abnormalities of lymphocyte subsets and changes with therapy.

    PubMed Central

    Freedman, D O; Lujan-Trangay, A; Steel, C; Gonzalez-Peralta, C; Nutman, T B

    1991-01-01

    To help define the immunoregulatory defects in patients with onchocerciasis, flow cytometric analysis of circulating lymphocyte subpopulations was performed in parallel with functional assays. No significant differences in CD4/CD8 ratios were seen when microfilariae-positive individuals from Guatemala were compared with Guatemalan controls. However, the infected individuals had significantly increased numbers of circulating CD4+CD45RA+ lymphocytes (mean 38.3%) when compared with controls (mean 16.0%). Coexpression of the activation marker HLA-DR was significantly increased on CD4+ cells from infected individuals. In contrast, no up-regulation of HLA-DR was seen on CD8+ or CD19+ cells. At 1 year after initiation of treatment with semiannual doses of the microfilaricide ivermectin, there were significant increases (P less than 0.05) in the percentage of CD4+CD45RA- cells, the percentage of CD4+HLA-DR+ cells, and mitogen-induced lymphokine production (IL-2, IL-4). Despite these changes, parasite-specific IL-2 and IL-4 production which had been undetectable before treatment did not manifest itself even by the 2-yr follow-up. Defects in the T-cell activation pathway in Onchocerca volvulus-infected individuals may thus exist at several independent points; a state of parasite antigen-specific tolerance appears to remain even after the relative reversal of other generalized immunoregulatory defects. PMID:1829096

  9. Aging of immune system: Immune signature from peripheral blood lymphocyte subsets in 1068 healthy adults

    PubMed Central

    Qin, Ling; Jing, Xie; Qiu, Zhifeng; Cao, Wei; Jiao, Yang; Routy, Jean-Pierre; Li, Taisheng

    2016-01-01

    Aging is a major risk factor for several conditions including neurodegenerative, cardiovascular diseases and cancer. Functional impairments in cellular pathways controlling genomic stability, and immune control have been identified. Biomarker of immune senescence is needed to improve vaccine response and to develop therapy to improve immune control. To identify phenotypic signature of circulating immune cells with aging, we enrolled 1068 Chinese healthy volunteers ranging from 18 to 80 years old. The decreased naïve CD4+ and CD8+ T cells, increased memory CD4+ or CD8+ T cells, loss of CD28 expression on T cells and reverse trend of CD38 and HLA-DR, were significant for aging of immune system. Conversely, the absolute counts and percentage of NK cells and CD19+B cells maintained stable in aging individuals. The Chinese reference ranges of absolute counts and percentage of peripheral lymphocyte in this study might be useful for future clinical evaluation. PMID:26886066

  10. Serum immunoglobulins and lymphocyte subset distributions in children and adults living in communities assessed for lead and cadmium exposure

    SciTech Connect

    Sarasua, S.M.; Vogt, R.F.; Henderson, L.O.; Jones, P.A.; Lybarger, J.A.

    2000-05-12

    This study assessed the impact of environmental cadmium and lead exposure on the immune system of more than 2,000 children and adults. Serum immunoglobulins [immunoglobulins (lg) A, G, and M] and peripheral blood lymphocyte phenotypes (T cells, B cells, NK cells, and CD4/CD8 subsets) were measured in a total of 2041 children and adults who lived either in sites with elevated soil levels of cadmium and lead (n = 1,561) or in comparison communities (n = 480). The blood lead and urine cadmium levels of participants were somewhat higher than national average mean blood lead levels were 7 {micro}g/dl for participants aged 6--35 mo; 6 {micro}g/dl for participants aged 36--71 mo, 4 {micro}g/dl for participants aged 6--15 yr; and 4.3 {micro}g/dl for participants aged 16--75 yr. Multivariate analysis indicated no marked differences in any of the immune marker distributions attributed to lead for adults or children over 3 yr of age. However, in children under age 3, increased blood lead levels, principally those over 15 {micro}g/dl were associated with increases in IgA, IgC, IgM, and circulating B/lymphocytes. Youth adults urine cadmium levels over 1.5 {micro}g/g were associated with higher levels of IgA and circulating B.

  11. Changes in lymphocyte subsets and macrophage functions from high, short-term dietary ethanol in C57/BL6 mice

    SciTech Connect

    Watson, R.R.; Prabhala, R.H.; Abril, E.; Smith, T.L.

    1988-01-01

    Chronic administration of a diet containing 7% ethanol (36% of total calories) for 8 days to male C57/BL6 mice resulted in significant changes in functioning of macrophages. Peritoneal exudate macrophages from the ethanol-fed mice released more tumor cell cytotoxic materials upon culturing in vitro than cells from controls. However, peritoneal exudate cells continued to respond to exogenous beta carotene in vitro to produce additional cytotoxic materials. Phagocytosis of sheep red blood cells in vitro was suppressed in cells from ethanol treated mice. The number of splenic lymphocytes of various subsets was significantly changed by the ethanol exposure. Total T cells and T suppressor cells were lower, with a significant decrease in B cells containing IgM on their surface. The percentage of spleen cells showing markers for macrophage functions and their activation were significantly reduced. It is concluded that short-term chronic consumption of dietary ethanol, which was sufficient to produce physical dependence, results in significant alterations in lymphocyte subtypes and suppression of some macrophage functions.

  12. CD4+ CCR5+ and CD4+ CCR3+ lymphocyte subset and monocyte apoptosis in patients with acute visceral leishmaniasis

    PubMed Central

    Potestio, Marcella; D'Agostino, Pietro; Romano, Giuseppina Colonna; Milano, Salvatore; Ferlazzo, Viviana; Aquino, Alessandra; Di Bella, Gloria; Caruso, Rosalba; Gambino, Giuseppe; Vitale, Giustina; Mansueto, Serafino; Cillari, Enrico

    2004-01-01

    The potential involvement of apoptosis in the pathogenesis of visceral leishmaniasis (VL) was examined by studying spontaneous and Leishmania antigen (LAg)-induced apoptosis using cryopreserved peripheral blood mononuclear cells (PBMC) of Sicilian patients with VL. Results indicate that monocytes and T lymphocytes from acute VL patients show a significantly higher level of apoptosis compared with that observed in healed subjects. The percentage of apoptotic cells was higher in monocytes than in T lymphocytes. T cells involved in programmed cell death (PCD) were mainly of the CD4+ phenotype. In particular, the T helper 1-type (Th1) subset, as evaluated by chemokine receptor-5 (CCR5) expression, is involved in this process. Cell death in Th1-type uses a CD95-mediated mechanism. Furthermore, Th1-type CCR5+ cells are prone to cell suicide in an autocrine or paracrine way, as attested by enhanced expression of CD95L in acute VL patients. The reduction in Th1-type cells by apoptosis was confirmed by the decrease in interferon-γ secretion. In conclusion, apoptosis of monocytes, CD4+ and CD4+ CCR5+ T cells could be involved in the failure of cell mediated immunity that is responsible for severe immune-depression in VL. PMID:15379987

  13. T lymphocyte subsets in leprosy. A study of 24 Egyptian patients.

    PubMed

    Ashamalla, L; Yang, S J; Michel, M; Rafla, L

    1989-09-01

    Phenotypic analysis was done on 24 Egyptian leprosy patients and 11 healthy controls. The type of leprosy, duration of disease at the time of testing, and age were found to affect T cell subset distribution. As compared with controls, neural leprosy tended to have a decreased total T cell percentage, borderline leprosy an increased T suppressor cell percentage, and reactional borderline leprosy an increased T helper/suppressor ratio. Patients with the disease for less than 1 year had a higher mean percentage of T suppressor cells and a lower mean T helper/suppressor ratio than patients with leprosy for more than 1 year. The same was true in older (50-70 years old) versus younger (12-41 years old) patients. PMID:2528512

  14. Differential requirement of CD28 costimulation for activation of murine CD8+ intestinal intraepithelial lymphocyte subsets and lymph node cells.

    PubMed

    Gelfanov, V; Lai, Y G; Gelfanova, V; Dong, J Y; Su, J P; Liao, N S

    1995-07-01

    The CD8+CD4- (CD8+) murine small intestinal intraepithelial lymphocytes (IELs) contain two subpopulations, one expressing alpha alpha-CD8 homodimers and another alpha beta-CD8 heterodimers. In this study, plate-bound anti-TCR beta-chain (TCR-beta) mAb alone or combined with anti-CD28 mAb is used as a model system to study activation requirement of these two CD8+ IEL subsets. In contrast to CD8+ lymph node (LN) cells that require both TCR and CD28 triggering for activation, alpha beta-CD8+ IELs proliferate and produce IL-2 and IFN-gamma when stimulated with anti-TCR-beta mAb alone, and soluble CTLA-4 Ig has no effect on their responses. On the other hand, alpha alpha-CD8+ IELs neither make IL-2 or IFN-gamma nor proliferate even when both stimuli are provided. However, alpha alpha-CD8+ IELs are capable of proliferation in both CD8+ IEL subsets is lower than in CD8+ LN cells, which contributes to the weaker and delayed response of CD8+ IELs. PMID:7602124

  15. Age-Related Differences in Percentages of Regulatory and Effector T Lymphocytes and Their Subsets in Healthy Individuals and Characteristic STAT1/STAT5 Signalling Response in Helper T Lymphocytes

    PubMed Central

    Holcar, Marija; Goropevšek, Aleš; Ihan, Alojz; Avčin, Tadej

    2015-01-01

    The dynamic process of the development of the immune system can in itself result in age-related immune malfunctions. In this study, we analysed lymphocyte subsets in the peripheral blood of 60 healthy donors, divided into groups of children, adolescents, and adults, focusing on effector (Teff) and regulatory (Treg) T lymphocytes and STAT1/STAT5 signalling response in helper T lymphocytes (Th) in adults, using flow cytometry. Our results demonstrate a decrease in the percentage of total Tregs and an increase in the percentage of total Teffs with age and a consequential immense increase in the Teff/Treg ratio. The increase of Teffs was most apparent in Th1, Th1Th17, and Th17CD161− subsets. Significant Th lymphocyte STAT1 expression differences were observed between children and adolescents, which were associated with the decrease in activated Tregs. Higher expression of STAT1 was found in FoxP3hi than in FoxP3low Th lymphocytes, while significant IL-2 induced STAT5 phosphorylation differences were found among the subsets of Th lymphocytes in adults. Our study demonstrates age-related changes in circulating Teff and Treg, as well as significant differences in STAT5/STAT1 signalling among FoxP3+ Th lymphocytes, providing new advances in the understanding of immunosenescence. PMID:26525134

  16. Age-Related Differences in Percentages of Regulatory and Effector T Lymphocytes and Their Subsets in Healthy Individuals and Characteristic STAT1/STAT5 Signalling Response in Helper T Lymphocytes.

    PubMed

    Holcar, Marija; Goropevšek, Aleš; Ihan, Alojz; Avčin, Tadej

    2015-01-01

    The dynamic process of the development of the immune system can in itself result in age-related immune malfunctions. In this study, we analysed lymphocyte subsets in the peripheral blood of 60 healthy donors, divided into groups of children, adolescents, and adults, focusing on effector (Teff) and regulatory (Treg) T lymphocytes and STAT1/STAT5 signalling response in helper T lymphocytes (Th) in adults, using flow cytometry. Our results demonstrate a decrease in the percentage of total Tregs and an increase in the percentage of total Teffs with age and a consequential immense increase in the Teff/Treg ratio. The increase of Teffs was most apparent in Th1, Th1Th17, and Th17CD161- subsets. Significant Th lymphocyte STAT1 expression differences were observed between children and adolescents, which were associated with the decrease in activated Tregs. Higher expression of STAT1 was found in FoxP3hi than in FoxP3low Th lymphocytes, while significant IL-2 induced STAT5 phosphorylation differences were found among the subsets of Th lymphocytes in adults. Our study demonstrates age-related changes in circulating Teff and Treg, as well as significant differences in STAT5/STAT1 signalling among FoxP3+ Th lymphocytes, providing new advances in the understanding of immunosenescence. PMID:26525134

  17. Lymphocyte subset characterization associated with persistent hepatitis C virus infection and subsequent progression of liver fibrosis.

    PubMed

    Yoshida, Kengo; Ohishi, Waka; Nakashima, Eiji; Fujiwara, Saeko; Akahoshi, Masazumi; Kasagi, Fumiyoshi; Chayama, Kazuaki; Hakoda, Masayuki; Kyoizumi, Seishi; Nakachi, Kei; Hayashi, Tomonori; Kusunoki, Yoichiro

    2011-10-01

    This study aims to deepen the understanding of lymphocyte phenotypes related to the course of hepatitis C virus (HCV) infection and progression of liver fibrosis in a cohort of atomic bomb survivors. The study subjects comprise 3 groups: 162 HCV persistently infected, 145 spontaneously cleared, and 3,511 uninfected individuals. We observed increased percentages of peripheral blood T(H)1 and total CD8 T cells and decreased percentages of natural killer (NK) cells in the HCV persistence group compared with the other 2 groups after adjustment for age, gender, and radiation exposure dose. Subsequently, we determined that increased T(H)1 cell percentages in the HCV persistence group were significantly associated with an accelerated time-course reduction in platelet counts-accelerated progression of liver fibrosis-whereas T(C)1 and NK cell percentages were inversely associated with progression. This study suggests that T(H)1 immunity is enhanced by persistent HCV infection and that percentages of peripheral T(H)1, T(C)1, and NK cells may help predict progression of liver fibrosis. PMID:21712057

  18. T cell subsets differently regulate osteogenic differentiation of human mesenchymal stromal cells in vitro.

    PubMed

    Grassi, Francesco; Cattini, Luca; Gambari, Laura; Manferdini, Cristina; Piacentini, Anna; Gabusi, Elena; Facchini, Andrea; Lisignoli, Gina

    2016-04-01

    T lymphocytes play a key role in the regulation of bone homeostasis and bone healing. The inflammatory response at the site of bone injury is essential to the initiation of the bone repair program; however, an uncontrolled exposure to inflammatory environment has a negative effect on tissue regeneration - indeed, activated T cells were shown to inhibit osteogenic differentiation on human mesenchymal stromal cells (MSCs). Whether resting T cells can induce osteogenic differentiation of MSCs and what role specific T cells subset play in this process is still elusive. In this study, we sought to analyse the osteogenic gene expression profile of whole T cells, CD4 and CD8 T cells isolated from healthy donors and investigated whether secreted factors from each group modulate osteogenic differentiation of human MSCs. Gene expression profiling identified a pool of 51 genes involved at various stages in bone growth which are expressed above detectable levels in CD4 and CD8 T cells. Most genes of this pool were expressed at higher levels in the CD4 subset. In vitro mineralization assays revealed that conditioned medium from CD4 T cells, but not from CD8 cells, significantly increased mineralization in osteogenic cultures of human MSCs; furthermore, mRNA expression of Runt-related transcription factor 2 (RUNX-2), osteocalcin (OC), bone sialoprotein (BSP) and alkaline phosphatase (ALP) in MSCs was significantly upregulated in the presence of CD4-conditioned medium but not with that obtained from CD8. The results show a differential role for CD4 and CD8 T cells in supporting bone formation and identify an osteogenic gene signature of each subset. Copyright © 2013 John Wiley & Sons, Ltd. PMID:23653421

  19. Modulation of Cytokine Expression and Lymphocyte Subsets during the Periparturient Period in Dairy Cows Naturally Infected with Mycobacterium avium subsp. paratuberculosis

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The objective of this study was to evaluate cytokine gene expression and populations of lymphocyte subsets in periparturient dairy cows naturally infected with Mycobacterium avium subsp. paratuberculosis (MAP). Blood was collected from noninfected, subclinical, and clinical MAP-infected dairy cows ...

  20. Human Decidual Natural Killer Cells Are a Unique NK Cell Subset with Immunomodulatory Potential

    PubMed Central

    Koopman, Louise A.; Kopcow, Hernan D.; Rybalov, Basya; Boyson, Jonathan E.; Orange, Jordan S.; Schatz, Frederick; Masch, Rachel; Lockwood, Charles J.; Schachter, Asher D.; Park, Peter J.; Strominger, Jack L.

    2003-01-01

    Natural killer cells constitute 50–90% of lymphocytes in human uterine decidua in early pregnancy. Here, CD56bright uterine decidual NK (dNK) cells were compared with the CD56bright and CD56dim peripheral NK cell subsets by microarray analysis, with verification of results by flow cytometry and RT-PCR. Among the ∼10,000 genes studied, 278 genes showed at least a threefold change with P ≤ 0.001 when comparing the dNK and peripheral NK cell subsets, most displaying increased expression in dNK cells. The largest number of these encoded surface proteins, including the unusual lectinlike receptors NKG2E and Ly-49L, several killer cell Ig-like receptors, the integrin subunits αD, αX, β1, and β5, and multiple tetraspanins (CD9, CD151, CD53, CD63, and TSPAN-5). Additionally, two secreted proteins, galectin-1 and progestagen-associated protein 14, known to have immunomodulatory functions, were selectively expressed in dNK cells. PMID:14568979

  1. Radiosensitivity of human natural killer cells: Binding and cytotoxic activities of natural killer cell subsets

    SciTech Connect

    Rana, R.; Vitale, M.; Mazzotti, G.; Manzoli, L.; Papa, S. )

    1990-10-01

    The sensitivity of human natural killer (NK) cell activities (both binding and killing) after exposure of peripheral blood mononuclear cells to different doses of gamma radiation was studied. A panel of monoclonal antibodies was used to identify the NK and T-lymphocyte subsets and to evaluate their radiosensitivity. Peripheral blood mononuclear cells were irradiated with low (2-6 Gy) and high (10-30 Gy) doses and NK cell binding and cytotoxic activity against K562 target cells were studied after 3 h and 48 h in culture. The primary damage to NK cell activity was identified at the postbinding level and affected mainly the lytic machinery. After 48 h culture postirradiation, an overall depression of cytotoxic activity was observed, but ionizing radiation produced either a selection of the more cytotoxic NK cell subsets, which therefore might be considered more resistant to radiation damage than the less cytotoxic NK cells, or a long-term stimulation of cytotoxic activity in surviving cells.

  2. Evidence for genetic restriction in the suppression of erythropoiesis by a unique subset of T lymphocytes in man.

    PubMed Central

    Lipton, J M; Nadler, L M; Canellos, G P; Kudisch, M; Reiss, C S; Nathan, D G

    1983-01-01

    The suppression of erythropoiesis by lymphocytes from patients with a T cell lymphoproliferative syndrome and pure erythrocyte aplasia has been previously demonstrated. To study the nature of the suppressor cell and possible genetic restriction of this suppression, we investigated a patient with severe anemia, splenomegaly, lymphocytosis, and erythroid aplasia. A 3-mo course of low-dose daily oral cyclophosphamide achieved a complete remission for over 12 mo. The surface phenotype of his lymphocytes was analyzed by means of antibodies to lineage, differentiation, and activation-specific surface antigens. The cells expressed mature T cell antigens T3, T8, and T11, while lacking T1. Immature T cell, B cell, and the monocyte-specific antigen Mo2 were absent, while Mo1, a monocyte-associated antigen not normally expressed on T cells, was present. T10 and Ia expressed as activation antigens were also present. The cells, cryopreserved at diagnosis, were thawed and co-cultured in plasma clot with patient remission marrow samples at T cell/bone marrow ratios of 1:1 and 2:1. There was nearly 90% suppression of erythroid colony-forming unit expression and 60% suppression of erythroid burst-forming unit expression at 2:1 T cell to bone marrow ratios and somewhat less suppression at 1:1. Granulocyte/macrophage progenitor expression was unaffected. Erythroid progenitor differentiation in the marrows of two HLA identical siblings was similarly suppressed. The cells were co-cultured with the marrows of nine nonrelated donors to investigate the potential genetic restriction of this suppression. Colony suppression equal to that observed in the marrow of the patient and his siblings was found in studies of two partially HLA identical individuals. No suppression was detected in marrow co-cultures of two entirely HLA dissimilar individuals. These results show that suppression of erythropoiesis by a unique subset of T8, Mo1, Ia-positive lymphocytes isolated from a patient with

  3. Aryl hydrocarbon mono-oxygenase activity in human lymphocytes

    SciTech Connect

    Griffin, G.D.; Schuresko, D.D.

    1981-06-01

    Aryl hydrocarbon mono-oxygenase (AHM), an enzyme of key importance in metabolism of xenobiotic chemicals such as polynuclear aromatic hydrocarbons (PNA), is present in human lymphocytes. Studies investing the relation of activity of AHM in human lymphocytes to parameters such as disease state, PNA exposure, in vitro mitogen stimulation, etc. have been summarized in this report. Some studies have demonstrated increased AHM activity in lymphocytes from cigarette smokers (compared to nonsmokers), and in lung cancer patients when compared to appropriate control groups. These observations are confused by extreme variability in human lymphocyte AHM activities, such variability arising from factors such as genetic variation in AHM activity, variation in in vitro culture conditions which affect AHM activity, and the problematical relationship of common AHM assays to actual PNA metabolism taking place in lymphocytes. If some of the foregoing problems can be adequately addressed, lymphocyte AHM activity could hold the promise of being a useful biomarker system for human PNA exposure.

  4. Mutations in TLR/MYD88 pathway identify a subset of young chronic lymphocytic leukemia patients with favorable outcome.

    PubMed

    Martínez-Trillos, Alejandra; Pinyol, Magda; Navarro, Alba; Aymerich, Marta; Jares, Pedro; Juan, Manel; Rozman, María; Colomer, Dolors; Delgado, Julio; Giné, Eva; González-Díaz, Marcos; Hernández-Rivas, Jesús M; Colado, Enrique; Rayón, Consolación; Payer, Angel R; Terol, Maria José; Navarro, Blanca; Quesada, Victor; Puente, Xosé S; Rozman, Ciril; López-Otín, Carlos; Campo, Elías; López-Guillermo, Armando; Villamor, Neus

    2014-06-12

    Mutations in Toll-like receptor (TLR) and myeloid differentiation primary response 88 (MYD88) genes have been found in chronic lymphocytic leukemia (CLL) at low frequency. We analyzed the incidence, clinicobiological characteristics, and outcome of patients with TLR/MYD88 mutations in 587 CLL patients. Twenty-three patients (3.9%) had mutations, 19 in MYD88 (one with concurrent IRAK1 mutation), 2 TLR2 (one with concomitant TLR6 mutation), 1 IRAK1, and 1 TLR5. No mutations were found in IRAK2 and IRAK4. TLR/MYD88-mutated CLL overexpressed genes of the nuclear factor κB pathway. Patients with TLR/MYD88 mutations were significantly younger (83% age ≤50 years) than those with no mutations. TLR/MYD88 mutations were the most frequent in young patients. Patients with mutated TLR/MYD88 CLL had a higher frequency of mutated IGHV and low expression of CD38 and ZAP-70. Overall survival (OS) was better in TLR/MYD88-mutated than unmutated patients in the whole series (10-year OS, 100% vs 62%; P = .002), and in the subset of patients age ≤50 years (100% vs 70%; P = .02). In addition, relative OS of TLR/MYD88-mutated patients was similar to that in the age- and gender-matched population. In summary, TLR/MYD88 mutations identify a population of young CLL patients with favorable outcome. PMID:24782504

  5. Systematic and Cell Type-Specific Telomere Length Changes in Subsets of Lymphocytes

    PubMed Central

    Lin, Jue; Cheon, Joshua; Brown, Rashida; Coccia, Michael; Puterman, Eli; Aschbacher, Kirstin; Sinclair, Elizabeth; Epel, Elissa; Blackburn, Elizabeth H.

    2016-01-01

    Telomeres, the protective DNA-protein complexes at the ends of linear chromosomes, are important for genome stability. Leukocyte or peripheral blood mononuclear cell (PBMC) telomere length is a potential biomarker for human aging that integrates genetic, environmental, and lifestyle factors and is associated with mortality and risks for major diseases. However, only a limited number of studies have examined longitudinal changes of telomere length and few have reported data on sorted circulating immune cells. We examined the average telomere length (TL) in CD4+, CD8+CD28+, and CD8+CD28− T cells, B cells, and PBMCs, cross-sectionally and longitudinally, in a cohort of premenopausal women. We report that TL changes over 18 months were correlated among these three T cell types within the same participant. Additionally, PBMC TL change was also correlated with those of all three T cell types, and B cells. The rate of shortening for B cells was significantly greater than for the three T cell types. CD8+CD28− cells, despite having the shortest TL, showed significantly more rapid attrition when compared to CD8+CD28+ T cells. These results suggest systematically coordinated, yet cell type-specific responses to factors and pathways contribute to telomere length regulation. PMID:26977417

  6. CD38 expression labels an activated subset within chronic lymphocytic leukemia clones enriched in proliferating B cells

    PubMed Central

    Damle, Rajendra N.; Temburni, Sonal; Calissano, Carlo; Yancopoulos, Sophia; Banapour, Taraneh; Sison, Cristina; Allen, Steven L.; Rai, Kanti R.

    2007-01-01

    Chronic lymphocytic leukemia (CLL) cells are thought to have diminished cell-cycling capacity, a view challenged by their phenotypic resemblance to activated human B lymphocytes. The present study addresses the cell-cycling status of CLL cells, focusing on those leukemic cells expressing CD38, a molecule involved in signaling and activation that also serves as a prognostic marker in this disease. CD38+ and CD38− members of individual CLL clones were analyzed for coexpression of molecules associated with cellular activation (CD27, CD62L, and CD69), cell-cycle entry (Ki-67), signaling (ZAP-70), and protection from apoptosis (telomerase and Bcl-2). Regardless of the size of the CD38+ fraction within a CLL clone, CD38+ subclones are markedly enriched for expression of Ki-67, ZAP-70, human telomerase reverse transcriptase, and telomerase activity. Although the percentage of cells (approximately 2%) entering the cell cycle as defined by Ki-67 expression is small, the absolute number within a clone can be sizeable and is contained primarily within the CD38+ fraction. Despite these activation/proliferation differences, both CD38+ and CD38− fractions have similar telomere lengths, suggesting that CD38 expression is dynamic and transient. These findings may help explain why high percentages of CD38+ cells within clones are associated with poor clinical outcome. PMID:17684154

  7. The effect of 5-amino-3-methyl-4-isoxazolecarboxylic acid hydrazide on lymphocyte subsets and humoral immune response in SRBC-immunized mice.

    PubMed

    Drynda, Angelika; Obmińska-Mrukowicz, Bożena; Mączyński, Marcin; Ryng, Stanisław

    2015-04-01

    5-Amino-3-methyl-4-isoxazolecarboxylic acid hydrazide is a non-cytotoxic synthetic isoxazole derivative with considerable immunomodulatory properties demonstrated in in vitro experiments. The aim of this study was to investigate the influence of this compound, depending on the dosage and schedule of treatment, on lymphocyte subsets in non-immunized mice and humoral immune response in SRBC (sheep red blood cells)-immunized mice. An analysis of lymphocyte subsets was carried out by flow cytometry, using specific monoclonal antibodies stained with fluorescein isothiocyanate (FITC) or phycoerythrin (PE). In the SRBC-immunized mice, the influence of the compound on the humoral response was determined, depending on the time of administration relative to the antigen. The number of plaque forming cells (PFC) was determined by a local hemolysis technique in an agar gel. Total and 2-mercaptoethanol resistant serum agglutination titers were defined by active hemagglutination test carried out on microplates. The investigated hydrazide was able to modulate the percentage and absolute number of T lymphocyte subsets in the thymus, and T and B lymphocytes in the peripheral lymphatic organs. It also enhanced humoral immune response in SRBC-immunized mice by increasing the number of cells producing hemolytic anti-SRBC antibodies (PFC) and by augmenting the level of total and 2-mercaptoethanol resistant hemagglutinin. The present study showed modulatory effects of 5-amino-3-methyl-4-isoxazolecarboxylic acid hydrazide on lymphocyte subsets and humoral immune response in mice. This compound could be potentially useful for the treatment of autoimmune diseases, infections or as an adjuvant for boosting the efficacy of vaccines. PMID:25572572

  8. Different roles for CD4+ and CD8+ T lymphocytes and macrophage subsets in the control of a generalized virus infection.

    PubMed Central

    Karupiah, G; Buller, R M; Van Rooijen, N; Duarte, C J; Chen, J

    1996-01-01

    The importance of T-lymphocyte subsets in the control of poxvirus infections is controversial. To determine the relative contribution of lymphocyte subsets important for recovery from infection with ectromelia virus (EV), a natural murine poxvirus pathogen, C57BL/6 (B6) mice lacking functional CD8+ T cells because of disruption of the beta2-microglobulin gene or lacking functional CD4+ T cells because of disruption of the I-(A)beta gene, acutely depleted of CD8+ or CD4+ T cells with monoclonal antibody, or depleted of macrophage subsets by the macrophage suicide technique were used. Recovery from infection was strictly dependent on the effector functions of CD8+ T cells, in the absence of which 100% mortality resulted. This lymphocyte population had demonstrable antiviral activity early in the infection process even before class I major histocompatibility complex (MHC)-restricted CD8+ cytotoxic T-lymphocyte (CTL) activity was detectable. CD4+ T cells were found to be necessary for the generation of an optimal virus-specific, class I MHC-restricted CD8+ CTL response and contributed to virus clearance not involving cytolytic mechanisms. In both models of CD4+ T-cell deficiency, virus clearance was incomplete and persisted at low levels in most organs and at very high levels in the skin, but the animals did not die. The elimination of macrophage subpopulations impeded virus clearance, impaired the generation of class I MHC-restricted antiviral CTL response, and resulted in 100% mortality. These findings establish an absolute requirement for CD8+ and CD4+ T lymphocytes and macrophage subsets in the elimination of a natural murine poxvirus infection and support the idea that macrophages may be essential accessory cells for the generation of class I MHC-restricted antiviral CTL responses. PMID:8970949

  9. In vitro generated anti-tumor T lymphocytes exhibit distinct subsets mimicking in vivo antigen experienced cells

    PubMed Central

    Yang, Shicheng; Gattinoni, Luca; Liu, Fang; Ji, Yun; Yu, Zhiya; Restifo, Nicholas P.; Rosenberg, Steven A.; Morgan, Richard A.

    2011-01-01

    The T lymphocyte pool can be sub-divided into naïve (Tn), effector memory (Tem), and central memory (Tcm) T cells. In this study, we characterized in vitro short-term cultured anti-tumor human T lymphocytes generated by lentiviral transduction with an anti-tumor antigen TCR vector. Within two weeks of in vitro culture, the cultured T cells showed a Tcm-like phenotype illustrated by a high percentage of CD62L and CD45RO cells. When the cells were sorted into populations that were CD45RO+/CD62L− (Tem), CD45RO+/CD62L+ (Tcm) or CD45ROlow/CD62L+ (Tn) and co-cultured with antigen-matched tumor lines, the magnitude of cytokine release from these populations for IFNγ (TnTcm>Tem) mimicked the types of immune cell responses observed in vivo. In comparing cell-mediated effector function, Tn were found to be deficient (relative to Tcm and Tem) in the ability to form conjugates with tumor cells and subsequent lytic activity. Moreover, analysis of the gene expression profiles of the in vitro cultured and sorted T cell populations also demonstrated patterns consistent with their in vivo counterparts. When Tcm and Tem were tested for the ability to survive in vivo, Tcm displayed significantly increased engraftment and persistence in NOD/SCID/γc−/− mice. In general, a large percentage of in vitro generated anti-tumor T lymphocytes mimic a Tcm-like phenotype (based on phenotype, effector function, and increased persistence in vivo), which suggests that these Tcm-like cultured T cells may be optimal for adoptive immunotherapy. PMID:21305379

  10. Characterization of a chromosome-specific chimpanzee alpha satellite subset: Evolutionary relationship to subsets on human chromosomes

    SciTech Connect

    Warburton, P.E.; Gosden, J.; Lawson, D.

    1996-04-15

    Alpha satellite DNA is a tandemly repeated DNA family found at the centromeres of all primate chromosomes examined. The fundamental repeat units of alpha satellite DNA are diverged 169- to 172-bp monomers, often found to be organized in chromosome-specific higher-order repeat units. The chromosomes of human (Homo sapiens (HSA)), chimpanzee (Pan troglodytes (PTR) and Pan paniscus), and gorilla (Gorilla gorilla) share a remarkable similarity and synteny. It is of interest to ask if alpha satellite arrays at centromeres of homologous chromosomes between these species are closely related (evolving in an orthologous manner) or if the evolutionary processes that homogenize and spread these arrays within and between chromosomes result in nonorthologous evolution of arrays. By using PCR primers specific for human chromosome 17-specific alpha satellite DNA, we have amplified, cloned, and characterized a chromosome-specific subset from the PTR chimpanzee genome. Hybridization both on Southern blots and in situ as well as sequence analysis show that this subset is most closely related, as expected, to sequences on HSA 17. However, in situ hybridization reveals that this subset is not found on the homologous chromosome in chimpanzee (PTR 19), but instead on PTR 12, which is homologous to HSA 2p. 40 refs., 3 figs.

  11. Detection of TCD4+ subsets in human carotid atheroma.

    PubMed

    Oliveira, Rômulo Tadeu Dias; Silva, Rosiane Maria; Teo, Fabio Haach; Mineiro, Marcela Franco; Ferreira, Maria Carolina; Altemani, Albina; Mamoni, Ronei Luciano; Menezes, Fábio Husseman; Blotta, Maria Heloisa Souza Lima

    2013-04-01

    Activated TCD4(+) cells are detected in human atherosclerotic plaques which indicate their participation in disease progression and destabilization. Among these cells, IFN-γ-producing T cells (TH1) are recognized as having a pro-atherogenic role. Recently, the IL-17-producing T helper lineage of cells (TH17) has been identified in atherosclerotic lesions. They have been linked to atheroma development through the production of pro-inflammatory mediators present in these lesions. Furthermore, IL-22 producing TCD4(+) cells (TH22) have been identified in the atheromatous environment, but their presence and function has not been investigated. The aim of this study was to analyze the immune response mediated by pro-inflammatory subtypes of TCD4(+) cells in atheromatous lesions. Atherosclerotic plaques of 57 patients with critical stenosis of carotid submitted to endarterectomy were evaluated. Three carotid fragments from organ donors were used as control. mRNA analysis showed expression of TH1 (IFN-γ, T-bet, IL-2, IL-12p35, TNF-α and IL-18); TH2 (GATA-3); TH17 (IL-17A, IL-17RA, Rorγt, TGF-β, IL-6, IL-1β, IL-23p19, CCL20, CCR4 and CCR6) and TH22 (IL-22 and Ahr) related markers. Asymptomatic patients showed higher expression of mRNA of IL-10, TGF-β, CCR4 and GATA-3 when compared to symptomatic ones. Immunohistochemistry analysis showed higher levels of IL-23, TGF-β, IL-1β and IL-18 in macrophages and foam cells in unstable lesions compared to stable and control ones. In vitro stimulation of atheroma cells induced IL-17 and IFN-γ production. Finally we were able to detect, the following subpopulations of TCD3(+) cells: TCD4(+) IFN-γ(+), TCD4(+)IL-17(+), TCD4(+)IL-4(+), TCD4(+)IL-22(+) and double positive cells (IFN-γ/IL-17(+), IFN-γ/IL-22(+) or IL-17/IL-22(+)). Our results showed the presence of distinct TCD4(+) cells subsets in human carotid lesions and suggest that interactions among them may contribute to the atheroma progression and destabilization. PMID

  12. In vitro effects of flunarizine on human lymphocytes.

    PubMed

    Brohée, D; Piro, P; Kennes, B; Nève, P

    1986-01-01

    Flunarizine, a slow-channel calcium entry blocker used as a vasodilator, interferes in vitro with human lymphocyte functions. It prevents lymphocytes from capping sheep erythrocytes, an effect which is probably due to the disconnection of the membrane from its cytoskeletal control. Flunarizine antagonizes both colchicine and cytochalasin B effects upon capping. Although the mitogen-induced lymphocyte stimulation has been shown to be sensitive to calcium depletion or calcium entry blocking by Verapamil, an enhanced response to phytohaemagglutinin A was observed with flunarizine. This suggests a differential sensitivity of the lymphocytes to calcium-entry blockers. PMID:3731875

  13. Mechanisms of inhibition of Cryptococcus neoformans by human lymphocytes.

    PubMed Central

    Levitz, S M; North, E A; Dupont, M P; Harrison, T S

    1995-01-01

    Recently, our laboratory and others have demonstrated that human peripheral blood T and NK lymphocytes directly inhibit the growth of Cryptococcus neoformans. In this study, we further define the conditions under which lymphocyte-mediated fungistasis against C. neoformans occurs and examine whether mechanisms implicated in lymphocyte-mediated activities against other target cells are also involved in anticryptococcal activity. The addition of whole or broken heat-killed C. neoformans modestly inhibited lymphocyte-mediated fungistasis, whereas other particulates had no effect. The hydroxyl radical scavenger catechin, but not diethyl urea or propyl gallate, profoundly inhibited fungistasis. Salicylic acid inhibited fungistasis in a dose-dependent fashion. However, two other cyclooxygenase inhibitors, piroxicam and indomethacin, had no effect, suggesting that the mechanism of inhibition by salicylic acid was cyclooxygenase independent. Reagent prostaglandin E2, at concentrations shown by others to inhibit NK cell-mediated bactericidal and tumorlytic activities, had no effect on lymphocyte-mediated fungistasis. The addition of selected monoclonal antibodies or ligands reactive with receptors on human lymphocytes had no significant effect on lymphocyte-mediated fungistasis. Acapsular, small-capsuled, and large-capsuled C. neoformans organisms were inhibited by lymphocytes to an approximately equal extent. These data demonstrate that lymphocyte-mediated activity against C. neoformans proceeds regardless of the presence of capsule and by mechanisms at least in part dissimilar from those seen with other target cells. PMID:7642290

  14. Human Monocyte Subsets at Homeostasis and Their Perturbation in Numbers and Function in Filarial Infection

    PubMed Central

    Moore, Vanessa; Bennuru, Sasisekhar; McDonald-Fleming, Renee; Ganesan, Sundar; Cotton, Rachel; Anuradha, Rajamanickam; Babu, Subash; Nutman, Thomas B.

    2014-01-01

    To characterize the function and plasticity of the major human circulating monocyte populations and to explore their role in systemic helminth infection, highly purified (by flow-based sorting) human monocyte subsets (CD14hi/CD16neg [classical], CD14+ or hi/CD16med [intermediate], and CD14neg/CD16hi [nonclassical]) were examined at homeostasis and after activation. Among these three subsets the classical and intermediate subsets were found to be the major sources of inflammatory and regulatory cytokines, as well as cytokines/chemokines associated with alternative activation, whereas the nonclassical and classical populations demonstrated an ability to transmigrate through endothelial monolayers. Moreover, it was primarily the classical subset that was the most efficient in promoting autologous T cell proliferation. The distribution of these subsets changed in the context of a systemic helminth (Wuchereria bancrofti) infection such that patent infection altered the frequency and distribution of these monocyte subsets with the nonclassical monocytes being expanded (almost 2-fold) in filarial infection. To understand further the filarial/monocyte interface, in vitro modeling demonstrated that the classical subset internalized filarial antigens more efficiently than the other two subsets but that the parasite-driven regulatory cytokine interleukin-10 was exclusively coming from the intermediate subset. Our data suggest that monocyte subsets have a differential function at homeostasis and in response to helminth parasites. PMID:25114121

  15. Long Intergenic Non-Coding RNAs: Novel Drivers of Human Lymphocyte Differentiation

    PubMed Central

    Panzeri, Ilaria; Rossetti, Grazisa; Abrignani, Sergio; Pagani, Massimiliano

    2015-01-01

    Upon recognition of a foreign antigen, CD4+ naïve T lymphocytes proliferate and differentiate into subsets with distinct functions. This process is fundamental for the effective immune system function, as CD4+ T cells orchestrate both the innate and adaptive immune response. Traditionally, this differentiation event has been regarded as the acquisition of an irreversible cell fate so that memory and effector CD4+ T subsets were considered terminally differentiated cells or lineages. Consequently, these lineages are conventionally defined thanks to their prototypical set of cytokines and transcription factors. However, recent findings suggest that CD4+ T lymphocytes possess a remarkable phenotypic plasticity, as they can often re-direct their functional program depending on the milieu they encounter. Therefore, new questions are now compelling such as which are the molecular determinants underlying plasticity and stability and how the balance between these two opposite forces drives the cell fate. As already mentioned, in some cases, the mere expression of cytokines and master regulators could not fully explain lymphocytes plasticity. We should consider other layers of regulation, including epigenetic factors such as the modulation of chromatin state or the transcription of non-coding RNAs, whose high cell-specificity give a hint on their involvement in cell fate determination. In this review, we will focus on the recent advances in understanding CD4+ T lymphocytes subsets specification from an epigenetic point of view. In particular, we will emphasize the emerging importance of non-coding RNAs as key players in these differentiation events. We will also present here new data from our laboratory highlighting the contribution of long non-coding RNAs in driving human CD4+ T lymphocytes differentiation. PMID:25926836

  16. Tempol protects human lymphocytes from genotoxicity induced by cisplatin.

    PubMed

    Khabour, Omar F; Alzoubi, Karem H; Mfady, Doa'a S; Alasseiri, Mohammed; Hasheesh, Taghrid F

    2014-01-01

    The use of cisplatin in treatments of human malignancies is limited by its side effects that include DNA damage and the subsequent risk of developing secondary cancer. In this study, we examined the possible protective effect of Tempol against DNA damage induced by cisplatin in human lymphocytes using chromosomal aberrations (CAs) and sister chromatid exchanges (SCEs) assays. Cisplatin induced significant elevation in the frequencies of CAs and SCEs in cultured human lymphocytes (P < 0.01). Treatment of lymphocytes with Tempol significantly lowered CAs and SCEs induced by cisplatin. Tempol alone did not affect spontaneous levels of SCEs and CAs observed in the control group (P > 0.05). In conclusion, Tempol protects human lymphocytes against genotoxicity induced by the anticancer drug cisplatin. PMID:24955171

  17. Altered cytokine production by specific human peripheral blood cell subsets immediately following space flight

    NASA Technical Reports Server (NTRS)

    Crucian, B. E.; Cubbage, M. L.; Sams, C. F.

    2000-01-01

    In this study, flow cytometry was used to positively identify the specific lymphocyte subsets exhibiting space flight-induced alterations in cytokine production. Whole blood samples were collected from 27 astronauts at three points (one preflight, two postflight) surrounding four space shuttle missions. Assays performed included serum/urine stress hormones, white blood cell (WBC) phenotyping, and intracellular cytokine production following mitogenic stimulation. Absolute levels of peripheral granulocytes were significantly elevated following space flight, but the levels of circulating lymphocytes and monocytes were unchanged. Lymphocyte subset analysis demonstrated a decreased percentage of T cells, whereas percentages of B cells and natural killer (NK) cells remained unchanged after flight. Nearly all the astronauts exhibited an increased CD4/CD8 T cell ratio. Assessment of naive (CD45RA+) vs. memory (CD45RO+) CD4+ T cell subsets was ambiguous, and subjects tended to group within specific missions. Although no significant trend was seen in absolute monocyte levels, a significant decrease in the percentage of the CD14+ CD16+ monocytes was seen following space flight in all subjects tested. T cell (CD3+) production of interleukin-2 (IL-2) was significantly decreased after space flight, as was IL-2 production by both CD4+ and CD8+ T cell subsets. Production of interferon-gamma (IFN-gamma) was not altered by space flight for the CD8+ cell subset, but there was a significant decrease in IFN-gamma production for the CD4+ T cell subset. Serum and urine stress hormone analysis indicated significant physiologic stresses in astronauts following space flight. Altered peripheral leukocyte subsets, altered serum and urine stress hormone levels, and altered T cell cytokine secretion profiles were all observed postflight. In addition, there appeared to be differential susceptibility to space flight regarding cytokine secretion by T cell subsets. These alterations may be the

  18. Mitogenic effect of Parkia speciosa seed lectin on human lymphocytes.

    PubMed

    Suvachittanont, W; Jaranchavanapet, P

    2000-12-01

    Mitogenic activity of a lectin, purified from Parkia speciosa seeds, on the isolated peripheral blood lymphocytes taken from normal blood donors and patients with esophageal carcinoma was examined using [3H]thymidine incorporation. The lectin increases the incorporation of [3H]thymidine into DNA of human lymphocytes. The activity of the lectin increased as its concentration was increased and then declined once the concentration passed an optimum point. The stimulant effect was also expressed using a proliferation index (PI): the ratio of [3H]thymidine incorporated into lymphocytes in the presence and absence of the lectin. The mitogenic activity of the lectin is comparable to those of the known T-cell mitogens, such as concanavalin A, phytohaemagglutinin, and pokeweed mitogen. Only slightly less responsiveness was observed in the case of lymphocytes from esophageal cancer compared to lymphocytes from normal donors. PMID:11199124

  19. Adenosine metabolism in phytohemagglutinin-stimulated human lymphocytes.

    PubMed Central

    Snyder, F F; Mendelsohn, J; Seegmiller, J E

    1976-01-01

    The association of a human genetic deficiency of adenosine deaminase activity with combined immunodeficiency prompted a study of the effects of adenosine and of inhibition of adenosine deaminase activity on human lymphocyte transformation and a detailed study of adenosine metabolism throughout phytohemagglutinin-induced blastogenesis. The adenosine deaminase inhibitor, coformycin, at a concentration that inhibited adenosine deaminase activity more than 95%, or 50 muM adenosine, did not prevent blastogenesis by criteria of morphology or thymidine incorporation into acid-precipitable material. The combination of coformycin and adenosine, however, substantially reduced both the viable cell count and the incorporation of thymidine into DNA in phytohemagglutinin-stimulated lymphocytes. Incubation of lymphocytes with phytohemagglutinin for 72 h produced a 12-fold increase in the rate of deamination and a 6-fold increase in phosphorylation of adenosine by intact lymphocytes. There was no change in the apparent affinity for adenosine with either deamination or phosphorylation. The increased rates of metabolism, apparent as early as 3 h after addition of mitogen, may be due to increased entry of the nucleoside into stimulated lymphocytes. Increased adenosine metabolism was not due to changes in total enzyme activity; after 72 h in culture, the ratios of specific activities in extracts of stimulated to unstimulated lymphocytes were essentially unchanged for adenosine kinase, 0.92, and decreased for adenosine deaminase, 0.44. As much as 38% of the initial lymphocyte adenosine deaminase activity accumulated extracellularly after a 72-h culture with phytohemagglutinin. In phytohemagglutinin-stimulated lymphocytes, the principal route of adenosine metabolism was phosphorylation at less than 5 muM adenosine, and deamination at concentrations greater than 5 muM. In unstimulated lymphocytes, deamination was the principal route of adenosine metabolism over the range of adenosine

  20. Mutant Huntingtin Does Not Affect the Intrinsic Phenotype of Human Huntington’s Disease T Lymphocytes

    PubMed Central

    Miller, James R. C.; Träger, Ulrike; Andre, Ralph; Tabrizi, Sarah J.

    2015-01-01

    Huntington’s disease is a fatal neurodegenerative condition caused by a CAG repeat expansion in the huntingtin gene. The peripheral innate immune system is dysregulated in Huntington’s disease and may contribute to its pathogenesis. However, it is not clear whether or to what extent the adaptive immune system is also involved. Here, we carry out the first comprehensive investigation of human ex vivo T lymphocytes in Huntington’s disease, focusing on the frequency of a range of T lymphocyte subsets, as well as analysis of proliferation, cytokine production and gene transcription. In contrast to the innate immune system, the intrinsic phenotype of T lymphocytes does not appear to be affected by the presence of mutant huntingtin, with Huntington’s disease T lymphocytes exhibiting no significant functional differences compared to control cells. The transcriptional profile of T lymphocytes also does not appear to be significantly affected, suggesting that peripheral immune dysfunction in Huntington’s disease is likely to be mediated primarily by the innate rather than the adaptive immune system. This study increases our understanding of the effects of Huntington’s disease on peripheral tissues, while further demonstrating the differential effects of the mutant protein on different but related cell types. Finally, this study suggests that the potential use of novel therapeutics aimed at modulating the Huntington’s disease innate immune system should not be extended to include the adaptive immune system. PMID:26529236

  1. Activation of human lymphocytes by supernatants from human thymic epithelium.

    PubMed Central

    Goust, J M; Vesole, D H; Fudenberg, H H

    1979-01-01

    Supernatants from human thymic epithelial cells (TS) were found to have a mitogenic effect on cultured human peripheral blood mononuclear cells and to potentiate their responses to lectins. This was not observed with culture supernatants from the human cell lines AV-3 and HeLa or from the murine cell line L-929. The maximum potentiating effects were observed with pokeweed mitogen (PWM) and phytohaemagglutinin (PHA), whereas the response to concanavalin A (Con A) was only slightly enhanced. TS also potentiated the mixed lymphocyte culture (MLC) response of normal T cells and thymocytes cultured with mitomycin C-treated B lymphoid cell lines. The mitogenic effect of TS was time-dependent and paralleled the appearance of lymphoid colonies in semi-solid agar. Chromatographical separation of concentrated serum-free TS on Sephadex G-100 yielded an active fraction of molecular weight 15,000--25,000 which had all the activities of unseparated TS. PMID:160851

  2. Activation of human lymphocytes by supernatants from human thymic epithelium.

    PubMed

    Goust, J M; Vesole, D H; Fudenberg, H H

    1979-11-01

    Supernatants from human thymic epithelial cells (TS) were found to have a mitogenic effect on cultured human peripheral blood mononuclear cells and to potentiate their responses to lectins. This was not observed with culture supernatants from the human cell lines AV-3 and HeLa or from the murine cell line L-929. The maximum potentiating effects were observed with pokeweed mitogen (PWM) and phytohaemagglutinin (PHA), whereas the response to concanavalin A (Con A) was only slightly enhanced. TS also potentiated the mixed lymphocyte culture (MLC) response of normal T cells and thymocytes cultured with mitomycin C-treated B lymphoid cell lines. The mitogenic effect of TS was time-dependent and paralleled the appearance of lymphoid colonies in semi-solid agar. Chromatographical separation of concentrated serum-free TS on Sephadex G-100 yielded an active fraction of molecular weight 15,000--25,000 which had all the activities of unseparated TS. PMID:160851

  3. Delineation of a novel dendritic-like subset in human spleen

    PubMed Central

    Petvises, Sawang; Talaulikar, Dipti; O'Neill, Helen C

    2016-01-01

    Dendritic cells (DCs) and monocyte subpopulations present in the human spleen were analyzed by flow cytometry in an attempt to identify the presence of a novel dendritic-like cell subset described previously in mice and named L-DCs. In this study, an equivalent of this novel murine subset was characterized in the human spleen, thus increasing our knowledge of the antigen-presenting cell types present in the human spleen. Human L-DCs were identified as a hCD11c+hCD11b+HLA-DR−hCD86+ subset in the spleen, along with the previously described subsets of hCD1c+ DCs, hCD123+ plasmacytoid DCs (pDCs), hCD16+ DCs and hCD141+ DCs. Three subsets of monocytes were also characterized. DC and monocyte subsets in human spleen had phenotypes similar to those of subsets in human blood. In line with murine studies, the presence of L-DC progenitors within the spleen was also investigated. When human splenocytes depleted of T and B cells were cocultured with the murine stromal line 5G3, hematopoiesis ensued and hCD11c+HLA-DR+ and hCD11c+HLA-DR− cells were produced. The latter resemble L-DCs, which are also produced in murine spleen cocultures. Both subsets expressed hCD80 and hCD86, which identifies them as antigen-presenting cells, particularly DCs, and were highly endocytic. It is noteworthy that murine splenic stroma can serve as a support matrix for human hematopoiesis and DC production. These results support the hypothesis that 5G3 must express both cell-associated and soluble factors that can signal hematopoiesis in human and murine progenitors. PMID:25891217

  4. Delineation of a novel dendritic-like subset in human spleen.

    PubMed

    Petvises, Sawang; Talaulikar, Dipti; O'Neill, Helen C

    2016-07-01

    Dendritic cells (DCs) and monocyte subpopulations present in the human spleen were analyzed by flow cytometry in an attempt to identify the presence of a novel dendritic-like cell subset described previously in mice and named L-DCs. In this study, an equivalent of this novel murine subset was characterized in the human spleen, thus increasing our knowledge of the antigen-presenting cell types present in the human spleen. Human L-DCs were identified as a hCD11c(+)hCD11b(+)HLA-DR(-)hCD86(+) subset in the spleen, along with the previously described subsets of hCD1c(+) DCs, hCD123(+) plasmacytoid DCs (pDCs), hCD16(+) DCs and hCD141(+) DCs. Three subsets of monocytes were also characterized. DC and monocyte subsets in human spleen had phenotypes similar to those of subsets in human blood. In line with murine studies, the presence of L-DC progenitors within the spleen was also investigated. When human splenocytes depleted of T and B cells were cocultured with the murine stromal line 5G3, hematopoiesis ensued and hCD11c(+)HLA-DR(+) and hCD11c(+)HLA-DR(-) cells were produced. The latter resemble L-DCs, which are also produced in murine spleen cocultures. Both subsets expressed hCD80 and hCD86, which identifies them as antigen-presenting cells, particularly DCs, and were highly endocytic. It is noteworthy that murine splenic stroma can serve as a support matrix for human hematopoiesis and DC production. These results support the hypothesis that 5G3 must express both cell-associated and soluble factors that can signal hematopoiesis in human and murine progenitors. PMID:25891217

  5. Spontaneous release of interleukin 2 by lung T lymphocytes in active pulmonary sarcoidosis is primarily from the Leu3+DR+ T cell subset.

    PubMed Central

    Saltini, C; Spurzem, J R; Lee, J J; Pinkston, P; Crystal, R G

    1986-01-01

    The inflammation within the lower respiratory tract of individuals with pulmonary sarcoidosis is dominated by large numbers of helper T lymphocytes that proliferate and spontaneously release interleukin 2 (IL-2). To identify the lymphocyte subpopulation that releases IL-2 in this disorder, lung lymphocytes recovered by bronchoalveolar lavage were characterized using the monoclonal antibodies Leu4 (T lymphocyte), Leu3 (helper/inducer), Leu2 (suppressor/cytotoxic), and anti-HLA-DR, and separated by panning and flow cytometry. The majority of the IL-2 spontaneously released by T cells in the sarcoid lung was contributed by the Leu3+ cell population (Leu3+65 +/- 23 IL-2 units released/10(6) cells per 24 h; Leu2+ 9 +/- 8, P less than 0.04). Further characterization of the lung Leu3+ T cells in sarcoid demonstrated that 30 +/- 3% were expressing HLA-DR molecules on their surface compared with 6 +/- 1% in normals (P less than 0.01). Importantly, the subpopulation of Leu3+ lung T lymphocytes expressing a high intensity of HLA-DR molecules on their surface was responsible for the majority of the release of IL-2 in the sarcoid lung (Leu3+ high-intensity DR 42 +/- 17 U/10(6) cells per 24 h, Leu3+ low-intensity DR 8 +/- 1 U/10(6) cells per 24 h; P less than 0.01). Thus, the spontaneous release of IL-2 in the lung of sarcoid patients appears to be localized to a subset of Leu3+ high-intensity DR ("activated" lung helper/inducer) T lymphocytes. Because the sarcoid lung is characterized by markedly increased numbers of these cells, it is likely that this compartmentalized T cell population plays a major role in sustaining the exaggerated localized immune processes of this disorder. PMID:3486888

  6. Comparison of gene expression profiles between human and mouse monocyte subsets

    PubMed Central

    Ingersoll, Molly A.; Spanbroek, Rainer; Lottaz, Claudio; Gautier, Emmanuel L.; Frankenberger, Marion; Hoffmann, Reinhard; Lang, Roland; Haniffa, Muzlifah; Collin, Matthew; Tacke, Frank; Habenicht, Andreas J. R.

    2010-01-01

    Blood of both humans and mice contains 2 main monocyte subsets. Here, we investigated the extent of their similarity using a microarray approach. Approximately 270 genes in humans and 550 genes in mice were differentially expressed between subsets by 2-fold or more. More than 130 of these gene expression differences were conserved between mouse and human monocyte subsets. We confirmed numerous of these differences at the cell surface protein level. Despite overall conservation, some molecules were conversely expressed between the 2 species' subsets, including CD36, CD9, and TREM-1. Other differences included a prominent peroxisome proliferator-activated receptor γ (PPARγ) signature in mouse monocytes, which is absent in humans, and strikingly opposed patterns of receptors involved in uptake of apoptotic cells and other phagocytic cargo between human and mouse monocyte subsets. Thus, whereas human and mouse monocyte subsets are far more broadly conserved than currently recognized, important differences between the species deserve consideration when models of human disease are studied in mice. PMID:19965649

  7. T lymphocytes subsets and cytokine pattern induced by vaccination against bovine brucellosis employing S19 calfhood vaccination and adult RB51 revaccination.

    PubMed

    Dorneles, Elaine M S; Teixeira-Carvalho, Andréa; Araújo, Márcio S S; Lima, Graciela Kunrath; Martins-Filho, Olindo A; Sriranganathan, Nammalwar; Lage, Andrey P

    2014-10-21

    The aims of this study were to address the protective immune response induced by S19 vaccination (n=10) and RB51 revaccination, in pregnant (n=9) and non-pregnant (n=10) S19 calfhood-vaccinated cattle as follows: evaluate the in vitro CD4(+) and CD8(+) T-lymphocytes specific proliferation, and in vitro expression of IFN-γ by CD4(+) and CD8(+) T-cells and IL-4 by CD4(+), CD8(+) and CD21(+) lymphocytes subset. Upon in vitro stimulation with γ-irradiated Brucella abortus 2308, blood mononuclear cells from S19 vaccinated and RB51 revaccinated cows exhibited significantly higher proliferation of CD4(+) and CD8(+) T-lymphocytes and CD4(+)IFN-γ(+) T-cells compared to non-vaccinated animals. RB51 revaccination, regardless of the pregnancy status, did not enhance the proliferation of CD4(+) or CD8(+) T-cells nor IFN-γ or IL-4 production. Data from the present study suggest that cattle's cellular immune response induced after brucellosis vaccination and revaccination is due to CD4(+) and CD8(+) T-lymphocytes, being CD4(+) T-cells the main source of IFN-γ. PMID:25218192

  8. Shen-Qi-Jie-Yu-Fang has antidepressant effects in a rodent model of postpartum depression by regulating the immune organs and subsets of T lymphocytes

    PubMed Central

    Qu, Miao; Tang, Qisheng; Li, Xiaoli; Zhao, Ruizhen; Li, Jingya; Xu, Hong; Gao, Yushan; Mao, Yingqiu

    2015-01-01

    Background Shen-Qi-Jie-Yu-Fang (SJ Fang) is a herbal preparation used in traditional Chinese medicine, and is a potentially important new therapeutic agent in postpartum depression (PPD). Previously, we have elucidated the effects of SJ Fang on hormone receptors and monoamine neurotransmitters involved in the hypothalamic-pituitary-adrenal and hypothalamic-pituitary-gonadal axes in PPD rats. However, the immune-modulating effects of SJ Fang in PPD are still unknown. In this study, we explored the effects of SJ Fang on the immune organs and subsets of T lymphocytes in PPD rats. Methods PPD was created in Sprague-Dawley rats by inducing hormone-simulated pregnancy followed by hormone withdrawal. After hormone withdrawal, the PPD rats were then treated with fluoxetine at 1, 2, and 4 weeks, and the SJ Fang rats were also treated at 1, 2, and 4 weeks. Depressive behavior in the rats was evaluated by the forced swim test, sucrose consumption test, and open field test. The thymus index and spleen index were calculated. Hematoxylin-eosin staining was used to identify pathological features in the thymus and spleen. CD3, CD4, and CD8 lymphocyte subsets were analyzed by flow cytometry. Results Both fluoxetine and SJ Fang decreased immobility time, increased sucrose consumption, an horizontal and vertical movement. After 4 weeks of treatment with fluoxetine or SJ Fang, the thymus index and spleen index were significantly higher than at baseline, and the morphology of the thymus and spleen were returning to normal. Two weeks after hormone withdrawal, subsets of T lymphocytes indicated a shift from immune activation to immune suppression, which was reversed by 4 weeks of treatment with fluoxetine or SJ Fang. Conclusion It is suggested that T-cell mediate immune responses which may play a role in the etiopathology of postpartum depression. SJ Fang had an antidepressant effect on the immune system in rats with PPD. PMID:26203247

  9. Circulating and in situ lymphocyte subsets and Langerhans cells in patients with compositae oleoresin dermatitis and increased ultraviolet A sensitivity during treatment with azathioprine

    SciTech Connect

    Baadsgaard, O.

    1986-04-01

    Circulating and in situ lymphocyte subsets and Langerhans cells in four patients with compositae oleoresin dermatitis and increased ultraviolet A sensitivity before and during treatment with azathioprine were estimated. It was found that the number of Leu 6+ Langerhans cells decreased during therapy. This decrease was accompanied by a reduction in the number of Leu 2a+, Leu 3a+, Leu 4+, DR+, and Leu M2+ cells in the blood and a reduction in the number of Leu 2a+, Leu 3a+, Leu 4+, and DR+ cells in the skin. Concomitantly with the changes in the number of immunocompetent cells, the eczema cleared.

  10. Human malignant melanoma-derived progestagen-associated endometrial protein immunosuppresses T lymphocytes in vitro.

    PubMed

    Ren, Suping; Chai, Lina; Wang, Chunyan; Li, Changlan; Ren, Qiquan; Yang, Lihua; Wang, Fumei; Qiao, Zhixin; Li, Weijing; He, Min; Riker, Adam I; Han, Ying; Yu, Qun

    2015-01-01

    Progestagen-associated endometrial protein (PAEP) is a glycoprotein of the lipocalin family that acts as a negative regulator of T cell receptor-mediated activation. However, the function of tumor-derived PAEP on the human immune system in the tumor microenvironment is unknown. PAEP is highly expressed in intermediate and thick primary melanomas (Breslow's 2.5mm or greater) and metastatic melanomas, correlating with its expression in daughter cell lines established in vitro. The current study investigates the role of melanoma cell-secreted PAEP protein in regulating T cell function. Upon the enrichment of CD3+, CD4+ and CD8+ T cells from human peripheral blood mononuclear cells, each subset was then mixed with either melanoma-derived PAEP protein or PAEP-poor supernatant of gene-silenced tumor cells. IL-2 and IFN-γ secretion of CD4+ T cells significantly decreased with the addition of PAEP-rich supernatant. And the addition of PAEP-positive cell supernatant to activated lymphocytes significantly inhibited lymphocyte proliferation and cytotoxic T cell activity, while increasing lymphocyte apoptosis. Our result suggests that melanoma cell-secreted PAEP protein immunosuppresses the activation, proliferation and cytotoxicity of T lymphocytes, which might partially explain the mechanism of immune tolerance induced by melanoma cells within the tumor microenvironment. PMID:25785839

  11. Association between gene and miRNA expression profiles and stereotyped subset #4 B-cell receptor in chronic lymphocytic leukemia.

    PubMed

    Maura, Francesco; Cutrona, Giovanna; Mosca, Laura; Matis, Serena; Lionetti, Marta; Fabris, Sonia; Agnelli, Luca; Colombo, Monica; Massucco, Carlotta; Ferracin, Manuela; Zagatti, Barbara; Reverberi, Daniele; Gentile, Massimo; Recchia, Anna Grazia; Bossio, Sabrina; Rossi, Davide; Gaidano, Gianluca; Molica, Stefano; Cortelezzi, Agostino; Di Raimondo, Francesco; Negrini, Massimo; Tassone, Pierfrancesco; Morabito, Fortunato; Ferrarini, Manlio; Neri, Antonino

    2015-01-01

    In this study we investigated specific biological and clinical features associated with chronic lymphocytic leukemia (CLL) patients carrying stereotyped BCR subset #4 (IGHV4-34) among a prospective cohort of 462 CLL/MBL patients in early stage (Binet A). All subset #4 patients (n = 16) were characterized by the IGHV mutated gene configuration, and absence of unfavorable cytogenetic lesions, NOTCH1 or SF3B1 mutations. Gene and miRNA expression profiling evidenced that the leukemic cells of subset #4 cases showed significant downregulation of WDFY4, MF2A and upregulation of PDGFA, FGFR1 and TFEC gene transcripts, as well as the upregulation of miR-497 and miR-29c. The transfection of miR-497 mimic in primary leukemic CLL cells induced a downregulation of BCL2, a known validated target of this miRNA. Our data identify biological characteristics associated with subset #4 patients, providing further evidence for the putative role of BCR in shaping the features of the tumor cells in CLL. PMID:25860243

  12. Tumor-derived exosomes regulate expression of immune function-related genes in human T cell subsets

    PubMed Central

    Muller, Laurent; Mitsuhashi, Masato; Simms, Patricia; Gooding, William E.; Whiteside, Theresa L.

    2016-01-01

    Tumor cell-derived exosomes (TEX) suppress functions of immune cells. Here, changes in the gene profiles of primary human T lymphocytes exposed in vitro to exosomes were evaluated. CD4+ Tconv, CD8+ T or CD4+ CD39+ Treg were isolated from normal donors’ peripheral blood and co-incubated with TEX or exosomes isolated from supernatants of cultured dendritic cells (DEX). Expression levels of 24–27 immune response-related genes in these T cells were quantified by qRT-PCR. In activated T cells, TEX and DEX up-regulated mRNA expression levels of multiple genes. Multifactorial data analysis of ΔCt values identified T cell activation and the immune cell type, but not exosome source, as factors regulating gene expression by exosomes. Treg were more sensitive to TEX-mediated effects than other T cell subsets. In Treg, TEX-mediated down-regulation of genes regulating the adenosine pathway translated into high expression of CD39 and increased adenosine production. TEX also induced up-regulation of inhibitory genes in CD4+ Tconv, which translated into a loss of CD69 on their surface and a functional decline. Exosomes are not internalized by T cells, but signals they carry and deliver to cell surface receptors modulate gene expression and functions of human T lymphocytes. PMID:26842680

  13. Interaction of nanosilver particles with human lymphocyte cells

    NASA Astrophysics Data System (ADS)

    Zhornik, Alena; Baranova, Ludmila; Volotovski, Igor; Chizhik, Sergey; Drozd, Elizaveta; Sudas, Margarita; Buu Ngo, Quoc; Chau Nguyen, Hoai; Huynh, Thi Ha; Hien Dao, Trong

    2015-01-01

    The damaging effects of nanoparticles were hypothesized to be the oxidative stress caused by the formation of reactive oxygen species and initiation of inflammatory reactions. In this context a study on the effects of nanosilver particles on the formation of reactive oxygen species in human lymphocyte culture was carried out. The obtained results showed that fluorescence intensity considerably increased after cells had interacted with nanosilver particles of varying concentrations, indicating the formation of reactive oxygen species and their accumulation in lymphocyte cells. Morphological study of the lymphocyte cells under the effects of nanosilver particles showed that the change in morphology depends on the concentration and size of nanosilver particles: for a size ≤20 nm the lymphocyte cell significantly shrank with pronounced differences in the morphological structure of the cell membrane, but for a size ≥200 nm no change was observed.

  14. Human gamma interferon production by cytotoxic T lymphocytes sensitized during hepatitis A virus infection

    SciTech Connect

    Maier, K.; Gabriel, P.; Koscielniak, E.; Stierhof, Y.D.; Wiedmann, K.H.; Flehmig, B.; Vallbracht, A.

    1988-10-01

    The production of interferon (IFN) during a chromium-51 release assay with hepatitis A virus (HAV)-infected fibroblasts and autologous peripheral blood lymphocytes from patients with acute HAV infection was studied to determine whether IFN plays a role in immunopathogenesis of hepatitis A infection in humans. Skin fibroblasts of eight patients after acute HAV infection and from two control persons without history of current of past HAV infection were infected with HAV. Peripheral blood lymphocytes were collected at different times after the onset of icterus and tested in a chromium-51 release assay against autologous HAV-infected skin fibroblasts for their cytolytic and IFN-producing activity. The IFN produced during the assay was characterized and found to have the properties of human gamma IFN. Cytotoxicity and gamma IFN release were virus specific. The cell types responsible for both functions were characterized and found to be in the HLA-dependent T8/sup +/ lymphocyte subset. Considering that gamma IFN has an antiviral effect on persistent HAV infection in vitro and that it probably accounts for stimulation of HLA class I antigen expression on hepatocytes, these experimental results presented here demonstrate that human gamma IFN produced by HAV-specific T cells may participate in pathogenesis of hepatitis A infection in humans.

  15. Effect of controlled ozone exposure on human lymphocyte function

    SciTech Connect

    Peterson, M.L.; Smialowicz, R.; Harder, S.; Ketcham, B.; House, D.

    1981-04-01

    The effects of ozone (O/sub 3/) on cell-mediated immunity were studied in 16 human subjects exposed to 1176 ..mu..g/m/sup 3/ O/sub 3/ (0.6 ppM) for 2 h in an environmentally controlled exposure chamber. Venous blood samples were taken before and immediately after controlled air and O/sub 3/ exposures, as well as at 72 h, 2 and 4 weeks, and at one random time at least 1 month after treatment. The relative frequency of T lymphocytes in blood and the in vitro blastogenic response of lymphocytes to phytohemagglutinin (PHA), concanavalin A (Con A), pokeweed mitogen (PWM), and Candida albicans were determined. During the course of the experiment, no statistically significant changes were observed in the number of T lymphocytes that form spontaneous rosettes with sheep erythrocytes. The response of T lymphocytes to PHA was significantly reduced (P < 0.05) in samples taken at 2 and 4 weeks, following O/sub 3/ exposure. Normal response to PHA was observed at 2 months post-O/sub 3/ exposure. No statistically significant changes in lymphocyte responses to Con A, PWM, or Candida were seen. These results show that one 2 h exposure of humans to 0.6 ppM O/sub 3/ may lead to a transient suppression of the PHA-stimulated blastogenic transformation of peripheral blood lymphocytes. The data indicate that the blastogenic response to PHA of human lymphocytes is exquisitely sensitive to O/sub 3/ exposure and could serve as a bioassay for evaluating subtle changes in cellular immunity induced by O/sub 3/ and possibly other pollutants.

  16. T Lymphocyte Density and Distribution in Human Colorectal Mucosa, and Inefficiency of Current Cell Isolation Protocols

    PubMed Central

    Preza, Gloria Cuevas; Yang, Otto O.; Elliott, Julie; Anton, Peter A.; Ochoa, Maria T.

    2015-01-01

    Mucosal tissues are critical immune effector sites containing complex populations of leukocytes in a tissue microenvironment that remains incompletely understood. We identify and quantify in human distal colorectal tissue absolute mucosal CD3+ lymphocytes, including CD4+ and CD8+ subsets, by direct visualization using immunohistochemistry (IHC), immunofluorescence (IF), and an automated counting protocol (r2=0.90). Sigmoid and rectal mucosal tissues are both densely packed with T lymphocytes in the mucosal compartment. Both compartments had similar densities of CD3+ T lymphocytes with 37,400 ± 2,801 cells/mm3 and 33,700 ± 4,324 cell/mm3, respectively. Sigmoid mucosa contained 57% CD3+CD4+ and 40% CD3+CD8+ T lymphocytes which calculates to 21,300 ± 1,476/mm3 and 15,000 ± 275/mm3 T lymphocytes, respectively. Rectal mucosa had 57% CD3+CD4+ and 42% CD3+CD8+ or 21,577 ± 332, and 17,090 ± 1,206 cells/mm3, respectively. By comparison, sigmoid mucosal biopsies subjected to conventional collagenase digestion, mononuclear cell (MMC) isolation and staining for flow cytometry yielded 4,549 ± 381/mm3 and 2,708 ± 245/mm3 CD4+ and CD8+ T lymphocytes. These data suggest only ~20.7% recovery compared to IHC results for these markers. Further studies will determine if this reflects a selective bias in only CD3+, CD4+ and CD8+ T cells or can be generalized to all flow-analyzed cells from mucosal tissues for phenotyping and functional testing. PMID:25856343

  17. Experimental allergic encephalomyelitis in rhesus monkeys: II. Treatment of EAE with anti-T lymphocyte subset monoclonal antibodies.

    PubMed Central

    Van Lambalgen, R; Jonker, M

    1987-01-01

    Experimental allergic encephalomyelitis was induced in rhesus monkeys by immunization with bovine brain homogenate emulsified in complete Freund's adjuvant. Four monkeys were treated with anti-CD4 (OKT4+4A) monoclonal antibodies after the onset of clinical signs. One monkey developed a chronic-progressive course of EAE and was killed after a significantly prolonged disease of 19 days. One monkey had a relapse and survived with stable neurological signs. Two monkeys fully recovered. OKT4+4A treatment resulted in a short-term clearance of CD4+ lymphocytes and a reduction in granulocytes. Granulocytes may be attracted to the brain by chemotactic factors produced by CD4+ lymphocytes and are responsible for the development of the lethal granulocytic lesions. Clearance of CD4+ lymphocytes successfully prevented granulocytes from migrating to the brain. Nuclear magnetic resonance imaging showed extensive lesions during an acute attack, but these lesions became undetectable when the monkeys recovered. These results indicate that treatment with OKT4+4A can successfully reverse clinical signs of EAE. Four untreated monkeys and one monkey treated with OKT8F died of acute EAE within 3 days of the onset of clinical signs. Images Fig. 5 PMID:3498570

  18. Human blood dendritic cell subsets exhibit discriminative pattern recognition receptor profiles

    PubMed Central

    Lundberg, Kristina; Rydnert, Frida; Greiff, Lennart; Lindstedt, Malin

    2014-01-01

    Dendritic cells (DCs) operate as the link between innate and adaptive immunity. Their expression of pattern recognition receptors (PRRs), such as Toll-like receptors (TLRs) and C-type lectin receptors (CLRs), enables antigen recognition and mediates appropriate immune responses. Distinct subsets of human DCs have been identified; however their expression of PRRs is not fully clarified. Expressions of CLRs by DC subpopulations, in particular, remain elusive. This study aimed to identify and compare PRR expressions on human blood DC subsets, including CD1c+, CD141+ and CD16+ myeloid DCs and CD123+ plasmacytoid DCs, in order to understand their capacity to recognize different antigens as well as their responsiveness to PRR-directed targeting. Whole blood was obtained from 13 allergic and six non-allergic individuals. Mononuclear cells were purified and multi-colour flow cytometry was used to assess the expression of 10 CLRs and two TLRs on distinct DC subsets. PRR expression levels were shown to differ between DC subsets for each PRR assessed. Furthermore, principal component analysis and random forest test demonstrated that the PRR profiles were discriminative between DC subsets. Interestingly, CLEC9A was expressed at lower levels by CD141+ DCs from allergic compared with non-allergic donors. The subset-specific PRR expression profiles suggests individual responsiveness to PRR-targeting and supports functional specialization. PMID:24444310

  19. Human blood dendritic cell subsets exhibit discriminative pattern recognition receptor profiles.

    PubMed

    Lundberg, Kristina; Rydnert, Frida; Greiff, Lennart; Lindstedt, Malin

    2014-06-01

    Dendritic cells (DCs) operate as the link between innate and adaptive immunity. Their expression of pattern recognition receptors (PRRs), such as Toll-like receptors (TLRs) and C-type lectin receptors (CLRs), enables antigen recognition and mediates appropriate immune responses. Distinct subsets of human DCs have been identified; however their expression of PRRs is not fully clarified. Expressions of CLRs by DC subpopulations, in particular, remain elusive. This study aimed to identify and compare PRR expressions on human blood DC subsets, including CD1c(+) , CD141(+) and CD16(+) myeloid DCs and CD123(+) plasmacytoid DCs, in order to understand their capacity to recognize different antigens as well as their responsiveness to PRR-directed targeting. Whole blood was obtained from 13 allergic and six non-allergic individuals. Mononuclear cells were purified and multi-colour flow cytometry was used to assess the expression of 10 CLRs and two TLRs on distinct DC subsets. PRR expression levels were shown to differ between DC subsets for each PRR assessed. Furthermore, principal component analysis and random forest test demonstrated that the PRR profiles were discriminative between DC subsets. Interestingly, CLEC9A was expressed at lower levels by CD141(+) DCs from allergic compared with non-allergic donors. The subset-specific PRR expression profiles suggests individual responsiveness to PRR-targeting and supports functional specialization. PMID:24444310

  20. Effects of Beryllium on Human Serum Immunoglobulin and Lymphocyte Subpopulation

    PubMed Central

    Kim, DaeSeong; Won, Yong Lim; Kang, Seong-Kyu

    2013-01-01

    To investigate the effects of short-term exposure of beryllium on the human immune system, the proportion of T-lymphocytes such as CD3+, CD4+, CD8+, CD95, and NK cells, andthe proportion of B cells and TNFα level in peripheral blood and immunoglobulins in the serum of 43 exposed workers and 34 healthy control subjects were studied. External exposure to beryllium was measured by atomic absorption spectrometer as recommended by the NIOSH analytical method 7300. T lymphocyte subpopulation analysis was carried out with flow cytometer. The working duration of exposed workers was less than 3 months and the mean ambient beryllium level was 3.4 μg/m3, 112.3 μg/m3, and 2.3 μg/m3 in molding (furnace), deforming (grinding), and sorting processes, respectively (cited from Kim et al., 2008). However, ambient beryllium level after process change was non-detectable (< 0.1 μg/m3). The number of T lymphocytes and the amount of immunoglobulins in the beryllium-exposed workers and control subjects were not significantly different, except for the total number of lymphocytes and CD95 (APO1/FAS). The total number of lymphocytes was higher in the beryllium-exposed individuals than in the healthy control subjects. Multiple logistic regression analysis showed lymphocytes to be affected by beryllium exposure (odd ratio = 7.293; p < 0.001). These results show that short-term exposure to beryllium does not induce immune dysfunction but is probably associated with lymphocytes proliferation. PMID:24278637

  1. Rabies virus infects mouse and human lymphocytes and induces apoptosis.

    PubMed Central

    Thoulouze, M I; Lafage, M; Montano-Hirose, J A; Lafon, M

    1997-01-01

    Attenuated and highly neurovirulent rabies virus strains have distinct cellular tropisms. Highly neurovirulent strains such as the challenge virus standard (CVS) are highly neurotropic, whereas the attenuated strain ERA also infects nonneuronal cells. We report that both rabies virus strains infect activated murine lymphocytes and the human lymphoblastoid Jurkat T-cell line in vitro. The lymphocytes are more permissive to the attenuated ERA rabies virus strain than to the CVS strain in both cases. We also report that in contrast to that of the CVS strain, ERA viral replication induces apoptosis of infected Jurkat T cells, and cell death is concomitant with viral glycoprotein expression, suggesting that this protein has a role in the induction of apoptosis. Our data indicate that (i) rabies virus infects lymphocytes, (ii) lymphocyte infection with the attenuated rabies virus strain causes apoptosis, and (iii) apoptosis does not hinder rabies virus production. In contrast to CVS, ERA rabies virus and other attenuated rabies virus vaccines stimulate a strong immune response and are efficient live vaccines. The paradoxical finding that a rabies virus triggers a strong immune response despite the fact that it infects lymphocytes and induces apoptosis is discussed in terms of the function of apoptosis in the immune response. PMID:9311815

  2. Analysis of purine metabolic enzymes in human CD4 Leu 8- and CD4 Leu 8+ lymphocyte subpopulations.

    PubMed

    Fernandez-Mejia, C; Polmar, S H; Peralta-Zaragoza, O; Madrid-Marina, V

    1993-02-01

    1. Specific activities of adenosine deaminase, purine nucleoside phosphorylase, adenosine kinase, 5'-nucleotidase, S-adenosyl-L-homocysteine hydrolase, AMP deaminase, adenine phosphoribosyl transferase, and hypoxanthine phosphoribosyl transferase were analyzed in human CD4 T-lymphocyte subsets. 2. CD4 Leu 8- (helper/inducer) and CD4 Leu 8+ (suppressor/inducer) subpopulations were obtained by panning or fluorescence activated cell sorting techniques using specific monoclonal antibodies. 3. A 45% decrease of 5'-NT AMP activity in the CD4 Leu 8- cells (suppressor/inducer) compared with CD4 total cell population. 4. No statistical significant differences in enzyme activity were found between the subsets analyzed in other purine enzymes. 5. These results suggest that the distribution of purine metabolic enzymes is homogeneous in CD4 Leu 8- and CD4 Leu 8+ T-lymphocyte subpopulations. PMID:8444317

  3. Altered Cytokine Production By Specific Human Peripheral Blood Cell Subsets Immediately Following Spaceflight

    NASA Technical Reports Server (NTRS)

    Crucian, Brian E.; Cubbage, Michael L.; Sams, Clarence F.

    1999-01-01

    In this study, we have attempted to combine standard immunological assays with the cellular resolving power of the flow cytometer to positively identify the specific cell types involved in spaceflight-induced immune alterations. We have obtained whole blood samples from 27 astronauts collected at three timepoints (L-10, R+0 and R+3) surrounding four recent space shuttle missions. The duration of these missions ranged from 10 to 18 days. Assays performed included serum/urine cortisol, comprehensive subset phenotyping, assessment of cellular activation markers and intracellular cytokine production following mitogenic stimulation. Absolute levels of peripheral granulocytes were significantly elevated following spaceflight, but the levels of circulating lymphocytes and monocytes were unchanged. Lymphocyte subset analysis demonstrated trends towards a decreased percentage of T cells and an increased percentage of B cells. Nearly all of the astronauts exhibited an increased CD4:CD8 ratio, which was dramatic in some individuals. Assessment of memory (CD45RA+) vs. naive (CD45RO+) CD4+ T cell subsets was more ambiguous, with subjects tending to group more as a flight crew. All subjects from one mission demonstrated an increased CD45RA:CD45RO ratio, while all subjects from another Mission demonstrated a decreased ratio. While no significant trend was seen in the monocyte population as defined by scatter, a decreased percentage of the CD14+ CD16+ monocyte subset was seen following spaceflight in all subjects tested. In general, most of the cellular changes described above which were assessed at R+O and compared to L-10 trended to pre-flight levels by R+3. Although no significant differences were seen in the expression of the cellular activation markers CD69 and CD25 following exposure to microgravity, significant alterations were seen in cytokine production in response to mitogenic activation for specific subsets. T cell (CD3+) production of IL-2 was significantly decreased

  4. Influence of ethanol on human T-lymphocyte migration

    SciTech Connect

    Kaelin, R.M.; Semerjian, A.; Center, D.M.; Bernardo, J.

    1984-11-01

    Because ethanol consumption is associated with increased susceptibility to infection, an examination was made of the effects of ethanol and its metabolite acetaldehyde on human T-lymphocyte migration, an important functional component of cellular inflammatory responses. With a modified Boyden chamber system, ethanol at 0.25% and 0.50% (vol/vol) inhibited spontaneous motility of human T-lymphocytes, in a noncytotoxic manner, to 65% +/- 7% (mean +/- SEM) and 62% +/- 7% of control values of migration, respectively. When T-lymphocyte migration was stimulated by colchicine (10/sup -5/ mol/L), incubation with ethanol (0.25% and 0.50%, vol/vol) decreased migration to 80% +/- 4% and 66% +/- 8% of control values, respectively. Similar degrees of inhibition of migration were obtained with acetaldehyde at concentrations five to 10 times less than ethanol. Ethanol was similarly capable of inhibiting T cell migration induced by dibutyryl cyclic guanosine monophosphate, but it had no effect on stimulated migration induced by a human chemokinetic lymphokine. The study demonstrates that ethanol, at concentrations achievable in vivo, is capable of depressing T-lymphocyte migration. This effect might contribute to the immunosuppression associated with ethanol consumption. 36 references, 4 figures.

  5. Immunomodulating activity of seaweed extract on human lymphocytes in vitro.

    PubMed

    Shan, B E; Yoshida, Y; Kuroda, E; Yamashita, U

    1999-01-01

    Effect of eight kinds of seaweed extract (SWE) on human lymphocytes was studied in vitro. The extracts of Hizikiafusiformis and Meristotheca papulosa (green) markedly stimulated human lymphocytes to proliferate, whereas Eucheuma muricatum and Meristotheca papulosa (red) weakly stimulated proliferation. The responder cells are T cells, because T cells purified by sheep red blood cell (SRBC) rosette-formation were significantly stimulated with SWE, but B cells were not. These extracts enhanced the induction of cytotoxic T lymphocyte (CTL) activity, but failed to enhance natural killer (NK) cell activity. These extracts had a stimulatory effect on immunoglobulin (Ig) production by B cells and tumor necrosis factor (TNF) production by monocytes. The activity of Hizikia fusiformis associated with polysaccharides which were extracted with ethanol and purified by ion-exchange and gel filtration chromatography, whose molecular weight was about 100 kDa. These results suggest that SWE has an immunomodulating activity on human lymphocytes and this ability might be useful for clinical application to treat several diseases such as tumors. PMID:10411282

  6. The role of a human antigen specific T8+ cell subset in antigen presentation, helper function and contrasuppression.

    PubMed Central

    Lehner, T; Avery, J; Jones, T

    1985-01-01

    Regulation of the human immune response was studied by sequential separation of subsets of T cells, followed by assessment of their helper and suppressor functions in a series of reconstitution experiments. T8+ lymphocytes were separated by panning on streptococcal antigen (SA) coated plates into T8+ SA-adherent cells (T8+SA+) and T8+ SA-non-adherent cells (T8+SA-). The helper and suppressor functions of the T8+SA+ and T8+SA- cells, reconstituted with T4+ helper cells were then studied by a direct antibody forming cell assay. T4+ cells will not induce helper activity by 1000 ng SA alone but require the accessory function of monocytes (Mo). However, replacing Mo by T8+SA+ cells will elicit a similar helper activity by T4+ cells and SA as that induced by Mo. In addition to the antigen-specific presentation and induction of helper activity, the T8+SA+ subset displays the properties of antigen-specific contrasuppressor cells. Thus, reconstitution of T4+ cells and T8+SA- (suppressor cells) with T8+SA+ and 1000 ng SA induces helper and no suppressor activity. Substitution of Mo for the T8+SA+ cells converts the helper to a predominantly suppressor-cell function. T8+SA- cells elicit suppression with 1 ng SA in the absence of accessory cells and reconstitution with Mo, T8+SA+ or T4+ cells failed to affect the suppressor activity. Total reconstitution of the four principle subsets of T4+, T8+SA+, T8+SA- cells and Mo elicited similar antigen dose-dependent responses as those of the unseparated mononuclear cells. It seems that all four cell subsets are required for optimal immunoregulation. We suggest that the T8+SA+ can present antigen to T4+ helper cells and induce helper activity, but in addition these cells can prevent the suppressor subset of T8+ cells from inhibiting T4+ helper cells and function as contrasuppressor cells. The mechanism of these functions is not known but HLA class II antigens might play an essential role in antigen binding, presentation and

  7. MicroRNA expression profiling of human blood monocyte subsets highlights functional differences

    PubMed Central

    Dang, Truong-Minh; Wong, Wing-Cheong; Ong, Siew-Min; Li, Peng; Lum, Josephine; Chen, Jinmiao; Poidinger, Michael; Zolezzi, Francesca; Wong, Siew-Cheng

    2015-01-01

    Within human blood there are two subsets of monocytes that can be identified by differential expression of CD16. Although numerous phenotypic and functional differences between the subsets have been described, little is known of the mechanisms underlying the distinctive properties of the two subsets. MicroRNAs (miRNAs) are small non-coding RNAs that can regulate gene expression through promoting mRNA degradation or repressing translation, leading to alterations in cellular processes. Their potential influence on the functions of monocyte subsets has not been investigated. In this study, we employed microarray analysis to define the miRNA expression profile of human monocyte subsets. We identified 66 miRNAs that were differentially expressed (DE) between CD16+ and CD16− monocytes. Gene ontology analysis revealed that the predicted targets of the DE miRNAs were predominantly associated with cell death and cellular movement. We validated the functional impacts of selected DE miRNAs in CD16− monocytes, over-expression of miR-432 significantly increases apoptosis, and inhibiting miR-19a significantly reduces cell motility. Furthermore, we found that miR-345, another DE miRNA directly targets the transcription factor RelA in monocytes, which resulted in the differential expression of RelA in monocyte subsets. This implicates miR-345 indirect regulation of many genes downstream of RelA, including important inflammatory mediators. Together, our data show that DE miRNAs could contribute substantially to regulating the functions of human blood monocytes. PMID:25707426

  8. Purine nucleoside modulation of functions of human lymphocytes.

    PubMed

    Priebe, T; Platsoucas, C D; Seki, H; Fox, F E; Nelson, J A

    1990-09-01

    The accumulation of endogenous substrates in patients with adenosine deaminase deficiency or purine nucleoside phosphorylase deficiency is believed to be responsible for the immunodeficiency observed in these patients. To identify the lymphocyte populations that are most susceptible to these substrates, we investigated the effect of their nucleoside analogs on a number of T and B cell functions of human lymphocytes. We found that tubercidin (Tub), 2-chloro 2'deoxyadenosine (2CldA), 2-fluoro adenine arabinoside-5'phosphate (FaraAMP), and 9-beta-D-arabinosyl guanine (AraGua) inhibited the proliferative responses of human peripheral blood mononuclear cells (PBMC) to polyclonal activators (PHA, OKT3 mab) or to allogeneic PBMC in mixed lymphocyte cultures (MLC). Addition of recombinant IL-2 from the beginning of the culture did not alter the inhibition by Tub of the proliferative responses of PBMC. These purine nucleoside analogs also inhibited the proliferative responses of purified human peripheral blood CD4+ and CD8+ T cells to PHA and of purified B cells to SAC. The concentrations of these nucleosides required to achieve a given degree of inhibition of proliferative responses of T lymphocyte subpopulations or B cells was similar, suggesting that these analogs do not exhibit any selectivity for these purified lymphocyte populations. Tub and FaraAMP, respectively, inhibited and enhanced, at the effector phase, both NK cytotoxicity and specific T cell-mediated cytotoxicity. In contrast to these findings, LAK cytotoxicity at the effector phase was not significantly inhibited by Tub, and was not enhanced by FaraAMP. Both analogs inhibited rIL-2-induced proliferative responses of PBMC, but did not affect the generation of LAK cytotoxicity (induction phase) against the K562 targets when added at the beginning of the culture. This suggests that DNA synthesis is not required for LAK cell induction. Both Tub and FaraAMP inhibited immunoglobulin production (IgG and IgM) by

  9. De novo transcriptome profiling of highly purified human lymphocytes primary cells

    PubMed Central

    Bonnal, Raoul J.P.; Ranzani, Valeria; Arrigoni, Alberto; Curti, Serena; Panzeri, Ilaria; Gruarin, Paola; Abrignani, Sergio; Rossetti, Grazisa; Pagani, Massimiliano

    2015-01-01

    To help better understand the role of long noncoding RNAs in the human immune system, we recently generated a comprehensive RNA-seq data set using 63 RNA samples from 13 subsets of T (CD4+ naive, CD4+ TH1, CD4+ TH2, CD4+ TH17, CD4+ Treg, CD4+ TCM, CD4+ TEM, CD8+ TCM, CD8+ TEM, CD8+ naive) and B (B naive, B memory, B CD5+) lymphocytes. There were five biological replicates for each subset except for CD8+ TCM and B CD5+ populations that included 4 replicates. RNA-Seq data were generated by an Illumina HiScanSQ sequencer using the TruSeq v3 Cluster kit. 2.192 billion of paired-ends reads, 2×100 bp, were sequenced and after filtering a total of about 1.7 billion reads were mapped. Using different de novo transcriptome reconstruction techniques over 500 previously unknown lincRNAs were identified. The current data set could be exploited to drive the functional characterization of lincRNAs, identify novel genes and regulatory networks associated with specific cells subsets of the human immune system. PMID:26451251

  10. Rapid activation of spleen dendritic cell subsets following lymphocytic choriomeningitis virus infection of mice: analysis of the involvement of type 1 IFN.

    PubMed

    Montoya, Maria; Edwards, Matthew J; Reid, Delyth M; Borrow, Persephone

    2005-02-15

    In this study, we report the dynamic changes in activation and functions that occur in spleen dendritic cell (sDC) subsets following infection of mice with a natural murine pathogen, lymphocytic choriomeningitis virus (LCMV). Within 24 h postinfection (pi), sDCs acquired the ability to stimulate naive LCMV-specific CD8+ T cells ex vivo. Conventional (CD11chigh CD8+ and CD4+) sDC subsets rapidly up-regulated expression of costimulatory molecules and began to produce proinflammatory cytokines. Their tendency to undergo apoptosis ex vivo simultaneously increased, and in vivo the number of conventional DCs in the spleen decreased markedly, dropping approximately 2-fold by day 3 pi. Conversely, the number of plasmacytoid (CD11clowB220+) DCs in the spleen increased, so that they constituted almost 40% of sDCs by day 3 pi. Type 1 IFN production was up-regulated in plasmacytoid DCs by 24 h pi. Analysis of DC activation and maturation in mice unable to respond to type 1 IFNs implicated these cytokines in driving infection-associated phenotypic activation of conventional DCs and their enhanced tendency to undergo apoptosis, but also indicated the existence of type 1 IFN-independent pathways for the functional maturation of DCs during LCMV infection. PMID:15699111

  11. Stable growth transformation of human T lymphocytes by herpesvirus saimiri.

    PubMed Central

    Biesinger, B; Müller-Fleckenstein, I; Simmer, B; Lang, G; Wittmann, S; Platzer, E; Desrosiers, R C; Fleckenstein, B

    1992-01-01

    Herpesvirus saimiri induces T-cell lymphomas in various species of New World monkeys and in rabbits, and it is able to immortalize monkey T lymphocytes in vitro. Sequences responsible for these effects have been localized to a region of the genome that varies significantly among the virus subgroups A, B, and C. We now report that infection of human blood lymphocytes and thymocytes with strains of subgroup C, in contrast to viruses of the other subgroups, yields continuously proliferating T-cell lines with the phenotype of mature CD4- or CD8-positive cells. Infection with strains of Herpes-virus saimiri subgroup C can thus be used to generate human T-cell lines for a variety of immunological and developmental studies. Images PMID:1313581

  12. Telomerase Contributes to Fludarabine Resistance in Primary Human Leukemic Lymphocytes

    PubMed Central

    Shawi, May; Chu, Tsz Wai; Martinez-Marignac, Veronica; Yu, Y.; Gryaznov, Sergei M.; Johnston, James B.; Lees-Miller, Susan P.; Assouline, Sarit E.; Autexier, Chantal; Aloyz, Raquel

    2013-01-01

    We report that Imetelstat, a telomerase inhibitor that binds to the RNA component of telomerase (hTR), can sensitize primary CLL lymphocytes to fludarabine in vitro. This effect was observed in lymphocytes from clinically resistant cases and with cytogenetic abnormalities associated with bad prognosis. Imetelstat mediated-sensitization to fludarabine was not associated with telomerase activity, but with the basal expression of Ku80. Since both Imetelstat and Ku80 bind hTR, we assessed 1) if Ku80 and Imetelstat alter each other's binding to hTR in vitro and 2) the effect of an oligonucleotide complementary to the Ku binding site in hTR (Ku oligo) on the survival of primary CLL lymphocytes exposed to fludarabine. We show that Imetelstat interferes with the binding of Ku70/80 (Ku) to hTR and that the Ku oligo can sensitize CLL lymphocytes to FLU. Our results suggest that Ku binding to hTR may contribute to fludarabine resistance in CLL lmphocytes. This is the first report highlighting the potentially broad effectiveness of Imetelstat in CLL, and the potential biological and clinical implications of a functional interaction between Ku and hTR in primary human cancer cells. PMID:23922990

  13. Telomerase contributes to fludarabine resistance in primary human leukemic lymphocytes.

    PubMed

    Shawi, May; Chu, Tsz Wai; Martinez-Marignac, Veronica; Yu, Y; Gryaznov, Sergei M; Johnston, James B; Lees-Miller, Susan P; Assouline, Sarit E; Autexier, Chantal; Aloyz, Raquel

    2013-01-01

    We report that Imetelstat, a telomerase inhibitor that binds to the RNA component of telomerase (hTR), can sensitize primary CLL lymphocytes to fludarabine in vitro. This effect was observed in lymphocytes from clinically resistant cases and with cytogenetic abnormalities associated with bad prognosis. Imetelstat mediated-sensitization to fludarabine was not associated with telomerase activity, but with the basal expression of Ku80. Since both Imetelstat and Ku80 bind hTR, we assessed 1) if Ku80 and Imetelstat alter each other's binding to hTR in vitro and 2) the effect of an oligonucleotide complementary to the Ku binding site in hTR (Ku oligo) on the survival of primary CLL lymphocytes exposed to fludarabine. We show that Imetelstat interferes with the binding of Ku70/80 (Ku) to hTR and that the Ku oligo can sensitize CLL lymphocytes to FLU. Our results suggest that Ku binding to hTR may contribute to fludarabine resistance in CLL lmphocytes. This is the first report highlighting the potentially broad effectiveness of Imetelstat in CLL, and the potential biological and clinical implications of a functional interaction between Ku and hTR in primary human cancer cells. PMID:23922990

  14. Chrysin induces apoptosis in peripheral blood lymphocytes isolated from human chronic lymphocytic leukemia.

    PubMed

    Zaric, Milan; Mitrovic, Marina; Nikolic, Ivana; Baskic, Dejan; Popovic, Suzana; Djurdjevic, Predrag; Milosavljevic, Zoran; Zelen, Ivanka

    2015-01-01

    Chronic lymphocytic leukemia (CLL) develops due to an imbalance between apoptosis and proliferation of B lymphocytes. Chrysin induced apoptosis in leukemia cell lines such as U937, MO7e, THP-1 and HL-60, but there has not yet been data demonstrating the apoptotic effect of chrysin on CLL cells. Therefore, in our investigation we examined the cytotoxicity of chrysin against two leukemia cell lines, MOLT-4 and JVM-13, peripheral blood lymphocytes isolated from B-CLL patients and peripheral blood mononuclear cells (PBMC) from healthy individuals in vitro. The effect of chrysin on viability of MOLT-4 and JVM-13 cell lines, B-CLL cells derived from 28 patients and PBMC from 16 healthy subjects was determined by MTT assay. The type of cell death induced by chrysin was verified by Annexin V/7AAD assay and acridine orange and ethidium bromide (AO/EB) staining assay. Intracellular localisation and endogenic expression of apoptotic proteins including Bax, Bcl-2, cytochrome c and caspase-3 were determined by flow cytometry and fluorescent microscopy. Our results demonstrated that exposure of MOLT-4, JVM-13 cell lines and B-CLL cells to the concentration of chrysin of 10μM and higher selectively decreased viability of cells in this cell population, but not in the PBMC derived from healthy subjects; LC50 values of chrysin for B-CLL cells were 51μM for 24 hours and 32μM for 48 hours of incubation, respectively. Our findings demonstrated that chrysin induces the activation of proapoptotic Bax and decreases the expression of antiapoptotic Bcl-2 protein, releases cytochrome c from mitochondria into cytosol and cleavages/activates caspase-3, subsequently leading to the activation of apoptosis of B-CLL cells. Together, these findings suggest that chrysin selectively induces apoptosis of peripheral blood lymphocytes isolated from human chronic lymphocytic leukemia patients via mitochondrial pathway in vitro and that it might have a promising role as a potential future antileukemic

  15. Cellular and humoral immune reactions in chronic active liver disease. II. Lymphocyte subsets and viral antigens in liver biopsies of patients with acute and chronic hepatitis B.

    PubMed Central

    Eggink, H F; Houthoff, H J; Huitema, S; Wolters, G; Poppema, S; Gips, C H

    1984-01-01

    The characteristics and distribution of the inflammatory infiltrate in liver biopsies of 25 patients with hepatitis B viral (HBV) infection were studied in relation to the distribution and expression of HBV antigens. Mononuclear subsets were characterized with monoclonal (OKT, OKM, Leu) antibodies to surface antigens. For the demonstration of viral antigens directly conjugated antibodies to surface (HBsAg), core (HBcAg) and 'e' (HBeAg) antigen were used. For the study of mutual relations all methods were performed on serial cut tissue sections. In chronic active hepatitis B (CAH-B, n = 12) OKT8+ lymphocytes of T cell origin were the only cell type present in areas with liver cell degeneration and T cell cytotoxicity appears to be the only immune mechanism. In chronic persistent hepatitis B (CPH-B, n = 7) the only conspicuous feature was the presence of many Leu 3+ lymphocytes of the helper/inducer population in the portal tracts. In acute hepatitis B (AHB, n = 6) OKT8+ cells of non-T origin (OKT1-,3-) and Leu 7+ cells of presumed natural killer (NK) potential predominated in the areas with liver cell necrosis, and non-T cell cytotoxicity appears to be the predominant immune mechanism. In none of these disease entities a positive spatial relation could be established between the cytotoxic cells and the demonstrable expression of HBV antigens in hepatocytes. It is concluded that differences in immunological reaction pattern may explain the different course in the three forms of HBV infection studied. Images Fig. 1 Fig. 2 PMID:6713726

  16. In vitro effect of fenthion on human lymphocytes

    SciTech Connect

    Rani, M.V.U. ); Rao, M.S. )

    1991-08-01

    Fenthion is an organophosphorus insecticide which is extensively used in control of leaf hoppers, cutworms, mites on vegetable crops. It has been reported that organophosphorus pesticides cause a significant increase in sister chromatid exchanges in mammalian cell lines. A significant increase of chromosomal aberrations has been reported in rural population exposed to pesticides. Organosphosphorus pesticides malathion, diazinon, dimethoate, phosdrin and dursban induced sister chromatid exchanges in human lymphoid cells. Exchange type of aberration has been reported in fluoriculturist who were exposed to organophosphorus, organochlorine pesticides. In the present investigation an attempt has been made to evaluate the cytogenetic effect of fenthion in human lymphocyte cultures in vitro.

  17. A Progenitor Cell Expressing Transcription Factor RORγt Generates All Human Innate Lymphoid Cell Subsets.

    PubMed

    Scoville, Steven D; Mundy-Bosse, Bethany L; Zhang, Michael H; Chen, Li; Zhang, Xiaoli; Keller, Karen A; Hughes, Tiffany; Chen, Luxi; Cheng, Stephanie; Bergin, Stephen M; Mao, Hsiaoyin C; McClory, Susan; Yu, Jianhua; Carson, William E; Caligiuri, Michael A; Freud, Aharon G

    2016-05-17

    The current model of murine innate lymphoid cell (ILC) development holds that mouse ILCs are derived downstream of the common lymphoid progenitor through lineage-restricted progenitors. However, corresponding lineage-restricted progenitors in humans have yet to be discovered. Here we identified a progenitor population in human secondary lymphoid tissues (SLTs) that expressed the transcription factor RORγt and was unique in its ability to generate all known ILC subsets, including natural killer (NK) cells, but not other leukocyte populations. In contrast to murine fate-mapping data, which indicate that only ILC3s express Rorγt, these human progenitor cells as well as human peripheral blood NK cells and all mature ILC populations expressed RORγt. Thus, all human ILCs can be generated through an RORγt(+) developmental pathway from a common progenitor in SLTs. These findings help establish the developmental signals and pathways involved in human ILC development. PMID:27178467

  18. Electrostimulation of rat callus cells and human lymphocytes in vitro

    SciTech Connect

    Aro, H.; Eerola, E.; Aho, A.J.; Penttinen, R.

    1984-01-01

    Asymmetrical pulsing low voltage current was supplied via electrodes to cultured rat fracture callus cells and human peripheral blood lymphocytes. The (/sup 3/H)thymidine incorporation of the callus cells and 5-(/sup 125/I)iodo-2'-deoxyuridine incorporation of the lymphocytes were determined. The growth pattern of callus cells (estimated by cellular density) did not respond to electrical stimulation. However, the uptake of (/sup 3/H)thymidine was increased at the early phase of cell proliferation and inhibited at later phases of proliferation. The (/sup 3/H)thymidine uptake of confluent callus cell cultures did not respond to electrical stimulation. Lymphocytes reacted in a similar way; stimulated cells took up more DNA precursor than control cells at the early phase of stimulation. During cell division, induced by the mitogens phytohemagglutinin and Concanavalin-A, the uptake of DNA precursor by stimulated cells was constantly inhibited. The results suggest that electrical stimuli affect the uptake mechanisms of cell membranes. The duality of the effect seems to be dependent on the cell cycle.

  19. Immunohistochemical identification of cytotoxic lymphocytes using human perforin monoclonal antibody.

    PubMed Central

    Hameed, A.; Olsen, K. J.; Cheng, L.; Fox, W. M.; Hruban, R. H.; Podack, E. R.

    1992-01-01

    Perforin is a potent cytolytic pore-forming protein expressed in cytoplasmic granules of cytotoxic T lymphocytes and natural killer cells. A new monoclonal antibody raised against human perforin was used to detect both in vitro and in vivo perforin expression in cytotoxic cells. Immunohistochemical analysis of human peripheral blood mononuclear cells cultured in recombinant interleukin-2 (rIL-2) showed strong granular cytoplasmic staining of the IL-2 activated cytotoxic cells. Fresh-frozen tissue sections from patients with heart allograft rejection were also stained. Strong granular cytoplasmic staining of the mononuclear inflammatory infiltrate characteristic for perforin in cardiac allograft rejection was observed. The detection and quantitative analysis of perforin-associated cytotoxic cells by the human anti-perforin monoclonal antibody will help to evaluate the significance of these functionally distinct cytotoxic cells in human tissue. Images Figure 1 PMID:1374586

  20. Immunoregulatory properties of vasoactive intestinal peptide in human T cell subsets: implications for rheumatoid arthritis.

    PubMed

    Gutiérrez-Cañas, Irene; Juarranz, Yasmina; Santiago, Begoña; Martínez, Carmen; Gomariz, Rosa P; Pablos, José Luis; Leceta, Javier

    2008-03-01

    Rheumatoid arthritis (RA) is a chronic inflammatory autoimmune disease whose pathogenesis is not completely understood. Unbalanced Th1/Th2 T-cell polarization has been suggested to play a pathogenetic role and therefore, modulation of T-cell polarization is a potential therapeutic target. Vasoactive intestinal peptide (VIP) is a broadly distributed peptide that exerts anti-inflammatory and immunomodulatory effects, in the collagen-induced arthritis (CIA) murine model of RA, and ex vivo, in synovial cells from RA patients. In the present study, we have found that polyclonal stimulation of peripheral blood lymphocytes (PBL) from RA patients produces higher levels of inflammatory mediators and lower levels of Th1 cytokines than PBL from healthy controls; moreover, VIP has negligible effects on inflammatory mediators and Th1 cytokines produced by PBL from healthy controls but favours Th2 profile and enhanced IL-10 production after stimulation. VIP increases the levels of IL-10 and IL-4 in the supernatant of human CD4(+)CD45RA(+) cells cultured in a non-conditioned or a Th2-conditioned situation. In contrast, VIP does not modify the production of these cytokines in a Th1-conditioned medium. In summary, VIP can differentially modify the functional capacity of human lymphocytes by inducing Th2/Treg differentiation depending on their previous phenotype. PMID:17951026

  1. Primary Human Leukocyte Subsets Differentially Express Vaccinia Virus Receptors Enriched in Lipid Rafts

    PubMed Central

    Byrd, Daniel; Amet, Tohti; Hu, Ningjie; Lan, Jie; Hu, Sishun

    2013-01-01

    Poxviruses, including vaccinia virus (VV) and canarypox virus (ALVAC), do not indiscriminately infect all cell types of the primary human leukocytes (PHLs) that they encounter but instead demonstrate an extremely strong bias toward infection of monocytes and monocyte lineage cells. We studied the specific molecular events that determine the VV tropism for major PHL subsets including monocytes, B cells, neutrophils, NK cells, and T cells. We found that VV exhibited an extremely strong bias of cell surface protein-dependent binding to monocytes, B cells, and activated T cells to a similar degree and to neutrophils to a much lesser extent. Resting T cells and resting NK cells exhibited only trace amounts of VV binding. Activated T cells, however, became permissive to VV binding, infection, and replication, while activated NK cells still resisted VV binding. VV binding strongly colocalized with lipid rafts on the surfaces of all VV binding-susceptible PHL subsets, even when lipid rafts were relocated to cell uropods upon cell polarization. Immunosera raised against detergent-resistant membranes (DRMs) from monocytes or activated T cells, but not resting T cells, effectively cross-blocked VV binding to and infection of PHL subsets. CD29 and CD98, two lipid raft-associated membrane proteins that had been found to be important for VV entry into HeLa cells, had no effect on VV binding to and infection of primary activated T cells. Our data indicate that PHL subsets express VV protein receptors enriched in lipid rafts and that receptors are cross-presented on all susceptible PHLs. PMID:23785200

  2. The interaction of human macrophage subsets with silicone as a biomaterial.

    PubMed

    Vijaya Bhaskar, Thanga Bhuvanesh; Ma, Nan; Lendlein, Andreas; Roch, Toralf

    2015-01-01

    Silicones are widely used as biomaterials for medical devices such as extracorporeal equipments. However, there is often conflicting evidence about their supposed cell- and histocompatibility. Macrophages could mediate silicone-induced adverse responses such as foreign body reaction and fibrous encapsulation. The polarization behaviour of macrophages could determine the clinical outcome after implantation of biomaterials. Induction of classically activated macrophages (CAM) may induce and support uncontrolled inflammatory responses and undesired material degradation. In contrast, polarization into alternatively activated macrophages (AAM) is assumed to support healing processes and implant integration.This study compared the interaction of non-polarized macrophages (M0), CAM, and AAM with commercially available tissue culture polystyrene (TCP) and a medical grade silicone-based biomaterial, regarding the secretion of inflammatory mediators such as cytokines and chemokines. Firstly, by using the Limulus amoebocyte lysate (LAL) test the silicone films were shown to be free of soluble endotoxins, which is the prerequisite to investigate their interaction with primary immune cells. Primary human monocyte-derived macrophages (M0) were polarized into CAM and AAM by addition of suitable differentiation factors. These macrophage subsets were incubated on the materials for 24 hours and their viability and cytokine secretion was assessed. In comparison to TCP, cell adhesion was lower on silicone after 24 hours for all three macrophage subsets. However, compared to TCP, silicone induced higher levels of certain inflammatory and chemotactic cytokines in M0, CAM, and AAM macrophage subsets.Conclusively, it was shown that silicone has the ability to induce a pro-inflammatory state to different magnitudes dependent on the macrophage subsets. This priming of the macrophage phenotype by silicone could explain the incidence of severe foreign body complications observed in vivo. PMID

  3. DEXAMETHASONE DIFFERENTIALLY DOWN-REGULATES L-SELECTIN (CD62L) EXPRESSION BY BOVINE LYMPHOCYTE SUBSETS IN VIVO AND DEPLETES THE INTESTINAL MUCOSA OF INTRAEPITHELIAL GAMMA DELTA CELLS

    Technology Transfer Automated Retrieval System (TEKTRAN)

    A hallmark of immune modulation by dexamethasone (DEX) is the suppression of lymphocyte proliferation. DEX also alters the expression of adhesion molecules on bovine lymphocytes. Although intraepithelial lymphocytes (IEL) mainly are non-proliferating cells, we hypothesized that DEX treatment of ca...

  4. Transcriptional specialization of human dendritic cell subsets in response to microbial vaccines

    PubMed Central

    Banchereau, Romain; Baldwin, Nicole; Cepika, Alma-Martina; Athale, Shruti; Xue, Yaming; Yu, Chun I; Metang, Patrick; Cheruku, Abhilasha; Berthier, Isabelle; Gayet, Ingrid; Wang, Yuanyuan; Ohouo, Marina; Snipes, LuAnn; Xu, Hui; Obermoser, Gerlinde; Blankenship, Derek; Oh, Sangkon; Ramilo, Octavio; Chaussabel, Damien; Banchereau, Jacques; Palucka, Karolina; Pascual, Virginia

    2014-01-01

    The mechanisms by which microbial vaccines interact with human APCs remain elusive. Herein, we describe the transcriptional programs induced in human DCs by pathogens, innate receptor ligands and vaccines. Exposure of DCs to influenza, Salmonella enterica and Staphylococcus aureus allows us to build a modular framework containing 204 transcript clusters. We use this framework to characterize the responses of human monocytes, monocyte-derived DCs and blood DC subsets to 13 vaccines. Different vaccines induce distinct transcriptional programs based on pathogen type, adjuvant formulation and APC targeted. Fluzone, Pneumovax and Gardasil, respectively, activate monocyte-derived DCs, monocytes and CD1c+ blood DCs, highlighting APC specialization in response to vaccines. Finally, the blood signatures from individuals vaccinated with Fluzone or infected with influenza reveal a signature of adaptive immunity activation following vaccination and symptomatic infections, but not asymptomatic infections. These data, offered with a web interface, may guide the development of improved vaccines. PMID:25335753

  5. Human lymphocyte polymorphisms detected by quantitative two-dimensional electrophoresis

    SciTech Connect

    Goldman, D.; Merril, C.R.

    1983-09-01

    A survey of 186 soluble lymphocyte proteins for genetic polymorphism was carried out utilizing two-dimensional electrophoresis of /sup 14/C-labeled phytohemagglutinin (PHA)-stimulated human lymphocyte proteins. Nineteen of these proteins exhibited positional variation consistent with independent genetic polymorphism in a primary sample of 28 individuals. Each of these polymorphisms was characterized by quantitative gene-dosage dependence insofar as the heterozygous phenotype expressed approximately 50% of each allelic gene product as was seen in homozygotes. Patterns observed were also identical in monozygotic twins, replicate samples, and replicate gels. The three expected phenotypes (two homozygotes and a heterozygote) were observed in each of 10 of these polymorphisms while the remaining nine had one of the homozygous classes absent. The presence of the three phenotypes, the demonstration of gene-dosage dependence, and our own and previous pedigree analysis of certain of these polymorphisms supports the genetic basis of these variants. Based on this data, the frequency of polymorphic loci for man is: P . 19/186 . .102, and the average heterozygosity is .024. This estimate is approximately 1/3 to 1/2 the rate of polymorphism previously estimated for man in other studies using one-dimensional electrophoresis of isozyme loci. The newly described polymorphisms and others which should be detectable in larger protein surveys with two-dimensional electrophoresis hold promise as genetic markers of the human genome for use in gene mapping and pedigree analyses.

  6. slan/M-DC8+ cells constitute a distinct subset of dendritic cells in human tonsils

    PubMed Central

    Micheletti, Alessandra; Finotti, Giulia; Calzetti, Federica; Lonardi, Silvia; Zoratti, Elisa; Bugatti, Mattia; Stefini, Stefania; Vermi, William; Cassatella, Marco A.

    2016-01-01

    Human blood dendritic cells (DCs) include three main distinct subsets, namely the CD1c+ and CD141+ myeloid DCs (mDCs) and the CD303+ plasmacytoid DCs (pDCs). More recently, a population of slan/M-DC8+ cells, also known as “slanDCs”, has been described in blood and detected even in inflamed secondary lymphoid organs and non-lymphoid tissues. Nevertheless, hallmarks of slan/M-DC8+ cells in tissues are poorly defined. Herein, we report a detailed characterization of the phenotype and function of slan/M-DC8+ cells present in human tonsils. We found that tonsil slan/M-DC8+ cells represent a unique DC cell population, distinct from their circulating counterpart and also from all other tonsil DC and monocyte/macrophage subsets. Phenotypically, slan/M-DC8+ cells in tonsils display a CD11c+HLA-DR+CD14+CD11bdim/negCD16dim/negCX3CR1dim/neg marker repertoire, while functionally they exhibit an efficient antigen presentation capacity and a constitutive secretion of TNFα. Notably, such DC phenotype and functions are substantially reproduced by culturing blood slan/M-DC8+ cells in tonsil-derived conditioned medium (TDCM), further supporting the hypothesis of a full DC-like differentiation program occurring within the tonsil microenvironment. Taken together, our data suggest that blood slan/M-DC8+ cells are immediate precursors of a previously unrecognizedcompetent DC subset in tonsils, and pave the way for further characterization of slan/M-DC8+ cells in other tissues. PMID:26695549

  7. Monoclonal antibodies to antigens on human neutrophils, activated T lymphocytes, and acute leukemia blast cells

    SciTech Connect

    Miterev, G.Yu.; Burova, G.F.; Puzhitskaya, M.S.; Danilevich, S.V.; Bulycheva, T.I.

    1987-11-01

    The authors describe the production of two mouse hybridomas secreting monoclonal antibodies to antigenic determinants of the surface membranes of human neutrophils, activated T lymphocytes, and acute leukemic blast cells. The degree of lymphocyte stimulation was estimated from incorporation of /sup 3/H-thymidine with parallel microculture. Monoclonal antibodies of supernatants of hybridoma cultures shown here reacted in both immunofluorescence test and cytotoxicity test with surface membrane antigens on the majority of neutrophils and PHA-activated peripheral blood lymphocytes from healthy subjects, but did not give positive reactions with unactivated lymphocytes, adherent monocytes, erythrocytes, and alloantigen-stimulated lymphocytes.

  8. Evaluation of gamma-Induced Apoptosis in Human Peripheral Blood Lymphocytes

    SciTech Connect

    Baranova, Elena; Boreyko, Alla; Ravnachka, Ivanka; Saveleva, Maria

    2010-01-05

    Several experiments have been performed to study regularities in the induction of apoptotic cells in human lymphocytes by {sup 60}Cogamma-rays at different times after irradiation. Apoptosis induction by {sup 60}Cogamma-rays in human lymphocytes in different cell cycle phases (G{sub 0}, S, G{sub 1}, and G{sub 2}) has been studied. The maximal apoptosis output in lymphocyte cells was observed in the S phase. Modifying effect of replicative and reparative DNA synthesis inhibitors - 1-beta-D-arabinofuranosylcytosine (Ara-C) and hydroxyurea (Hu) - on the kinetics of {sup 60}Cogamma-rays induced apoptosis in human lymphocytes has been studied.

  9. CHARACTERIZATION OF NORMAL HUMAN LUNG LYMPHOCYTES AND INTERLEUKIN-2-INDUCED LUNG T CELL LINES

    EPA Science Inventory

    Lymphocytes from the lower respiratory tract were obtained by bronchoalveolar lavage of healthy, non-smoking individuals. arious monoclonal antibodies characterizing activated T cells, helper-inducer and suppressor-inducer T cell subsets, and naive versus memory cells were used t...

  10. Radiosensitivity of CD4 or CD8 positive human T-lymphocytes by an in vitro colony formation assay

    SciTech Connect

    Nakamura, N.; Kusunoki, Y.; Akiyama, M. )

    1990-08-01

    The recent development of an in vitro lymphocyte colony assay makes it possible to examine variations in the radiosensitivity of humans using peripheral blood lymphocytes (PBL) instead of the skin fibroblast assay. Our recent study showed that most of the colonies consisted of lymphocytes bearing CD4 or CD8 antigens. Since the fraction of CD4+ and CD8+ cells in PBL differs among individuals, we suspected that individual radiosensitivity might be biased by the different subset frequencies if the dose-survival curves of the CD4+ and CD8+ cells were different from each other. In the present study, CD4+ (helper/inducer T) and CD8+ (suppressor/cytotoxic T) lymphocytes were isolated from PBL and their dose-survival curves were determined. The results showed that the D10 (dose required to reduce the surviving fraction to 10%) was similar for these two types of cells (3.13 +/- 0.10 Gy (mean +/- SD) for CD4+, 3.34 +/- 0.50 Gy for CD8+ and 3.14 +/- 0.17 Gy for the unsorted cells), supporting the use of the whole PBL population for the screening of individuals with altered radiosensitivity.

  11. Differential in vivo activation of monocyte subsets during low-grade inflammation through experimental endotoxemia in humans.

    PubMed

    Thaler, B; Hohensinner, P J; Krychtiuk, K A; Matzneller, P; Koller, L; Brekalo, M; Maurer, G; Huber, K; Zeitlinger, M; Jilma, B; Wojta, J; Speidl, W S

    2016-01-01

    Human monocytes are a heterogeneous cell population, which can be divided into a classical (CD14++CD16-), a non-classical (CD14+CD16+), and an intermediate (CD14++CD16+) subset. We hypothesized that low-grade inflammation may differentially affect monocyte subsets. We used a human lipopolysaccharide (LPS) infusion model to mimic low-grade inflammation to identify, which monocyte subsets are preferentially activated under these conditions. Monocyte subsets were identified by staining for CD14 and CD16, activation status of monocytes was analyzed by staining for CD11b and a novel in situ mRNA hybridization approach to detect IL-6 and IL-8 specific mRNA at the single-cell level by flow cytometry. After LPS challenge, cell numbers of monocyte subsets dropped after 2 h with cell numbers recovering after 6 h. Distribution of monocyte subsets was skewed dramatically towards the intermediate subset after 24 h. Furthermore, intermediate monocytes displayed the largest increase of CD11b expression after 2 h. Finally, IL-6 and IL-8 mRNA levels increased in intermediate and non-classical monocytes after 6 h whereas these mRNA levels in classical monocytes changed only marginally. In conclusion, our data indicates that the main responding subset of monocytes to standardized low-grade inflammation induced by LPS in humans is the CD14++CD16+ intermediate subset followed by the CD14+CD16+ non-classical monocyte subset. Circulating classical monocytes showed comparably less reaction to LPS challenge in vivo. PMID:27444882

  12. Differential in vivo activation of monocyte subsets during low-grade inflammation through experimental endotoxemia in humans

    PubMed Central

    Thaler, B.; Hohensinner, P. J.; Krychtiuk, K. A.; Matzneller, P.; Koller, L.; Brekalo, M.; Maurer, G.; Huber, K.; Zeitlinger, M.; Jilma, B.; Wojta, J.; Speidl, W. S.

    2016-01-01

    Human monocytes are a heterogeneous cell population, which can be divided into a classical (CD14++CD16−), a non-classical (CD14+CD16+), and an intermediate (CD14++CD16+) subset. We hypothesized that low-grade inflammation may differentially affect monocyte subsets. We used a human lipopolysaccharide (LPS) infusion model to mimic low-grade inflammation to identify, which monocyte subsets are preferentially activated under these conditions. Monocyte subsets were identified by staining for CD14 and CD16, activation status of monocytes was analyzed by staining for CD11b and a novel in situ mRNA hybridization approach to detect IL-6 and IL-8 specific mRNA at the single-cell level by flow cytometry. After LPS challenge, cell numbers of monocyte subsets dropped after 2 h with cell numbers recovering after 6 h. Distribution of monocyte subsets was skewed dramatically towards the intermediate subset after 24 h. Furthermore, intermediate monocytes displayed the largest increase of CD11b expression after 2 h. Finally, IL-6 and IL-8 mRNA levels increased in intermediate and non-classical monocytes after 6 h whereas these mRNA levels in classical monocytes changed only marginally. In conclusion, our data indicates that the main responding subset of monocytes to standardized low-grade inflammation induced by LPS in humans is the CD14++CD16+ intermediate subset followed by the CD14+CD16+ non-classical monocyte subset. Circulating classical monocytes showed comparably less reaction to LPS challenge in vivo. PMID:27444882

  13. Effects of budlein A on human neutrophils and lymphocytes

    PubMed Central

    KNOB, Carollinie Dias; SILVA, Milena; GASPAROTO, Thaís Helena; OLIVEIRA, Carine Ervolino; AMÔR, Nádia Ghinelli; ARAKAWA, Nilton Syogo; COSTA, Fernando Batista; CAMPANELLI, Ana Paula

    2016-01-01

    ABSTRACT Sesquiterpene lactones (SLs) are the active constituents of a variety of medicinal plants used in traditional medicine for the treatment of inflammatory diseases and other ailments. Objective In this study, we evaluated whether budlein A modulates the activation of innate and adaptive immune cells such as neutrophils and lymphocytes. Material and Methods Our research group has investigated several plant species and several compounds have been isolated, identified, and their medical potential evaluated. Budlein A is a SL isolated from the species Aldama buddlejiformis and A. robusta (Asteraceae) and shows anti-inflammatory and anti-nociceptive activities. Advances in understanding how plant-derived substances modulate the activation of innate and adaptive immune cells have led to the development of new therapies for human diseases. Results Budlein A inhibited MPO activity, IL-6, CXCL8, IL-10, and IL-12 production and induces neutrophil apoptosis. In contrast, budlein A inhibited lymphocyte proliferation and IL-2, IL-10, TGF-β, and IFN-γ production, but it did not lead to cell death. Conclusions Collectively, our results indicate that budlein A shows distinct immunomodulatory effects on immune cells. PMID:27383709

  14. Unstable high molecular weight inverted repetitive DNA in human lymphocytes.

    PubMed Central

    Rogers, J C; Rucinsky, T E

    1982-01-01

    About 1% of newly synthesized DNA from PHA-stimulated human lymphocytes can be isolated as large (up to 90 kilobase pairs) double stranded fragments that resist sequential alkali and heat denaturation steps but are not closed circular. By electron microscopy about 1% have single-strand hairpin loops at one end and therefore present inverted repetitive sequences (IR-DNA). Most of the remainder have a blunt-appearing double-strand terminus at both ends (78%) or one end (18%). Indirect evidence indicates that these also are inverted complementary structures with terminal hairpin loops too small to be visualized: (1) Treatment with either a 5' or 3' single-strand exonuclease generates essentially only fragments with a single strand at one end; (2) with partial denaturation, the number of fragments with identifiable single-strand hairpin loops increases (to about 20%); (3) after S1 nuclease digestion, greater than 95% can be fully heat denatured. Cot analysis indicates that these fragments are derived from dispersed sites throughout the genome. Up to 25% of DNA released from lymphocytes during growth similarly resists denaturation, and released DNA and IR-DNA are both enriched in the same set of repetitive sequences. Thus at least a portion of IR-DNA appears to be unstable. Images PMID:7145706

  15. Spaceflight alters microtubules and increases apoptosis in human lymphocytes (Jurkat)

    NASA Technical Reports Server (NTRS)

    Lewis, M. L.; Reynolds, J. L.; Cubano, L. A.; Hatton, J. P.; Lawless, B. D.; Piepmeier, E. H.

    1998-01-01

    Alteration in cytoskeletal organization appears to underlie mechanisms of gravity sensitivity in space-flown cells. Human T lymphoblastoid cells (Jurkat) were flown on the Space Shuttle to test the hypothesis that growth responsiveness is associated with microtubule anomalies and mediated by apoptosis. Cell growth was stimulated in microgravity by increasing serum concentration. After 4 and 48 h, cells filtered from medium were fixed with formalin. Post-flight, confocal microscopy revealed diffuse, shortened microtubules extending from poorly defined microtubule organizing centers (MTOCs). In comparable ground controls, discrete microtubule filaments radiated from organized MTOCs and branched toward the cell membrane. At 4 h, 30% of flown, compared to 17% of ground, cells showed DNA condensation characteristic of apoptosis. Time-dependent increase of the apoptosis-associated Fas/ APO-1 protein in static flown, but not the in-flight 1 g centrifuged or ground controls, confirmed microgravity-associated apoptosis. By 48 h, ground cultures had increased by 40%. Flown populations did not increase, though some cells were cycling and actively metabolizing glucose. We conclude that cytoskeletal alteration, growth retardation, and metabolic changes in space-flown lymphocytes are concomitant with increased apoptosis and time-dependent elevation of Fas/APO-1 protein. We suggest that reduced growth response in lymphocytes during spaceflight is linked to apoptosis.

  16. In vitro sensitization of human lymphocytes to a myeloma cell-related antigen

    SciTech Connect

    Whitson, M.E.; Griffin, G.D.; Novelli, G.D.; Solomon, A.

    1981-01-01

    Peripheral blood lymphocytes from normal human donors were cocultivated with cells from two established human multiple myeloma cell lines, RPMI 8226 and K-737, and with lymphoblastoid cells from a third B cell line, RAMM. After a comparison of three methods of lymphocyte sensitization, a 6-day incubation protocol with equal numbers of normal lymphocytes and mitomycin C-treated tumor cells was selected. Cells fom the RPMI 8226 myeloma line stimulated the differentiation of lymphocytes into cytotoxic effector cells as measured by /sup 51/Cr release from labeled target cells. The RPMI 8226-sensitized lymphocytes were cytotoxic for myeloma cells (RPMI 8226 and K-737) and for lymphoblastoid cells (RAMM) but not for cells from human lung tumor lines (A549, A427, MB9812), a breast carcinoma line (ALAB), a normal diploid fibroblast line (HSBP), or normal lymphocytes.

  17. Stimulation of human lymphocytes by Herpes simplex virus antigens.

    PubMed Central

    Starr, S E; Karatela, S A; Shore, S L; Duffey, A; Nahmias, A J

    1975-01-01

    Lymphocytes from individuals with laboratory evidence of prior infection with herpes simplex virus (HSV) type 1 or type 2 demonstrated transformation (av antigens. Higher stimulation indexes were obtained when lymphocytes were incubated with the homologous as compared with the heterologous antigen. Higher mean lymphocyte stimulation indexes were also demonstrated in seropositive as compared with seronegative individuals. Lymphocytes from children with HSV-1 stomatitis usually became responsive to HSV-1 antigen within 2 to 6 weeks after the onset of illness. Lymphocytes from infants with neonatal HSV-2 infection were stimulated by HSV-2 antigen. PMID:163788

  18. The effect of adherent and phagocytic cells on human lymphocyte PHA responsiveness.

    PubMed

    Potter, M R; Moore, M

    1977-01-01

    The effect of small numbers of adherent and phagocytic cells on the human peripheral blood lymphocyte response to PHA was examined by depleting these cells from lymphocyte preparations. Lymphocyte preparations obtained by centrifugation on Ficoll--Triosil, which contained on average 85% lymphocytes, responded well to PHA. Depletion of cells adhering to nylon fibre, giving a population containing on average 95% lymphocytes, resulted in a considerably reduced response. Depletion of cells that adhered to plastic or ingested iron powder to give populations containing on average 90% lymphocytes, also reduced the PHA response, but to a lesser extent. Reduction in PHA responsiveness correlated with increasing lymphocyte purity. The responsiveness of nylon-column-filtered cells could be restored by adding a small number of cells from a monocyte-rich population. PMID:300303

  19. Communication between Human Dendritic Cell Subsets in Tuberculosis: Requirements for Naive CD4+ T Cell Stimulation

    PubMed Central

    Lozza, Laura; Farinacci, Maura; Bechtle, Marina; Stäber, Manuela; Zedler, Ulrike; Baiocchini, Andrea; del Nonno, Franca; Kaufmann, Stefan H. E.

    2014-01-01

    Human primary dendritic cells (DCs) are heterogeneous by phenotype, function, and tissue localization and distinct from inflammatory monocyte-derived DCs. Current information regarding the susceptibility and functional role of primary human DC subsets to Mycobacterium tuberculosis (Mtb) infection is limited. Here, we dissect the response of different primary DC subsets to Mtb infection. Myeloid CD11c+ cells and pDCs (C-type lectin 4C+ cells) were located in human lymph nodes (LNs) of tuberculosis (TB) patients by histochemistry. Rare CD141hi DCs (C-type lectin 9A+ cells) were also identified. Infection with live Mtb revealed a higher responsiveness of myeloid CD1c+ DCs compared to CD141hi DCs and pDCs. CD1c+ DCs produced interleukin (IL)-6, tumor necrosis factor α, and IL-1β but not IL-12p70, a cytokine important for Th1 activation and host defenses against Mtb. Yet, CD1c+ DCs were able to activate autologous naïve CD4+ T cells. By combining cell purification with fluorescence-activated cell sorting and gene expression profiling on rare cell populations, we detected in responding CD4+ T cells, genes related to effector-cytolytic functions and transcription factors associated with Th1, Th17, and Treg polarization, suggesting multifunctional properties in our experimental conditions. Finally, immunohistologic analyses revealed contact between CD11c+ cells and pDCs in LNs of TB patients and in vitro data suggest that cooperation between Mtb-infected CD1c+ DCs and pDCs favors stimulation of CD4+ T cells. PMID:25071784

  20. Effect of steady magnetic field on human lymphocytes

    SciTech Connect

    Mileva, M.; Ivanov, B.; Bulanova, M.; Pantev, T.

    1983-01-01

    Exposure to steady magnetic field (SMF) for different periods of time did not elicit statistically reliable increase in chromosome aberrations in human peripheral blood lymphocytes. Metaphase analysis of Crepis capilaris cells revealed that SMF (9 k0e, 200 0e/cm) for 2 days did not induce chromosome aberrations. Nor were any changes demonstrated in roots of beans, onions and L-fibroblasts of subcutaneous tissue of mice and Chinese hamsters. The obtained data are indicative of absence of cytogenetic effect of SMF. The level and spectrum of chromosome aberrations did not exceed the values for spontaneous chromatic fragments in cultures. Cytogenetic analysis of DEDE cells of the Chinese hamster revealed a mild mutagenic effect of SMF. Chromosomal aberrations were also demonstrated after exposure (5 min) of garlic roots.

  1. Human cytotoxic T lymphocytes against the Plasmodium falciparum circumsporozoite protein.

    PubMed Central

    Malik, A; Egan, J E; Houghten, R A; Sadoff, J C; Hoffman, S L

    1991-01-01

    Cytotoxic T lymphocytes (CTL) against the circumsporozoite (CS) protein of malaria sporozoites protect against malaria in rodents. Although there is interest in developing human vaccines that induce CTL against the Plasmodium falciparum CS protein, humans have never been shown to produce CTL against any Plasmodium species protein or other parasite protein. We report that when peripheral blood mononuclear cells (PBMC) from three of four volunteers immunized with irradiated P. falciparum sporozoites were stimulated in vitro with a recombinant vaccinia virus expressing the P. falciparum CS protein or a peptide including only amino acids 368-390 of the P. falciparum CS protein [CS-(368-390)], the PBMC lysed autologous Epstein-Barr virus-transformed B cells transfected with the P. falciparum CS protein gene or incubated with CS-(368-390) tricosapeptide. Activity was antigen specific, genetically restricted, and dependent on CD8+ T cells. In one volunteer, seven peptides reflecting amino acids 311-400 were tested, and, as in B10.BR mice, CTL activity was only associated with the CS-(368-390) peptide. Development of an assay for studying human CTL against the CS and other malaria proteins and a method for constructing target cells by direct gene transfection provide a foundation for studying the role of CTL in protection against malaria. PMID:1707538

  2. Chromosome Aberration in Human Blood Lymphocytes Exposed to Energetic Protons

    NASA Technical Reports Server (NTRS)

    Hada, M.; George, Kerry A.; Cucinotta, F. A.

    2008-01-01

    During space flight, astronauts are exposed to a space radiation consisting of high-energy protons, high charge and energy (HZE) nuclei, as well as secondary particles that are generated when the primary particles penetrate the spacecraft shielding. Secondary particles have a higher LET value than primary protons and therefore expected to have a higher relative biological effectiveness (RBE). To investigate this theory, we exposed human peripheral blood lymphocytes to protons with energies of 250 MeV, 800MeV, 2 GeV, or 2.5 GeV. LET values for these protons ranged from 0.4 to 0.2 keV/micrometer. and doses ranged from 0.2 to 3 Gy. Over this energy the probability of nuclear reaction leading to secondary radiation, and the multiplicity of reaction produces such as neutrons and mesons increases substantially. The effect of aluminum and polyethylene shielding was also assessed using the 2 GeV and 2.5GeV proton beams. After exposure lymphocytes were stimulated to divide and chromosomes were collected from cells in the first G2 and metaphase cell cycle after exposure using a chemical induced premature chromosome condensation (PCC) technique. Dose response data for chromosome damage was analyzed using the fluorescence in situ hybridization (FISH) chromosome painting technique. Selected samples were also analyzed with multicolor FISH (mFISH) and multicolor banding FISH (mBAND) techniques. Data indicates that the dose response for simple-type exchanges is similar for proton and gamma exposure, whereas protons induce higher yields of complex exchanges that are LET dependent. RBE values will be presented for each proton energy, and the effects of shielding and possible cytogenetic signatures of proton exposure will be discussed.

  3. Modeling adenovirus latency in human lymphocyte cell lines.

    PubMed

    Zhang, Yange; Huang, Wen; Ornelles, David A; Gooding, Linda R

    2010-09-01

    Species C adenovirus establishes a latent infection in lymphocytes of the tonsils and adenoids. To understand how this lytic virus is maintained in these cells, four human lymphocytic cell lines that support the entire virus life cycle were examined. The T-cell line Jurkat ceased proliferation and died shortly after virus infection. BJAB, Ramos (B cells), and KE37 (T cells) continued to divide at nearly normal rates while replicating the virus genome. Viral genome numbers peaked and then declined in BJAB cells below one genome per cell at 130 to 150 days postinfection. Ramos and KE37 cells maintained the virus genome at over 100 copies per cell over a comparable period of time. BJAB cells maintained the viral DNA as a monomeric episome. All three persistently infected cells lost expression of the cell surface coxsackie and adenovirus receptor (CAR) within 24 h postinfection, and CAR expression remained low for at least 340 days postinfection. CAR loss proceeded via a two-stage process. First, an initial loss of cell surface staining for CAR required virus late gene expression and a CAR-binding fiber protein even while CAR protein and mRNA levels remained high. Second, CAR mRNA disappeared at around 30 days postinfection and remained low even after virus DNA was lost from the cells. At late times postinfection (day 180), BJAB cells could not be reinfected with adenovirus, even when CAR was reintroduced to the cells via retroviral transduction, suggesting that the expression of multiple genes had been stably altered in these cells following infection. PMID:20573817

  4. Human dendritic cells and macrophages. In situ immunophenotypic definition of subsets that exhibit specific morphologic and microenvironmental characteristics.

    PubMed Central

    Wood, G. S.; Turner, R. R.; Shiurba, R. A.; Eng, L.; Warnke, R. A.

    1985-01-01

    Using a panel of monoclonal antibodies and antisera in situ, the authors have defined subsets of human dendritic cells and macrophages that exhibit specific morphologic and microenvironmental characteristics. All subsets contained cells that reacted with antibodies directed against HLA-A,B,C, HLA-Dr, leukocyte common, Leu-M3, and Leu-3(T4) antigens. R4/23 and anti-S100 defined three major subsets. R4/23+, S100- cells constituted the B-cell-related follicular dendritic cells, which were identified only within the germinal center/mantle microenvironment of lymphoid follicles. R4/23-, S100+ cells constituted the T-cell-related dendritic cell subset. Anti-Leu-6(T6) further subdivided this group into Leu-6(T6)- interdigitating cells within the T-cell microenvironments of lymphoid organs and Leu-6(T6)+ Langerhans cells found predominantly in epithelial microenvironments, especially the skin. R4/23-, S100- cells constituted the nondendritic tissue macrophage subset which was widely distributed, primarily outside of dendritic-cell microenvironments. These data indicate that although dendritic cells and macrophages share several common antigenic features, morphologically and microenvironmentally distinct subsets express distinct immunologic phenotypes. Such data may provide insight into the ontogeny and function of these subsets and constitute a basis for the comparison of normal dendritic cells and macrophages to various histiocytic proliferative disorders. Images Figure 1 Figure 2 p78-c Figure 3 Figure 4 Figure 5 PMID:3985124

  5. B Cell Anergy Modulated by TLR1/2 and the miR-17∼92 Cluster Underlies the Indolent Clinical Course of Chronic Lymphocytic Leukemia Stereotyped Subset #4.

    PubMed

    Ntoufa, Stavroula; Papakonstantinou, Nikos; Apollonio, Benedetta; Gounari, Maria; Galigalidou, Chrysi; Fonte, Eleonora; Anagnostopoulos, Achilles; Belessi, Chrysoula; Muzio, Marta; Ghia, Paolo; Stamatopoulos, Kostas

    2016-05-15

    Chronic lymphocytic leukemia (CLL) patients assigned to stereotyped subset #4 (mutated IGHV4-34/IGKV2-30 BCR Ig) display a particularly indolent disease course. Immunogenetic studies of the clonotypic BCR Ig of CLL subset #4 suggested a resemblance with B cells rendered anergic through chronic autoantigenic stimulation. In this article, we provide experimental evidence that subset #4 CLL cells show low IgG levels, constitutive ERK1/2 activation, and fail to either release intracellular Ca(2+) or activate MAPK signaling after BCR cross-linking, thus displaying a signature of B cell anergy at both biochemical and functional levels. Interestingly, TLR1/2 triggering restored BCR functionality, likely breaching the anergic state, and this was accompanied by induction of the miR-17∼92 cluster, whose members target critical BCR-associated molecules, including MAPKs. In conclusion, we demonstrate BCR anergy in CLL subset #4 and implicate TLR signaling and the miR-17∼92 cluster in the regulation of the anergic state. This detailed signaling profiling of subset #4 has implications for advanced understanding of the complex regulation of intracellular signaling pathways in CLL, currently a major therapeutic target of the disease. PMID:27059597

  6. Modulation of human lymphocyte mitogen responsiveness and interleukin-2 production by polymorphonuclear leukocytes.

    PubMed

    Lyte, M

    1990-06-01

    The response of human peripheral blood lymphocytes to the mitogenic lectins phytohemagglutinin (PHA) and pokeweed mitogen (PWM) was examined in the presence of autologous polymorphonuclear leukocytes (PMN). Experiments were performed at sub-optimal and optimal mitogen concentrations employing lymphocyte: PMN ratios over a three log cell concentration range. Increases of up to 25,000-fold in mitogen stimulated lymphocyte proliferation as determined by 3H-thymidine incorporation were observed in PMN supplemented lymphocyte cultures as compared to lymphocytes cultured in the absence of PMN or with irradiated lymphocytes serving as filler cells. Similar results were obtained for PHA stimulated IL-2 production. The degree of enhancement of lymphocyte reactivity by PMN was also shown to be dependent on the source of serum supplementation (autologous versus xenogeneic). These results indicate that cell ratio is a critical factor in examining lymphocyte-PMN interactions as well as serum supplementation used. Early reports which have indicated a suppressive or no effect of PMN on lymphocyte reactivity based on a single lymphocyte: PMN cell ratio may need to be re-evaluated. PMID:1967045

  7. Effects of flavonoids on human lymphocyte proliferative responses

    SciTech Connect

    Mookerjee, B.K.; Lee, T.P.; Logue, G.P.; Lippes, H.A.; Middleton, E.

    1986-01-01

    Flavonoids reversibly inhibit lymphocyte proliferative responses to phytomitogens, soluble antigens and phorbol esters by blocking an early event or events that follow stimulation. Quercetin and tangeretin inhibit thymidine transport in stimulated lymphocytes. These flavonoids reversibly inhibit antigen processing by monocytes and inhibit the expression of class II histocompatibility (DR) antigens in PBM cells.

  8. THE EFFECT OF OZONE ON HUMAN IMMUNITY: IN VITRO RESPONSIVENESS OF LYMPHOCYTES TO PHYTOHEMMAGGLUTININ

    EPA Science Inventory

    Peripheral blood lymphocytes from 20 human subjects exposed to 784 micrograms cu m ozone for 4 hours, and from 11 subjects exposed to clean air for the same length of time were studied for in vitro responsiveness to phytohemagglutinin (PHA). Thymus-derived (T) lymphocyte response...

  9. Viral Engineering of Chimeric Antigen Receptor Expression on Murine and Human T Lymphocytes.

    PubMed

    Hammill, Joanne A; Afsahi, Arya; Bramson, Jonathan L; Helsen, Christopher W

    2016-01-01

    The adoptive transfer of a bolus of tumor-specific T lymphocytes into cancer patients is a promising therapeutic strategy. In one approach, tumor specificity is conferred upon T cells via engineering expression of exogenous receptors, such as chimeric antigen receptors (CARs). Here, we describe the generation and production of both murine and human CAR-engineered T lymphocytes using retroviruses. PMID:27581020

  10. Vincristine-induced bystander effect in human lymphocytes.

    PubMed

    Testi, Serena; Azzarà, Alessia; Giovannini, Caterina; Lombardi, Sara; Piaggi, Simona; Facioni, Maria Sole; Scarpato, Roberto

    2016-07-01

    Bystander effect is a known radiobiological effect, widely described using ionizing radiations and which, more recently, has also been related to chemical mutagens. In this study, we aimed to assess whether or not a bystander response can be induced in cultured human peripheral lymphocytes by vincristine, a chemotherapeutic mutagen acting as spindle poison, and by mitomycin-C, an alkylating agent already known to induce this response in human lymphoblastoid cells. Designing a modified ad hoc protocol for the cytokinesis blocked micronucleus (MN) assay, we detected the presence of a dose-dependent bystander response in untreated cultures receiving the conditioned medium (CM) from mitomycin-C (MMC) or vincristine (VCR) treated cultures. In the case of MMC, MN frequencies, expressed as micronucleated binucleates, were: 13.5±1.41 at 6μM, 22±2.12 at 12μM or 28.25±5.13 at 15μM vs. a control value of 4.75±1.59. MN levels for VCR, expressed as micronucleated mononucleates were: 2.75±0.88 at 0.0μM, 27.25±2.30 at 0.4μM, 46.25±1.94 at 0.8μM, 98.25±7.25 at 1.6μM. To verify that no mutagen residual was transferred to recipient cultures together with the CM, we evaluated MN levels in cultures receiving the medium immediately after three washings following the chemical treatment (unconditioned medium). We further confirmed these results using a cell-mixing approach where untreated lymphocytes were co-cultured with donor cells treated with an effect-inducing dose of MMC or VCR. A distinct production pattern of both reactive oxygen species and soluble mediator proteins by treated cells may account for the differences observed in the manifestation of the bystander effect induced by VCR. In fact, we observed an increased level of ROS, IL-32 and TGF-β in the CM from VCR treated cultures, not present in MMC treated cultures. PMID:27050754

  11. T-lymphocyte induction of human monocyte angiotensin converting enzyme (ACE) is not dependent upon T-lymphocyte proliferation

    SciTech Connect

    Vuk-Pavlovic, Z.; Rohrbach, M.S.

    1986-03-05

    Human peripheral blood monocytes cultured in serum free media for seven days show a basal activity of the ectoenzyme ACE which is augmented 2-3 times by the presence of autologous peripheral blood T-lymphocytes. Since these two cell types are also involved in autologous mixed lymphocyte reaction if serum is present, the authors compared the ability of T-cells to stimulate ACE activity in the presence or absence of proliferation (measured by /sup 3/H-thymidine incorporation). By the seventh day, cultures with 5% AB/sup +/ serum showed significant increase in proliferation but no increase in ACE activity compared to the serum free cultures. Even higher proliferation rate achieved by co-culturing T-lymphocytes with allogeneic monocytes did not increase ACE production; on the contrary, ACE activity remained at the basal level. Monocyte-T-cell co-cultures stimulated with increasing concentrations of ConA or PHA showed dose dependent increases in proliferation but parallel decreases in ACE activity. Addition of soluble antigen (Candida albicans) also enhanced proliferation but not ACE synthesis. They conclude that T-lymphocyte induction of monocyte ACE is a result of cooperation between autologous cells which is not dependent upon T-cell proliferation.

  12. Revisiting the B-cell compartment in mouse and humans: more than one B-cell subset exists in the marginal zone and beyond

    PubMed Central

    2012-01-01

    The immunological roles of B-cells are being revealed as increasingly complex by functions that are largely beyond their commitment to differentiate into plasma cells and produce antibodies, the key molecular protagonists of innate immunity, and also by their compartmentalisation, a more recently acknowledged property of this immune cell category. For decades, B-cells have been recognised by their expression of an immunoglobulin that serves the function of an antigen receptor, which mediates intracellular signalling assisted by companion molecules. As such, B-cells were considered simple in their functioning compared to the other major type of immune cell, the T-lymphocytes, which comprise conventional T-lymphocyte subsets with seminal roles in homeostasis and pathology, and non-conventional T-lymphocyte subsets for which increasing knowledge is accumulating. Since the discovery that the B-cell family included two distinct categories — the non-conventional, or extrafollicular, B1 cells, that have mainly been characterised in the mouse; and the conventional, or lymph node type, B2 cells — plus the detailed description of the main B-cell regulator, FcγRIIb, and the function of CD40+ antigen presenting cells as committed/memory B-cells, progress in B-cell physiology has been slower than in other areas of immunology. Cellular and molecular tools have enabled the revival of innate immunity by allowing almost all aspects of cellular immunology to be re-visited. As such, B-cells were found to express “Pathogen Recognition Receptors” such as TLRs, and use them in concert with B-cell signalling during innate and adaptive immunity. An era of B-cell phenotypic and functional analysis thus began that encompassed the study of B-cell microanatomy principally in the lymph nodes, spleen and mucosae. The novel discovery of the differential localisation of B-cells with distinct phenotypes and functions revealed the compartmentalisation of B-cells. This review thus aims to

  13. Expression of Thomsen-Friedenreich (TF) antigens on lymphocytes. II. Loss of cryptic TF antigens during mitogenic activation of human T and B lymphocytes.

    PubMed

    Wolf, M F; Schmitt, H R; Schumacher, K

    1989-07-01

    Human peripheral blood lymphocytes (HPBL) were examined for the presence of cryptic Thomsen-Friedenreich (TF) antigens as detected by PNA or an anti-TF mAb (49H8) after neuraminidase treatment of the cell surface. Neither PNA nor the mAb bound to the cells before treatment with neuraminidase. After removal of surface sialic acid, all lymphocytes were PNA-reactive, and 85% of HPBL reacted with the mAb 49H8. Seventy-seven percent of nylon wool (NW)-eluted T cells (96% Leu 1+), 80% of enriched helper T cells (83% Leu 3a+), and 78% of suppressor/cytotoxic T cells (63% Leu 2a+) carried the cryptic TF determinant recognized by the mAb 49H8. Ninety-one percent of NW-adherent cells (68% Leu 10+, 5% Leu 1+) were also TF positive. In contrast to NW-eluted T cells which showed low to moderate mAb 49H8 binding, 48% of NW-adherent cells revealed strong binding of anti-TF mAb. With progressive activation of T cells by PHA, binding of mAb to the cryptic TF antigen completely disappeared on blast cells. The presence of TF antigens on small cells in the culture was only poorly affected. The same was observed for activation of B cells with PWM. On the other hand, binding sites for PNA did not change during blastogenesis. The disappearance of the particular, mAb 49H8-reactive TF antigen on T blast cells is not due to the loss of antigen in a distinct T cell subset, but occurs to an equal extent in the helper and suppressor/cytotoxic T cell subpopulations. Thus, the majority of peripheral T and B lymphocytes carries cryptic TF antigens. Activated T or B cell blasts, on the other hand, are deficient for the particular TF antigen detected by the mAb 49H8. We interpret these data as a modulation of certain TF antigens on effector cells in the course of lymphocyte activation. PMID:2786763

  14. Specific binding of antigen onto human T lymphocytes

    SciTech Connect

    Durandy, A.; Fischer, A.; Charron, D.; Griscelli, C.

    1986-05-01

    Human T lymphocytes sensitized to Candida albicans (CA) were shown to proliferate in cultures induced with mannan, a ramified polysaccharide extracted from the cell well of CA. We presently describe that, when we used strongly labeled (/sup 3/H)mannan, antigen-specific T blast cells were able to bind the labeled mannan on their membrane. The observations that irrelevant blast cells did not bind (/sup 3/H)mannan, and that mannan-specific blast cells did not bind tritiated pneumococcal polysaccharide SIII, indicate the specificity of mannan binding. Mannan binding was reversible and saturable. Mannan binding on T blast cells was inhibited by preincubation with monoclonal antibodies to T3 but not to other T cell-related molecules. The characteristics of this receptor suggest its identity with the T cell receptor for antigen. The direct binding of mannan could be either due to a cross-linking of the receptor by multivalent mannan or to a recognition of mannan in association with HLA-DQ molecules, as suggested by partial blocking of mannan binding using anti-HLA-DQ monoclonal antibodies.

  15. The Human Leukocyte Antigen–presented Ligandome of B Lymphocytes*

    PubMed Central

    Hassan, Chopie; Kester, Michel G. D.; de Ru, Arnoud H.; Hombrink, Pleun; Drijfhout, Jan Wouter; Nijveen, Harm; Leunissen, Jack A. M.; Heemskerk, Mirjam H. M.; Falkenburg, J. H. Frederik; van Veelen, Peter A.

    2013-01-01

    Peptides presented by human leukocyte antigen (HLA) molecules on the cell surface play a crucial role in adaptive immunology, mediating the communication between T cells and antigen presenting cells. Knowledge of these peptides is of pivotal importance in fundamental studies of T cell action and in cellular immunotherapy and transplantation. In this paper we present the in-depth identification and relative quantification of 14,500 peptide ligands constituting the HLA ligandome of B cells. This large number of identified ligands provides general insight into the presented peptide repertoire and antigen presentation. Our uniquely large set of HLA ligands allowed us to characterize in detail the peptides constituting the ligandome in terms of relative abundance, peptide length distribution, physicochemical properties, binding affinity to the HLA molecule, and presence of post-translational modifications. The presented B-lymphocyte ligandome is shown to be a rich source of information by the presence of minor histocompatibility antigens, virus-derived epitopes, and post-translationally modified HLA ligands, and it can be a good starting point for solving a wealth of specific immunological questions. These HLA ligands can form the basis for reversed immunology approaches to identify T cell epitopes based not on in silico predictions but on the bona fide eluted HLA ligandome. PMID:23481700

  16. Novel insights into the relationships between dendritic cell subsets in human and mouse revealed by genome-wide expression profiling

    PubMed Central

    Robbins, Scott H; Walzer, Thierry; Dembélé, Doulaye; Thibault, Christelle; Defays, Axel; Bessou, Gilles; Xu, Huichun; Vivier, Eric; Sellars, MacLean; Pierre, Philippe; Sharp, Franck R; Chan, Susan; Kastner, Philippe; Dalod, Marc

    2008-01-01

    Background Dendritic cells (DCs) are a complex group of cells that play a critical role in vertebrate immunity. Lymph-node resident DCs (LN-DCs) are subdivided into conventional DC (cDC) subsets (CD11b and CD8α in mouse; BDCA1 and BDCA3 in human) and plasmacytoid DCs (pDCs). It is currently unclear if these various DC populations belong to a unique hematopoietic lineage and if the subsets identified in the mouse and human systems are evolutionary homologs. To gain novel insights into these questions, we sought conserved genetic signatures for LN-DCs and in vitro derived granulocyte-macrophage colony stimulating factor (GM-CSF) DCs through the analysis of a compendium of genome-wide expression profiles of mouse or human leukocytes. Results We show through clustering analysis that all LN-DC subsets form a distinct branch within the leukocyte family tree, and reveal a transcriptomal signature evolutionarily conserved in all LN-DC subsets. Moreover, we identify a large gene expression program shared between mouse and human pDCs, and smaller conserved profiles shared between mouse and human LN-cDC subsets. Importantly, most of these genes have not been previously associated with DC function and many have unknown functions. Finally, we use compendium analysis to re-evaluate the classification of interferon-producing killer DCs, lin-CD16+HLA-DR+ cells and in vitro derived GM-CSF DCs, and show that these cells are more closely linked to natural killer and myeloid cells, respectively. Conclusion Our study provides a unique database resource for future investigation of the evolutionarily conserved molecular pathways governing the ontogeny and functions of leukocyte subsets, especially DCs. PMID:18218067

  17. Blastogenic response of human lymphocytes to antigens of Rothia dentocariosa.

    PubMed

    Fotos, P G; Gerencser, V F; Gerencser, M A

    1982-05-01

    Peripheral blood lymphocytes were isolated from 20 individuals with varying degrees of periodontal health and classified as either normal, having acute gingivitis (GV), or chronic periodontitis (PD). Crude cell wall and cytoplasmic antigens were derived from Rothia dentocariosa (RD), were applied to lymphocyte microcultures, and subjected to radioactive thymidine; the resulting lymphocyte blastogenesis (LB) was surveyed with a scintillation counter. All three groups displayed statistically similar levels of stimulation (F = 0.71), demonstrating that crude antigens of RD are not appreciably active in vitro studies of cell-mediated immunity (CMI), as measured by LB. PMID:6953091

  18. ESAM is a novel human hematopoietic stem cell marker associated with a subset of human leukemias.

    PubMed

    Ishibashi, Tomohiko; Yokota, Takafumi; Tanaka, Hirokazu; Ichii, Michiko; Sudo, Takao; Satoh, Yusuke; Doi, Yukiko; Ueda, Tomoaki; Tanimura, Akira; Hamanaka, Yuri; Ezoe, Sachiko; Shibayama, Hirohiko; Oritani, Kenji; Kanakura, Yuzuru

    2016-04-01

    Reliable markers are essential to increase our understanding of the biological features of human hematopoietic stem cells and to facilitate the application of hematopoietic stem cells in the field of transplantation and regenerative medicine. We previously identified endothelial cell-selective adhesion molecule (ESAM) as a novel functional marker of hematopoietic stem cells in mice. Here, we found that ESAM can also be used to purify human hematopoietic stem cells from all the currently available sources (adult bone marrow, mobilized peripheral blood, and cord blood). Multipotent colony-forming units and long-term hematopoietic-reconstituting cells in immunodeficient mice were found exclusively in the ESAM(High) fraction of CD34(+)CD38(-) cells. The CD34(+)CD38(-) fraction of cord blood and collagenase-treated bone marrow contained cells exhibiting extremely high expression of ESAM; these cells are likely to be related to the endothelial lineage. Leukemia cell lines of erythroid and megakaryocyte origin, but not those of myeloid or lymphoid descent, were ESAM positive. However, high ESAM expression was observed in some primary acute myeloid leukemia cells. Furthermore, KG-1a myeloid leukemia cells switched from ESAM negative to ESAM positive with repeated leukemia reconstitution in vivo. Thus, ESAM is a useful marker for studying both human hematopoietic stem cells and leukemia cells. PMID:26774386

  19. Serum BAFF and APRIL Levels, T-Lymphocyte Subsets, and Immunoglobulins after B-Cell Depletion Using the Monoclonal Anti-CD20 Antibody Rituximab in Myalgic Encephalopathy/Chronic Fatigue Syndrome.

    PubMed

    Lunde, Sigrid; Kristoffersen, Einar K; Sapkota, Dipak; Risa, Kristin; Dahl, Olav; Bruland, Ove; Mella, Olav; Fluge, Øystein

    2016-01-01

    Myalgic Encephalopathy/Chronic Fatigue Syndrome (ME/CFS) is a disease of unknown etiology. We have previously suggested clinical benefit from B-cell depletion using the monoclonal anti-CD20 antibody rituximab in a randomized and placebo-controlled study. Prolonged responses were then demonstrated in an open-label phase-II study with maintenance rituximab treatment. Using blood samples from patients in the previous two clinical trials, we investigated quantitative changes in T-lymphocyte subsets, in immunoglobulins, and in serum levels of two B-cell regulating cytokines during follow-up. B-lymphocyte activating factor of the tumor necrosis family (BAFF) in baseline serum samples was elevated in 70 ME/CFS patients as compared to 56 healthy controls (p = 0.011). There were no significant differences in baseline serum BAFF levels between patients with mild, moderate, or severe ME/CFS, or between responders and non-responders to rituximab. A proliferation-inducing ligand (APRIL) serum levels were not significantly different in ME/CFS patients compared to healthy controls at baseline, and no changes in serum levels were seen during follow-up. Immunophenotyping of peripheral blood T-lymphocyte subsets and T-cell activation markers at multiple time points during follow-up showed no significant differences over time, between rituximab and placebo groups, or between responders and non-responders to rituximab. Baseline serum IgG levels were significantly lower in patients with subsequent response after rituximab therapy compared to non-responders (p = 0.03). In the maintenance study, slight but significant reductions in mean serum immunoglobulin levels were observed at 24 months compared to baseline; IgG 10.6-9.5 g/L, IgA 1.8-1.5 g/L, and IgM 0.97-0.70 g/L. Although no functional assays were performed, the lack of significant associations of T- and NK-cell subset numbers with B-cell depletion, as well as the lack of associations to clinical responses, suggest that B

  20. Serum BAFF and APRIL Levels, T-Lymphocyte Subsets, and Immunoglobulins after B-Cell Depletion Using the Monoclonal Anti-CD20 Antibody Rituximab in Myalgic Encephalopathy/Chronic Fatigue Syndrome

    PubMed Central

    Lunde, Sigrid; Kristoffersen, Einar K.; Sapkota, Dipak; Risa, Kristin; Dahl, Olav; Bruland, Ove; Mella, Olav; Fluge, Øystein

    2016-01-01

    Myalgic Encephalopathy/Chronic Fatigue Syndrome (ME/CFS) is a disease of unknown etiology. We have previously suggested clinical benefit from B-cell depletion using the monoclonal anti-CD20 antibody rituximab in a randomized and placebo-controlled study. Prolonged responses were then demonstrated in an open-label phase-II study with maintenance rituximab treatment. Using blood samples from patients in the previous two clinical trials, we investigated quantitative changes in T-lymphocyte subsets, in immunoglobulins, and in serum levels of two B-cell regulating cytokines during follow-up. B-lymphocyte activating factor of the tumor necrosis family (BAFF) in baseline serum samples was elevated in 70 ME/CFS patients as compared to 56 healthy controls (p = 0.011). There were no significant differences in baseline serum BAFF levels between patients with mild, moderate, or severe ME/CFS, or between responders and non-responders to rituximab. A proliferation-inducing ligand (APRIL) serum levels were not significantly different in ME/CFS patients compared to healthy controls at baseline, and no changes in serum levels were seen during follow-up. Immunophenotyping of peripheral blood T-lymphocyte subsets and T-cell activation markers at multiple time points during follow-up showed no significant differences over time, between rituximab and placebo groups, or between responders and non-responders to rituximab. Baseline serum IgG levels were significantly lower in patients with subsequent response after rituximab therapy compared to non-responders (p = 0.03). In the maintenance study, slight but significant reductions in mean serum immunoglobulin levels were observed at 24 months compared to baseline; IgG 10.6–9.5 g/L, IgA 1.8–1.5 g/L, and IgM 0.97–0.70 g/L. Although no functional assays were performed, the lack of significant associations of T- and NK-cell subset numbers with B-cell depletion, as well as the lack of associations to clinical responses, suggest that B

  1. IL-36 receptor is expressed by human blood and intestinal T lymphocytes and is dose-dependently activated via IL-36β and induces CD4+ lymphocyte proliferation.

    PubMed

    Penha, Rafael; Higgins, John; Mutamba, Shilla; Barrow, Paul; Mahida, Yashwant; Foster, Neil

    2016-09-01

    We show that IL-36R is expressed by T (CD4+ and CD8+) and B (CD19+) lymphocytes in human blood and also by CD4+ T lymphocytes in the intestinal lamina propria. IL-36R protein was mostly stored in the cytoplasm of CD4 lymphocytes and B cells, during steady state conditions and the greatest expression of IL-36R mRNA was measured in CD4+ (T helper) lymphocytes. IL-36 β, which functions via IL-36R induced rapid and significant (P<0.05) proliferation of CD4+ lymphocytes, within 48h. IL-36R expression was also maintained on the surface of circulating CD4+ lymphocytes which enter the intestinal lamina propria. In conclusion our study is the first to show that (1) all human blood lymphocytes express IL-36R; (2) IL-36R expression is maintained by circulating CD4+ lymphocytes which enter the intestinal lamina propria and (3) IL-36R/IL-36 β induces rapid CD4 lymphocyte proliferation. The possible significance of these results in the context of human disease is discussed. PMID:27269181

  2. Human Herpesvirus 8 Induces Polyfunctional B Lymphocytes That Drive Kaposi’s Sarcoma

    PubMed Central

    Knowlton, Emilee R.; Rappocciolo, Giovanna; Piazza, Paolo; Lepone, Lauren M.; Nadgir, Sagar V.; Bullotta, Arlene; Berendam, Stella J.; Li, Jun; Reinhart, Todd A.; Jenkins, Frank J.

    2014-01-01

    ABSTRACT Kaposi’s sarcoma (KS) is an unusual neoplasia wherein the tumor consists primarily of endothelial cells infected with human herpesvirus 8 (HHV-8; Kaposi’s sarcoma-associated herpesvirus) that are not fully transformed but are instead driven to excess proliferation by inflammatory and angiogenic factors. This oncogenic process has been postulated but unproven to depend on a paracrine effect of an abnormal excess of host cytokines and chemokines produced by HHV-8-infected B lymphocytes. Using newly developed measures for intracellular detection of lytic cycle proteins and expression of cytokines and chemokines, we show that HHV-8 targets a range of naive B cell, IgM memory B cell, and plasma cell-like populations for infection and induction of interleukin-6, tumor necrosis factor alpha, macrophage inhibitory protein 1α, macrophage inhibitory protein 1β, and interleukin-8 in vitro and in the blood of HHV-8/HIV-1-coinfected subjects with KS. These B cell lineage subsets that support HHV-8 infection are highly polyfunctional, producing combinations of 2 to 5 of these cytokines and chemokines, with greater numbers in the blood of subjects with KS than in those without KS. Our study provides a new paradigm of B cell polyfunctionality and supports a key role for B cell-derived cytokines and chemokines produced during HHV-8 infection in the development of KS. PMID:25182322

  3. Human Lymphocytes Response to Low Gamma-ray Doses

    NASA Astrophysics Data System (ADS)

    Vega-Carrillo, Héctor René; Manzanares-Acuña, Eduardo; Bañuelos-Valenzuela, Rómulo

    2002-08-01

    Radiation and non-radiation workers lymphocytes were exposed to a low strength gamma-ray field to determine heat shock protein expression in function of radiation dose. Protein identification was carried out using mAb raised against Hsp25, Hsp60, Hsp70 and Hsp90; from these, only Hsp70 protein was detected before and after lymphocyte irradiation. In all cases, an increasing trend of relative amounts of Hsp70 in function to irradiation time was observed. After 70.5 uGy gamma-ray dose, radiation worker's lymphocytes expressed more Hsp70 protein, than non radiation workers' lymphocytes, indicating a larger tolerance to gamma rays (gammatolerance), due to an adaptation process developed by his labor condition.

  4. A Single-Cell Gene-Expression Profile Reveals Inter-Cellular Heterogeneity within Human Monocyte Subsets

    PubMed Central

    Gren, Susanne T.; Rasmussen, Thomas B.; Janciauskiene, Sabina; Håkansson, Katarina; Gerwien, Jens G.; Grip, Olof

    2015-01-01

    Human monocytes are a heterogeneous cell population classified into three different subsets: Classical CD14++CD16-, intermediate CD14++CD16+, and non-classical CD14+CD16++ monocytes. These subsets are distinguished by their differential expression of CD14 and CD16, and unique gene expression profile. So far, the variation in inter-cellular gene expression within the monocyte subsets is largely unknown. In this study, the cellular variation within each human monocyte subset from a single healthy donor was described by using a novel single-cell PCR gene-expression analysis tool. We investigated 86 different genes mainly encoding cell surface markers, and proteins involved in immune regulation. Within the three human monocyte subsets, our descriptive findings show multimodal expression of key immune response genes, such as CD40, NFⱪB1, RELA, TLR4, TLR8 and TLR9. Furthermore, we discovered one subgroup of cells within the classical monocytes, which showed alterations of 22 genes e.g. IRF8, CD40, CSF1R, NFⱪB1, RELA and TNF. Additionally one subgroup within the intermediate and non-classical monocytes also displayed distinct gene signatures by altered expression of 8 and 6 genes, respectively. Hence the three monocyte subsets can be further subdivided according to activation status and differentiation, independently of the traditional classification based on cell surface markers. Demonstrating the use and the ability to discover cell heterogeneity within defined populations of human monocytes is of great importance, and can be useful in unravelling inter-cellular variation in leukocyte populations, identifying subpopulations involved in disease pathogenesis and help tailor new therapies. PMID:26650546

  5. Experimental Study on Effect of Simulated Microgravity on Structural Chromosome Instability of Human Peripheral Blood Lymphocytes

    PubMed Central

    Wei, Lijun; Liu, Chuanpeng; Kang, Li; Liu, Yufeng; Shi, Shuliang; Wu, Qiong; Li, Yu

    2014-01-01

    Experimental study was made by keeping human peripheral blood lymphocytes under simulated microgravity in a Rotary Cell Culture System bioreactor to investigate the changes that occur in the number of chromosomes, the expression rate of chromosome fragile site, and the expressions of DNA replication- and repair-related genes. Experimental results indicate simulated microgravity has no effect on the numerical chromosome instability of human peripheral blood lymphocytes, but it enhances the structural chromosome instability of human peripheral blood lymphocytes through the inhibition of DNA replication and the reduction of DNA repair. So, the mechanism of chromosome fragile site induced by simulated microgravity can be explained using the changes that occur in the chromosome structure of human peripheral blood lymphocytes, the DNA replication and repair under the effect of simulated microgravity. PMID:24963972

  6. Experimental study on effect of simulated microgravity on structural chromosome instability of human peripheral blood lymphocytes.

    PubMed

    Wei, Lijun; Liu, Chuanpeng; Kang, Li; Liu, Yufeng; Shi, Shuliang; Wu, Qiong; Li, Yu

    2014-01-01

    Experimental study was made by keeping human peripheral blood lymphocytes under simulated microgravity in a Rotary Cell Culture System bioreactor to investigate the changes that occur in the number of chromosomes, the expression rate of chromosome fragile site, and the expressions of DNA replication- and repair-related genes. Experimental results indicate simulated microgravity has no effect on the numerical chromosome instability of human peripheral blood lymphocytes, but it enhances the structural chromosome instability of human peripheral blood lymphocytes through the inhibition of DNA replication and the reduction of DNA repair. So, the mechanism of chromosome fragile site induced by simulated microgravity can be explained using the changes that occur in the chromosome structure of human peripheral blood lymphocytes, the DNA replication and repair under the effect of simulated microgravity. PMID:24963972

  7. Phenotypic and functional consequences of herpesvirus saimiri infection of human CD8+ cytotoxic T lymphocytes.

    PubMed Central

    Berend, K R; Jung, J U; Boyle, T J; DiMaio, J M; Mungal, S A; Desrosiers, R C; Lyerly, H K

    1993-01-01

    Herpesvirus saimiri (HVS) was used to infect and transform human CD8+ cytotoxic T lymphocytes (CTL), and the phenotypic and functional consequences of HVS infection of CD8+ T lymphocytes were investigated. HVS-transformed CTL no longer require antigen restimulation yet maintain their phenotype and HLA-restricted cytolytic function and specificity. The ability of HVS to transform CTL may have an important role in the functional analysis of human antigen-specific CTL. Images PMID:8396687

  8. Identifying developmental toxicity pathways for a subset of ToxCast chemicals using human embryonic stem cells and metabolomics

    EPA Science Inventory

    Metabolomics analysis was performed on the supernatant of human embryonic stem (hES) cell cultures exposed to a blinded subset of 11 chemicals selected from the chemical library of EPA's ToxCast™ chemical screening and prioritization research project. Metabolites from hES cultur...

  9. A global look into human T cell subsets before and after cryopreservation using multiparametric flow cytometry and two-dimensional visualization analysis.

    PubMed

    Lemieux, Jennifer; Jobin, Christine; Simard, Carl; Néron, Sonia

    2016-07-01

    The cryopreservation of human lymphocytes is an essential step for the achievement of several cellular therapies. Besides, T cells are considered as promising actors in cancer therapy for their cytotoxic and regulatory properties. Consequently, the development of tools to monitor the impact of freezing and thawing processes on their fine distribution may be an asset to achieve quality control in cellular therapy. In this study, the phenotypes of freshly isolated human mononuclear cells were compared to those observed following one cycle of cryopreservation and rest periods 0h, 1h and 24h after thawing but before staining. T cells were scrutinized for their distribution according to naive, memory effector, regulatory and helper subsets. Flow cytometry analyses were done using eight-color antibody panels as proposed by the Human Immunophenotyping Consortium. Data were further analyzed by using conventional directed gating and clustering software, namely SPADE and viSNE. Overall, SPADE and viSNE tools were very efficient to monitor the outcome of PBMC populations and T cell subsets. T cells were more sensitive to cryopreservation than other cells. Our results indicated that submitting the thawed cells to a 1h rest period improved the detection of some cell markers when compared to fresh samples. In contrast, cells submitted to a 24h rest period, or to none, were less representative of fresh sample distribution. The heterogeneity of PBMC, as well as the effects of freeze-thaw cycle on their distribution, can be easily monitored by using SPADE and viSNE. PMID:27129808

  10. Signaling in Human and Murine Lymphocytes in Microgravity: Parallels and Contrasts

    NASA Technical Reports Server (NTRS)

    Neal, Pellis; Alamelu, Sundaresan; Kulkarni, A. D.; Yamauchi, K.

    2006-01-01

    Immune function in space undergoes dramatic changes, some of which are detrimental to lymphocyte function. These changes may lead to significant immune suppression. Studies with human lymphocytes both in space flight and with ground-based models (NASA in vitro ground-based microgravity analog) indicate that T cell activation is inhibited in microgravity. Other lymphocyte functions, such as locomotion, are also inhibited. There is about an 80 percent homology in the immune response of mice to that of humans. A murine model was investigated because of its ability to parallel some microgravity using hind limb suspension. In in vivo antiorthostatically (AOS)-suspended mice, T cell activation is greatly suppressed, with the majority of activation related cytokines being inhibited. PHA activation in lymphocytes derived from AOS mice (in vivo ground-based microgravity analog) is also suppressed. Calcium ionophore studies in human lymphocytes exposed to modeled microgravity indicate that the calcium pathways are probably unaffected in microgravity. IP3 (inositol triphosphate) receptor expression in both human and mouse lymphocytes cultured in modeled microgravity indicate no suppression of calcium signaling. In the human system, microgravity seems to inhibit signaling cascades either at the level of, or up-stream of, Protein Kinase C (PKC). In particular, a membrane event, such as phospholipase C gamma 1 activity in human lymphocytes is affected, with its direct upstream effector, LAT, being deficiently expressed. In the mouse pathway, LAT is undiminished while another critical intermediate, SLP-76, is diminished significantly. This study identifies critical stages in the human and mouse immune systems and in lymphocytes as a function of microgravity.

  11. Cryopreservation of human lymphocytes: a brief review and evaluation of an automated liquid nitrogen freezer.

    PubMed

    Glassman, A B; Bennett, C E

    1979-01-01

    Successful cryopreservation of human lymphocytes has been previously described. Cryopreserved lymphocytes are useful for a variety of in vitro immunologic studies. This study was performed to determine the applicability and/or advantages of using a programmable freezing system, and compares glycerol versus dimethyl sulfoxide (DMSO) at varying concentrations on post-thaw viability, E-rosetting and immunoglobulin fluorescence. Prefreeze T and B lymphocyte percentages were determined. Cells were then frozen in varying concentrations of glycerol and DMSO. Optimum cryoprotectant type and concentration was determined. Lymphocytes from seven individuals were frozen by the batch method in a mechanical freezer and with the automated liquid nitrogen injection system. Data on post-thaw T and B percentages and viability revealed 10% DMSO and liquid nitrogen control freezing method at 1 C/minute as the best conditions for lymphocyte preservation as reflected by post-thaw in vitro testing. PMID:373178

  12. Analysis of the in vitro effect of exogenous nitric oxide on human lymphocytes.

    PubMed

    Shoker, A S; Yang, H; Murabit, M A; Jamil, H; al-Ghoul, A; Okasha, K

    1997-06-01

    We investigated the role of endogenous or exogenous nitric oxide (NO) on human lymphocyte function. We used sodium nitroprusside, nitroglycerine, S-nitroso-N-acetylpenicillamine, sodium nitrite and S-nitroso-L-glutathione as NO-generating compounds. All agents were used at doses that do not produce direct cytotoxicity as measured by trypan blue exclusion as well as chromium-51 release assay. The immune responses examined were peripheral blood lymphocytes (PBL) proliferation and IL-2 production after activation with OKT3 and PHA; allogeneic mediated proliferation and cell mediated cytotoxicity (CML) in MLR; IgG and IgM production after PBL activation with Con-A; proliferation and expression of IFN-gamma and IL-4 mRNA after activation of allogeneic CD4+T cell clones. Cytokine mRNA expression was measured by reverse transcriptase PCR. Our results show that proliferating lymphocytes do not produce a detectable amount of NO as measured by the Griess reaction. In separate experiments, the addition of NG-monomethyl-L-arginine (L-NMMA) did not affect lymphocyte proliferation. Sodium nitroprusside and nitroglycerine exerted a dose dependent antimitogenic effect, inhibited cytokine production and expression, CML generation and antibody production. DNA gel electrophoresis showed no evidence for enhanced programmed cell death. The antimitogenic effect could not be blocked by the NO scavengers, hemoglobin or methylene blue. In contrast, the other nitric oxide generating compounds did not inhibit lymphocyte mitogenesis. The results suggest that human lymphocytes do not produce appreciable amounts of NO to affect lymphocyte mitogenesis. Sodium nitroprusside and nitroglycerine have a potent but nonspecific immunoinhibitory effect on human lymphocyte function by a mechanism other than NO production. In addition, pharmacological levels of NO do not inhibit human lymphocyte mitogenesis. PMID:9201699

  13. Diverse Gene Expression in Human Regulatory T Cell Subsets Uncovers Connection between Regulatory T Cell Genes and Suppressive Function.

    PubMed

    Hua, Jing; Davis, Scott P; Hill, Jonathan A; Yamagata, Tetsuya

    2015-10-15

    Regulatory T (Treg) cells have a critical role in the control of immunity, and their diverse subpopulations may allow adaptation to different types of immune responses. In this study, we analyzed human Treg cell subpopulations in the peripheral blood by performing genome-wide expression profiling of 40 Treg cell subsets from healthy donors. We found that the human peripheral blood Treg cell population is comprised of five major genomic subgroups, represented by 16 tractable subsets with a particular cell surface phenotype. These subsets possess a range of suppressive function and cytokine secretion and can exert a genomic footprint on target effector T (Teff) cells. Correlation analysis of variability in gene expression in the subsets identified several cell surface molecules associated with Treg suppressive function, and pharmacological interrogation revealed a set of genes having causative effect. The five genomic subgroups of Treg cells imposed a preserved pattern of gene expression on Teff cells, with a varying degree of genes being suppressed or induced. Notably, there was a cluster of genes induced by Treg cells that bolstered an autoinhibitory effect in Teff cells, and this induction appears to be governed by a different set of genes than ones involved in counteracting Teff activation. Our work shows an example of exploiting the diversity within human Treg cell subpopulations to dissect Treg cell biology. PMID:26371251

  14. Clinically used selective oestrogen receptor modulators increase LDL receptor activity in primary human lymphocytes

    PubMed Central

    Cerrato, F; Fernández-Suárez, M E; Alonso, R; Alonso, M; Vázquez, C; Pastor, O; Mata, P; Lasunción, M A; Gómez-Coronado, D

    2015-01-01

    Background and Purpose Treatment with selective oestrogen receptor modulators (SERMs) reduces low-density lipoprotein (LDL) cholesterol levels. We assessed the effect of tamoxifen, raloxifene and toremifene and their combinations with lovastatin on LDL receptor activity in lymphocytes from normolipidaemic and familial hypercholesterolaemic (FH) subjects, and human HepG2 hepatocytes and MOLT-4 lymphoblasts. Experimental Approach Lymphocytes were isolated from peripheral blood, treated with different compounds, and 1,1′-dioctadecyl-3,3,3,3′-tetramethylindocarbocyanine perchlorate (DiI)-labelled LDL uptake was analysed by flow cytometry. Key Results Tamoxifen, toremifene and raloxifene, in this order, stimulated DiI-LDL uptake by lymphocytes by inhibiting LDL-derived cholesterol trafficking and subsequent down-regulation of LDL receptor expression. Differently to what occurred in HepG2 and MOLT-4 cells, only tamoxifen consistently displayed a potentiating effect with lovastatin in primary lymphocytes. The SERM-mediated increase in LDL receptor activity was not altered by the anti-oestrogen ICI 182 780 nor was it reproduced by 17β-oestradiol. However, the tamoxifen-active metabolite endoxifen was equally effective as tamoxifen. The SERMs produced similar effects on LDL receptor activity in heterozygous FH lymphocytes as in normal lymphocytes, although none of them had a potentiating effect with lovastatin in heterozygous FH lymphocytes. The SERMs had no effect in homozygous FH lymphocytes. Conclusions and Implications Clinically used SERMs up-regulate LDL receptors in primary human lymphocytes. There is a mild enhancement between SERMs and lovastatin of lymphocyte LDLR activity, the potentiation being greater in HepG2 and MOLT-4 cells. The effect of SERMs is independent of oestrogen receptors but is preserved in the tamoxifen-active metabolite endoxifen. This mechanism may contribute to the cholesterol-lowering action of SERMs. PMID:25395200

  15. The human peripheral lymph node vascular addressin. An inducible endothelial antigen involved in lymphocyte homing.

    PubMed Central

    Michie, S. A.; Streeter, P. R.; Bolt, P. A.; Butcher, E. C.; Picker, L. J.

    1993-01-01

    The extravasation of blood-borne lymphocytes into organized lymphoid tissues and sites of chronic inflammation is directed in part by interactions of lymphocyte surface adhesion molecules, known as homing receptors, with tissue-selective endothelial ligands called vascular addressins. In mice and humans, lymphocyte L-selectin and the peripheral lymph node addressin (PNAd) form a homing receptor-endothelial ligand pair involved in lymphocyte traffic to peripheral lymph node (PLN). We have examined the tissue distribution and function of human PNAd, using monoclonal antibody MECA-79 and in vitro assays of L-selectin-dependent lymphocyte binding. We demonstrate that PNAd is expressed by human high endothelial venules (HEV) in lymphoid tissues which support lymphocyte adhesion via a PLN-associated recognition system. MECA-79 inhibits adhesion to these HEV of a cell line that binds predominantly via the PLN-homing receptor, L-selectin, but has no effect on adhesion by a mucosal HEV-binding cell line. Furthermore, MECA-79 blocks binding of human peripheral blood mononuclear cells to both PLN and tonsil HEV, but not significantly to HEV in the appendix. In addition, we demonstrate PNAd induction on venules at chronic inflammatory sites in humans, particularly sites with severe or long-standing chronic inflammatory involvement. These results confirm that PNAd functions as a PLN vascular addressin in humans, and that in addition to directing normal lymphocyte recirculation to lymph nodes and tonsils, this addressin likely participates in lymphocyte recruitment to sites of chronic inflammation. Images Figure 1 Figure 4 PMID:8256856

  16. Regulation of IgA secretion in polyclonally induced in vitro human lymphocyte cultures: the function of T and B cells from mesenteric lymph nodes and peripheral blood.

    PubMed Central

    Pang, G; Yeung, S; Clancy, R L; Cripps, A W; Hennessy, E J; Santhanam, A N

    1986-01-01

    Human gut-associated immunoregulatory events were studied in a pokeweed mitogen (PWM)-stimulated culture system using lymphocytes obtained from the mesenteric lymph nodes (MLN) of female subjects undergoing gastroplasty for obesity. Compared with peripheral blood lymphocytes, lymphocytes obtained from MLN secreted IgG, IgA and IgM isotypes that differ in pattern and distribution despite similar proportions of T cells and B cells expressing isotype-specific surface membrane immunoglobulin (SmIg). Among the isotypes secreted, IgA appeared to be increased relatively to other isotypes in MLN cultures. Crossover coculture experiments using T and B cells isolated from both MLN and blood by E-rosetting and cell panning procedures demonstrated that IgA was particularly sensitive to help and suppression exerted by MLN T cells and T cell subsets defined by monoclonal antibodies OKT4 and OKT8 respectively, when compared with similar subsets isolated from blood. The results presented provide a basis for study of gut handling of ingested antigen in man, and of disturbed immunoregulatory events in inflammatory and neoplastic disease of the human gut. PMID:2942320

  17. Divergence of human and nonhuman primate lymphocyte responses to bacterial superantigens.

    PubMed

    Bavari, S; Hunt, R E; Ulrich, R G

    1995-09-01

    We compared T cell responses of human, rhesus monkey (Macaca mulatta), and chimpanzee (Pan troglodytes) to four bacterial superantigens. When lymphocytes were cultured in media supplemented with species-specific sera, chimpanzee T cells were stimulated by lower doses of staphylococcal enterotoxin (SE) A and toxic shock syndrome toxin 1 (TSST1) than were human T cells, while chimpanzee responses to SEB and SEC1 were nearly equivalent to the human response. Interestingly, rhesus lymphocytes responded to 10,000 times lower amounts of SEA, SEB, and SEC1 and to 100 times lower concentrations of TSST1 than human cells. The greater sensitivity of rhesus T cells to these toxins was not a result of differences in class II binding affinities and was only partly attributable to the presence of anti-SE and TSST1 antibodies in human serum. These results suggest that rhesus T lymphocytes are more sensitive toward these bacterial superantigens than human T cells. PMID:7554446

  18. High-Efficiency Transfection of Primary Human and Mouse T Lymphocytes Using RNA Electroporation

    PubMed Central

    Zhao, Yangbing; Zheng, Zhili; Cohen, Cyrille J.; Gattinoni, Luca; Palmer, Douglas C.; Restifo, Nicholas P.; Rosenberg, Steven A.; Morgan, Richard A.

    2006-01-01

    The use of nonviral gene transfer methods in primary lymphocytes has been hampered by low gene transfer efficiency and high transfection-related toxicity. In this report, high gene transfection efficiency with low transfection-related toxicity was achieved by electroporation using in vitro-transcribed mRNA. Using these methods, >90% transgene expression with >80% viable cells was observed in stimulated primary human and murine T lymphocytes transfected with GFP or mCD62L. Electroporation of unstimulated human PBMCs or murine splenocytes with GFP RNA yielded 95 and 56% GFP+ cells, respectively. Electroporation of mRNA for NY-ESO-1, MART-1, and p53 antigen-specific TCRs into human T lymphocytes redirected these lymphocytes to recognize melanoma cell lines in an MHC-restricted manner. The onset of gene expression was rapid (within 30 min) and durable (up to 7 days postelectroporation) using both GFP and TCR-mediated recognition of target cells. There was no adverse effect observed on the T lymphocytes subjected to RNA electroporation evaluated by cell growth rate, annexin-V staining of apoptotic cells, BrdU incorporation, tumor antigen-specific recognition or antigen-specific TCR affinity. The results of this study indicate that mRNA electroporation provides a powerful tool to introduce genes into both human and murine primary T lymphocytes. PMID:16140584

  19. Migration of human lymphocytes. I. A model using the mouse as host.

    PubMed Central

    Morgan, K; Holt, P J

    1978-01-01

    The distribution of radioactivity after the intravenous injection of 51Cr-labelled human lymphocytes has been examined in normal mice, irradiated mice, mice treated with anti-platelet antiserum and in mice treated with colloidal carbon. Pre-treatment with carbon and anti-platelet antiserum appears to protect the human lymphocytes from uptake by the host's reticuloendothelial system (RES). Comparison of tissue radioactivity in carbon-treated mice after the injection of viable human lymphocytes with that found after the injection of dead cells and soluble or insoluble cell debris showed that radioactivity recovered in the spleen and lymph nodes is primarily due to the migration of viable lymphocytes into these tissues. Thus the measurement of radioactivity in lymph nodes of carbon-treated mice after the injection of 51Cr-labelled human lymphocytes can be used as a model of these lymphocytes' ability to migrate into the lymph nodes during recirculation and to study factors influencing this migration. PMID:721139

  20. Radioprotective effect of mefenamic acid against radiation-induced genotoxicity in human lymphocytes

    PubMed Central

    Nobakht, Reyhaneh; Ghasemi, Arash; Pourfallah, Tayyeb Allahverdi

    2015-01-01

    Purpose Mefenamic acid (MEF) as a non-steroidal anti-inflammatory drug is used as a medication for relieving of pain and inflammation. Radiation-induced inflammation process is involved in DNA damage and cell death. In this study, the radioprotective effect of MEF was investigated against genotoxicity induced by ionizing radiation in human blood lymphocytes. Materials and Methods Peripheral blood samples were collected from human volunteers and incubated with MEF at different concentrations (5, 10, 50, or 100 µM) for two hours. The whole blood was exposed to ionizing radiation at a dose 1.5 Gy. Lymphocytes were cultured with mitogenic stimulation to determine the micronuclei in cytokinesis blocked binucleated lymphocyte. Results A significant decreasing in the frequency of micronuclei was observed in human lymphocytes irradiated with MEF as compared to irradiated lymphocytes without MEF. The maximum decreasing in frequency of micronuclei was observed at 100 µM of MEF (38% decrease), providing maximal protection against ionizing radiation. Conclusion The radioprotective effect of MEF is probably related to anti-inflammatory property of MEF on human lymphocytes. PMID:26484310

  1. In vitro characteristics on human lymphocyte functions of a new immunomodulatory agent, a cyclic peptide, cyclomunine.

    PubMed Central

    Niaudet, P; Beaurain, G; Leibowitch, J; Bach, J F

    1980-01-01

    Cyclomunine, a cyclic peptide extracted from Fusarium equisiti, inhibits responses of human lymphocytes to mitogens, soluble antigens and allogeneic cells and the proliferation of lymphoblastoid cell lines. Cyclomunine has little effect on small lymphocytes but acts rather on lymphoblasts. It has no effect on fibroblasts and myeloid cells. Cyclomunine partially inhibits the generation of suppressor cells induced by Con A and the generation of cytotoxic T cells in a mixed lymphocyte culture and totally inhibits the in vitro synthesis of Ig by PBL. Cyclomunine merits consideration as a new in vitro anti-lymphoblastic agent. PMID:6451339

  2. Recognition of Major Histocompatibility Complex Antigens on Cultured Human Biliary Epithelial Cells by Alloreactive Lymphocytes

    PubMed Central

    Saidman, Susan L.; Duquesnoy, Rene J.; Zeevi, Adriana; Fung, John J.; Starzl, Thomas E.; Demetris, A. Jake

    2010-01-01

    We have developed an in vitro system to study the interactions between biliary epithelium and lymphocytes using cultured human biliary epithelial cells. No class II antigens were detected by immunoperoxidase staining of the normal biliary epithelial cells, but alloactivated lymphocyte culture supernatants were able to induce class II expression. The activity of the supernatants was blocked with an anti-γ-interferon monoclonal antibody. In addition, recombinant human γ-interferon alone induced the expression of class II antigens and increased the intensity of class I staining of cultured biliary epithelial cells. Biliary epithelial cell–induced proliferation of alloreactive T lymphocytes demonstrated that the major histocompatibility complex molecules carry functional lymphocyte-activating determinants. The recognition of major histocompatibility complex determinants was confirmed by monoclonal antibody–blocking studies and by stimulation of an alloreactive T-cell clone. However, the biliary epithelial cells were much less potent stimulators than arterial endothelial cells tested in the same assay system. PMID:1704868

  3. Cytotoxic human lymphocytes: from in vitro testing (1970s) to immunotherapy (1990s).

    PubMed

    Sinkovics, J; Horvath, J

    1993-01-01

    The senior author was the recipient of a contract (1-CP3-3292) from the National Cancer Institute, USA (NCI) in the early 1970s. The aim of NCI's targeted research program was the establishment of a tumour-specific human lymphocyte-mediated cytotoxicity assay. Neither lymphocyte growth factors nor monoclonal antibodies for lymphocyte typing were available. Tumour-specific populations of lymphocytes could not be maintained but their presence in ficoll-hypaque preparations of blood buffy coats or in primary cultures of tumours was clearly recognized. Another indiscriminately cytotoxic population of lymphocytes had usually overridden the tumour-specific population. In contradistinction to the ruling doctrine of the era, indiscriminately cytotoxic lymphocytes were readily found in the blood of tumour-bearing patients and healthy individuals (the senior author's lymphocytes were shown to practice indiscriminate cytotoxicity in 1971, an observation first interpreted as "immune surveillance at work" in an individual daily exposed to patients with metastatic cancers). Instead of converting the subject matter of the contract from a tumour-specific to a non-specific cytotoxicity assay, the NCI prematurely "phased it out" (but continued the project as intramural research). Nevertheless, many functions of cytotoxic lymphocytes that had become by now well established were foreshadowed during the early 1970s with the limited support of that NCI contract and funds from other sources. Here we recount those early observations; present the outlines of adoptive immunotherapy with various autologous lymphocyte populations and in a separate report in this volume give a technical description how these lymphocyte populations are prepared in the laboratory for therapeutic reinfusions into the patient. PMID:8191863

  4. Identification and characterization of human dendritic cell subsets in the steady state: a review of our current knowledge.

    PubMed

    Patel, Vineet Indrajit; Metcalf, Jordan Patrick

    2016-04-01

    Dendritic cells (DC) are generally categorized as a group of rare antigen presenting cells that are to the crucial development of immune responses to pathogens and also of tolerance to self-antigens. Therefore, having the ability to identify DC in specific tissues and to test their functional abilities in the steady state are scientific gaps needing attention. Research on primary human DC is lacking due to their rarity and the difficulty of obtaining tissue samples. However, recent findings have shown that several different DC subsets exist, and that these subsets vary both by markers expressed and functions depending on their specific microenvironment. After discriminating from other cell types, DC can be split into myeloid and plasmacytoid fractions. While plasmacytoid DC express definite markers, CD123 and BDCA-2, myeloid DC encompass several different subsets with overlapping markers expressed. Such markers include the blood DC antigens BDCA-1 and BDCA-3, along with Langerin, CD1a and CD14. Marker specificity is further reduced when accounting for microenvironmental differences, as observed in the blood, primary lymphoid tissues, skin and lungs. The mixed leukocyte reaction (MLR) has been used to measure the strength of antigen presentation by specific DC subsets. Surface markers and MLR require standardization to enable consistent identification of and comparisons between DC subsets. To alleviate these issues, researchers have begun comparing DC subsets at the transcriptional level. This has allowed degrees of relatedness to be determined between DC in different microenvironments, and should be a continued area of focus in years to come. PMID:26956785

  5. Establishment of a stable, inducible form of human immunodeficiency virus type 1 DNA in quiescent CD4 lymphocytes in vitro.

    PubMed Central

    Spina, C A; Guatelli, J C; Richman, D D

    1995-01-01

    Human immunodeficiency virus type 1 (HIV-1) possesses the ability to establish a complete infection in nondividing host cells. The capacity of HIV-1 to infect nondividing cells probably contributes significantly to its pathology in vivo, as reflected by infection of peripheral T lymphocytes, tissue macrophages, and microglial cells. However, the in vitro demonstration of the establishment of stable HIV-1 infection in quiescent T cells remains controversial. We have developed a primary T-cell model of acute HIV-1 infection of quiescent CD4 lymphocytes that demonstrates the development of a complete, reverse-transcribed form of virus that is stable for over 10 days in culture. To ensure that our primary cell culture was representative of a quiescent population, the CD4 lymphocyte targets were monitored for membrane expression of activation antigens and for shifts in cell cycle from G0/G1 to S/G2 phase. The presence of viral DNA fragments reflecting progressive reverse transcription was determined by PCR analysis. HIV entered primary CD4 cells rapidly, but viral DNA accumulated slowly in the resting cell cultures. DNA species containing regions of full-length reverse transcription were not detected until 3 to 5 days after infection. In parallel with the appearance of complete viral DNA, spliced RNA transcripts, predominantly of the nef species, were detected by reverse transcriptase PCR amplification. When infected CD4 cells were sorted on the basis of cell cycle analysis of DNA content, the accumulation of a complete viral DNA form was found to occur in both the purified G0/G1-phase cell subset and the cell fraction enriched for the minor S-phase subset. In contrast, spliced viral RNA products could be detected only in the enriched S-phase cell fraction. These results demonstrate that HIV-1 can infect and establish a complete, stable form of viral DNA in primary CD4 lymphocytes in vitro but is blocked from transcription in the absence of cell activation. The findings

  6. [Method for determining dopamine and morphine binding sites in lymphocytes from human peripheral blood].

    PubMed

    Gamaleia, N B; Kuz'mina, T I; Shostak, O A; Gamaleia, A A; Dmitrieva, I G

    2003-12-01

    A histochemical method was designed to detect the regions of binding the dopamine and morphine in human peripheral blood lymphocytes. It is based on incubating the suspension of lymphocytes and conjugated dopamine or morphine with bull serum albumin (BSA) marked by horse-radish peroxidase. After incubation, smears are prepared from the lymphocyte suspension, which are stained by diaminobenzidine in the presence of hydrogen peroxide for peroxidase. The light microscope with oil immersion is used to count the number of lymphocytes (from among 100 hundred of them), which contain the peroxidase granules. Smears from the lymphocyte suspension, which were incubated with the BSA-peroxidase conjugate, were controls. The binding of peroxidase-marked ligands of dopamine and mu-opioid receptors with lymphocytes was oppressed by the dose-dependant preliminary incubation with antagonists (haloperidol, naloxone), on the basis of which the presence of the ligand-receptor interaction can be suggested. The number of bindings of dopamine and morphine in lymphocytes was shown to be reliably higher in the alcoholic-intoxication state versus the healthy subjects without any signs of alcohol consumption. The designed method is simple enough in use and does not require any special equipment for the receptor detection in a moderate blood quantity. PMID:14971325

  7. Toxicity of methyl tertiary-butyl ether on human blood lymphocytes.

    PubMed

    Salimi, Ahmad; Vaghar-Moussavi, Mehrdad; Seydi, Enayatollah; Pourahmad, Jalal

    2016-05-01

    Methyl tertiary-butyl ether (MTBE) is a synthetic solvent widely used as oxygenate in unleaded gasoline. Few studies have addressed the cellular toxicity of MTBE on some cell lines, and so far, no comprehensive study has been conducted to investigate the probable immunotoxicity of this compound. In this study, the toxicity of MTBE on human blood lymphocytes was evaluated. Blood lymphocytes were isolated from healthy male volunteers' blood, using Ficoll polysaccharide followed by gradient centrifugation. Cell viability, reactive oxygen species (ROS) formation, lipid peroxidation, glutathione levels, and damage to mitochondria and lysosome were determined in blood lymphocytes after 6-h incubation with different concentrations of MTBE (0.1, 0.5, 1, and 2 mM). Our results showed that MTBE, in particular, decreased cell viability, which was associated with significant increase at intracellular ROS level and toxic alterations in mitochondria and lysosomes in human blood lymphocytes. Moreover, it was shown that MTBE strongly provoked lipid peroxidation and also depleted glutathione level at higher concentrations. Interestingly, MTBE exhibited its cytotoxic effects at low concentrations that may resemble to its concentrations in human blood following occupational and environmental exposure. It is therefore concluded that MTBE was capable of inducing oxidative stress and damage to mitochondria and lysosomes in human lymphocytes at concentrations ranging from 5 to 40 μg/L, which may be present in human blood as a result of environmental exposure. PMID:26797945

  8. Multicolor flow cytometry analysis of blood cell subsets in patients given total body irradiation before bone marrow transplantation

    SciTech Connect

    Clave, E.; Socie, G.; Carosella, E.

    1995-11-01

    Bone marrow transplantation has often been closely linked with accidental or intentional therapeutical irradiation. In both situations, study of the radiosensitivity of human blood cell subsets is of interest. Using one-color flow cytometry analysis of B lymphocytes, T cell subsets, and natural killer cells, we previously reported that lymphocyte subsets exhibit equal radiosensitivity. Taking advantage of recent developments in the knowledge of leukocyte differentiation antigens and flow cytometry technology we undertook a study of blood cell subsets to search for rare populations exhibiting different radiosensitivity. Thirty patients, who were delivered a 12 Gy fractionated total body irradiation as part of their conditioning regimen before transplantation for malignant disorders, were studied using multicolor flow cytometry. T and B lymphocytes showed a sharp, radiation-induced decrease, with the B lymphocytes (cluster of differentiation (CD) 19+) being the most sensitive. When analyzed by multicolor flow cytometry all major lymphocyte subsets appeared equally sensitive to the in vivo irradiation. Therefore, all major lymphocyte subsets sharing the helper phenotype (naive or memory) and the cytotoxic phenotype appeared equally sensitive to in vivo whole body irradiation. In parallel, the CD34+ cell subset remained basically unchanged after whole body irradiation. Finally, the CD3{minus}, 56+, 16+ natural killer cell subset was relatively radioresistant (91 and 74% of its initial value, after 2 and 4 Gy, respectively) as compared to other lymphocyte subsets. Our study provides evidence that T and B cell subsets seem to be highly radiosensitive in vivo. The CD34+ progenitor/stem cells and NK cells seem to be more radioresistant. This latter result might provide clues to the understanding of the pathophysiogeny of radiation-induced aplasia and of the engrafment/rejection process following bone marrow transplantation. 20 refs., 3 figs., 1 tab.

  9. A Comparative Study of the T Cell Stimulatory and Polarizing Capacity of Human Primary Blood Dendritic Cell Subsets

    PubMed Central

    Sittig, Simone P.; Bakdash, Ghaith; Weiden, Jorieke; Sköld, Annette E.; Tel, Jurjen; Figdor, Carl G.; de Vries, I. Jolanda M.

    2016-01-01

    Dendritic cells (DCs) are central players of immune responses; they become activated upon infection or inflammation and migrate to lymph nodes, where they can initiate an antigen-specific immune response by activating naive T cells. Two major types of naturally occurring DCs circulate in peripheral blood, namely, myeloid and plasmacytoid DCs (pDCs). Myeloid DCs (mDCs) can be subdivided based on the expression of either CD1c or CD141. These human DC subsets differ in surface marker expression, Toll-like receptor (TLR) repertoire, and transcriptional profile, suggesting functional differences between them. Here, we directly compared the capacity of human blood mDCs and pDCs to activate and polarize CD4+ T cells. CD141+ mDCs show an overall more mature phenotype over CD1c+ mDC and pDCs; they produce less IL-10 and more IL-12 than CD1c+ mDCs. Despite these differences, all subsets can induce the production of IFN-γ in naive CD4+ T cells. CD1c+ and CD141+ mDCs especially induce a strong T helper 1 profile. Importantly, naive CD4+ T cells are not polarized towards regulatory T cells by any subset. These findings further establish all three human blood DCs—despite their differences—as promising candidates for immunostimulatory effectors in cancer immunotherapy. PMID:27057096

  10. CXCR6 marks a novel subset of T-bet(lo)Eomes(hi) natural killer cells residing in human liver.

    PubMed

    Stegmann, Kerstin A; Robertson, Francis; Hansi, Navjyot; Gill, Upkar; Pallant, Celeste; Christophides, Theodoros; Pallett, Laura J; Peppa, Dimitra; Dunn, Claire; Fusai, Giuseppe; Male, Victoria; Davidson, Brian R; Kennedy, Patrick; Maini, Mala K

    2016-01-01

    Natural killer cells (NK) are highly enriched in the human liver, where they can regulate immunity and immunopathology. We probed them for a liver-resident subset, distinct from conventional bone-marrow-derived NK. CXCR6+ NK were strikingly enriched in healthy and diseased liver compared to blood (p < 0.0001). Human hepatic CXCR6+ NK had an immature phenotype (predominantly CD56(bright)CD16-CD57-), and expressed the tissue-residency marker CD69. CXCR6+ NK produced fewer cytotoxic mediators and pro-inflammatory cytokines than the non-liver-specific CXCR6- fraction. Instead CXCR6+ NK could upregulate TRAIL, a key death ligand in hepatitis pathogenesis. CXCR6 demarcated liver NK into two transcriptionally distinct populations: T-bet(hi)Eomes(lo)(CXCR6-) and T-bet(lo)Eomes(hi)(CXCR6+); the latter was virtually absent in the periphery. The small circulating CXCR6+ subset was predominantly T-bet(hi)Eomes(lo), suggesting its lineage was closer to CXCR6- peripheral than CXCR6+ liver NK. These data reveal a large subset of human liver-resident T-bet(lo)Eomes(hi) NK, distinguished by their surface expression of CXCR6, adapted for hepatic tolerance and inducible anti-viral immunity. PMID:27210614

  11. CXCR6 marks a novel subset of T-betloEomeshi natural killer cells residing in human liver

    PubMed Central

    Stegmann, Kerstin A.; Robertson, Francis; Hansi, Navjyot; Gill, Upkar; Pallant, Celeste; Christophides, Theodoros; Pallett, Laura J.; Peppa, Dimitra; Dunn, Claire; Fusai, Giuseppe; Male, Victoria; Davidson, Brian R.; Kennedy, Patrick; Maini, Mala K.

    2016-01-01

    Natural killer cells (NK) are highly enriched in the human liver, where they can regulate immunity and immunopathology. We probed them for a liver-resident subset, distinct from conventional bone-marrow-derived NK. CXCR6+ NK were strikingly enriched in healthy and diseased liver compared to blood (p < 0.0001). Human hepatic CXCR6+ NK had an immature phenotype (predominantly CD56brightCD16−CD57−), and expressed the tissue-residency marker CD69. CXCR6+ NK produced fewer cytotoxic mediators and pro-inflammatory cytokines than the non-liver-specific CXCR6− fraction. Instead CXCR6+ NK could upregulate TRAIL, a key death ligand in hepatitis pathogenesis. CXCR6 demarcated liver NK into two transcriptionally distinct populations: T-bethiEomeslo(CXCR6−) and T-betloEomeshi(CXCR6+); the latter was virtually absent in the periphery. The small circulating CXCR6+ subset was predominantly T-bethiEomeslo, suggesting its lineage was closer to CXCR6− peripheral than CXCR6+ liver NK. These data reveal a large subset of human liver-resident T-betloEomeshi NK, distinguished by their surface expression of CXCR6, adapted for hepatic tolerance and inducible anti-viral immunity. PMID:27210614

  12. Effects of doxycycline on haematology, blood chemistry and peripheral blood lymphocyte subsets of healthy dogs and dogs naturally infected with Ehrlichia canis.

    PubMed

    Villaescusa, A; García-Sancho, M; Rodríguez-Franco, F; Tesouro, M Á; Sainz, Á

    2015-06-01

    Canine monocytic ehrlichiosis (CME), caused by Ehrlichia canis, is a vector-borne disease with a worldwide distribution. It has been proposed that the pathogenesis, clinical severity and outcome of disease caused by Ehrlichia spp. can be attributed to the immune response rather than to any direct rickettsial effect. Moreover, doxycycline, the antimicrobial of choice for the treatment of CME, has immunomodulatory and anti-inflammatory properties associated with blood leukocyte proliferation function, cytokine synthesis, and matrix metalloproteinase activity. In order to assess the potential effects of doxycycline, dependent and independent of its antimicrobial activity, the present study compared changes in haematology, blood chemistry and circulating lymphocyte subpopulations in 12 healthy dogs and 20 dogs with CME after doxycycline therapy. Some changes were recorded only in the CME affected dogs, probably due to the antimicrobial effect of doxycycline. However, increases in mean corpuscular haemoglobin, mean corpuscular haemoglobin concentration, platelet count and α2-globulins, and decreased plasma creatinine were observed in both healthy and CME affected dogs. The absolute count of B lymphocytes (CD21(+)) increased initially, but then decreased until the end of the study period in both groups. A potential effect of doxycycline unrelated to its antimicrobial activity against E. canis is suggested, taking into account the results observed both in healthy dogs and in dogs with CME. PMID:25957920

  13. LincRNA landscape in human lymphocytes highlights regulation of T cell differentiation by linc-MAF-4

    PubMed Central

    Curti, Serena; Gruarin, Paola; Provasi, Elena; Sugliano, Elisa; Marconi, Maurizio; De Francesco, Raffaele; Geginat, Jens; Bodega, Beatrice; Abrignani, Sergio; Pagani, Massimiliano

    2014-01-01

    Long non-coding-RNAs are emerging as important regulators of cellular functions but little is known on their role in human immune system. Here we investigated long intergenic non-coding-RNAs (lincRNAs) in thirteen T and B lymphocyte subsets by RNA-seq analysis and de novo transcriptome reconstruction. Over five hundred new lincRNAs were identified and lincRNAs signatures were described. Expression of linc-MAF-4, a chromatin-associated TH1-specific lincRNA, was inversely correlated with MAF, a TH2-associated transcription factor. Linc-MAF-4 down-regulation skewed T cell differentiation toward TH2. We identified a long-distance interaction between linc-MAF-4 and MAF genomic regions, where linc-MAF-4 associates with LSD1 and EZH2, suggesting linc-MAF-4 regulated MAF transcription by recruitment of chromatin modifiers. Our results demonstrate a key role of lincRNAs in T lymphocyte differentiation. PMID:25621826

  14. Immunomodulation by neutrophil myeloperoxidase and hydrogen peroxide: differential susceptibility of human lymphocyte functions.

    PubMed

    el-Hag, A; Lipsky, P E; Bennett, M; Clark, R A

    1986-05-01

    The coexistence of activated polymorphonuclear leukocytes and lymphocytes in tumor masses and inflammatory tissues suggests the possibility of interaction between secreted neutrophil products and nearby lymphocytes. To test this hypothesis, we examined the effects of neutrophil myeloperoxidase and H2O2 on lymphocytes. Human peripheral blood mononuclear leukocytes were exposed to myeloperoxidase, an H2O2-generating system (glucose + glucose oxidase), and a halide, and were then tested for functional activities. Natural killer activity against K562 cells, lymphocyte proliferation in response to mitogens, and generation of immunoglobulin-secreting cells were all susceptible to oxidative injury by myeloperoxidase and H2O2. The degree as well as the mechanism of suppression was dependent on the glucose oxidase concentration (i.e., the rate of H2O2 delivery). At low H2O2 flux, myeloperoxidase was essential for induction of lymphocyte suppression; as the rate of H2O2 generation increased, suppression became myeloperoxidase-independent and was mediated by H2O2 alone. Various lymphocyte functions were differentially susceptible to oxidative injury by myeloperoxidase and H2O2. The proliferative response to poke-weed mitogen was the least sensitive, whereas antibody formation was the most sensitive. Proliferative responses to concanavalin A and phytohemagglutinin as well as natural killer activity displayed intermediate degrees of susceptibility. In all assays, lymphocyte viability was greater than 90%. Removal of monocytes from mononuclear leukocytes by adherence to glass increased susceptibility of lymphocytes to oxidative injury. Monocytes in proportions within the range present in peripheral blood mononuclear leukocytes protected lymphocyte functions against oxidative injury by myeloperoxidase and H2O2. This study demonstrates a differential susceptibility of various immune functions to oxidative injury by the neutrophil products myeloperoxidase and H2O2, and shows, in

  15. Specific high-affinity binding sites for a synthetic gliadin heptapeptide of human peripheral blood lymphocytes

    SciTech Connect

    Payan, D.G.; Horvath, K.; Graf, L.

    1987-03-23

    The synthetic peptide containing residues 43-49 of ..cap alpha..-gliadin, the major protein component of gluten, has previously been shown to inhibit the production of lymphokine activities by mononuclear leukocytes. The authors demonstrate using radiolabeled ..cap alpha..-gliadin(43-49) that human peripheral blood lymphocytes express approximately 20,000-25,000 surface receptors for this peptide, with a dissociation constant (K/sub D/) of 20 nM. In addition, binding is inhibited by naloxone and an enkephalin analog, thus confirming the functional correlate which demonstrates inhibition by these agents of ..cap alpha..-gliadin(43-49) functional effects. Furthermore, B-lymphocytes bind specifically a greater amount of (/sup 125/I)..cap alpha..-gliadin(43-49) than T-lymphocytes. The lymphocyte ..cap alpha..-gliadin(43-49) receptor may play an important role in mediating the immunological response to ..cap alpha..-gliadin. 16 references, 4 figures.

  16. T Lymphocyte Subsets and Cytokines in Rats Transplanted with Adipose-Derived Mesenchymal Stem Cells and Acellular Nerve for Repairing the Nerve Defects

    PubMed Central

    Chen, Ou; Chu, Ting-gang; Ding, Jian; Yu, Qing

    2015-01-01

    Objective The aim of this study was to explore the immunity in rats transplanted with adipose-derived mesenchymal stem cells (ADSCs) and acellular nerve (ACN) for repairing sciatic nerve defects. Methods ADSCs were isolated from the adipose tissues of Wistar rats. Sprague-Dawley rats were used to establish a sciatic nerve defect model and then divided into four groups, according to the following methods : Group A, allogenic nerve graft; Group B, allograft with ACN; Group C, allograft ADSCs+ACN, and Group D, nerve autograft. Results At the day before transplantation and 3, 7, 14, and 28 days after transplantation, orbital venous blood of the Sprague-Dawley rats in each group was collected to detect the proportion of CD3+, CD4+, and CD8+ subsets using flow cytometry and to determine the serum concentration of interleukin-2 (IL-2), tumor necrosis factor-α (TNF-α) and interferon-γ (IFN-γ) using enzyme-linked immunosorbent assay (ELISA). At each postoperative time point, the proportion of CD3+, CD4+, and CD8+ subsets and the serum concentration of IL-2, TNF-α, and IFN-γ in group C were all near to those in group B and group D, in which no statistically significant difference was observed. As compared with group A, the proportion of CD3+, CD4+, and CD8+ subsets and the serum concentration of IL-2, TNF-α, and IFN-γ were significantly reduced in group C (p<0.05). Conclusion The artificial nerve established with ADSCs and ACN has no obvious allograft rejection for repairing rat nerve defects. PMID:26361524

  17. COMPARATIVE GENOTOXIC RESPONSES TO ARSENITE IN GUINEA PIG, MOUSE, RAT AND HUMAN LYMPHOCYTES

    EPA Science Inventory

    Comparative genotoxic responses to arsenite in guinea pig, mouse, rat and human
    lymphocytes.

    Inorganic arsenic is a known human carcinogen causing skin, lung, and bladder cancer following chronic exposures. Yet, long-term laboratory animal carcinogenicity studies have ...

  18. Cytostatic and genotoxic effect of temephos in human lymphocytes and HepG2 cells.

    PubMed

    Benitez-Trinidad, A B; Herrera-Moreno, J F; Vázquez-Estrada, G; Verdín-Betancourt, F A; Sordo, M; Ostrosky-Wegman, P; Bernal-Hernández, Y Y; Medina-Díaz, I M; Barrón-Vivanco, B S; Robledo-Marenco, M L; Salazar, A M; Rojas-García, A E

    2015-06-01

    Temephos is an organophosphorus pesticide that is used in control campaigns against Aedes aegypti mosquitoes, which transmit dengue. In spite of the widespread use of temephos, few studies have examined its genotoxic potential. The aim of this study was to evaluate the cytotoxic, cytostatic and genotoxic effects of temephos in human lymphocytes and hepatoma cells (HepG2). The cytotoxicity was evaluated with simultaneous staining (FDA/EtBr). The cytostatic and genotoxic effects were evaluated using comet assays and the micronucleus technique. We found that temephos was not cytotoxic in either lymphocytes or HepG2 cells. Regarding the cytostatic effect in human lymphocytes, temephos (10 μM) caused a significant decrease in the percentage of binucleated cells and in the nuclear division index as well as an increase in the apoptotic cell frequency, which was not the case for HepG2 cells. The comet assay showed that temephos increased the DNA damage levels in human lymphocytes, but it did not increase the MN frequency. In contrast, in HepG2 cells, temephos increased the tail length, tail moment and MN frequency in HepG2 cells compared to control cells. In conclusion, temephos causes stable DNA damage in HepG2 cells but not in human lymphocytes. These findings suggest the importance of temephos biotransformation in its genotoxic effect. PMID:25746384

  19. Arctigenin from Arctium lappa inhibits interleukin-2 and interferon gene expression in primary human T lymphocytes

    PubMed Central

    2011-01-01

    Background Arctium lappa (Niubang), a Chinese herbal medicine, is used to treat tissue inflammation. This study investigates the effects of arctigenin (AC), isolated from A. lappa, on anti-CD3/CD28 Ab-stimulated cell proliferation and cytokine gene expression in primary human T lymphocytes. Methods Cell proliferation was determined with enzyme immunoassays and the tritiated thymidine uptake method. Cytokine production and gene expression were analyzed with reverse transcription-polymerase chain reaction. Results AC inhibited primary human T lymphocytes proliferation activated by anti-CD3/CD28 Ab. Cell viability test indicated that the inhibitory effects of AC on primary human T lymphocyte proliferation were not due to direct cytotoxicity. AC suppressed interleukin-2 (IL-2) and interferon-γ (IFN-γ) production in a concentration-dependent manner. Furthermore, AC decreased the IL-2 and IFN-γ gene expression in primary human T lymphocytes induced by anti-CD3/CD28 Ab. Reporter gene analyses revealed that AC decreased NF-AT-mediated reporter gene expression. Conclusion AC inhibited T lymphocyte proliferation and decreased the gene expression of IL-2, IFN-γ and NF-AT. PMID:21435270

  20. Ontogeny of Innate T Lymphocytes – Some Innate Lymphocytes are More Innate than Others

    PubMed Central

    Vermijlen, David; Prinz, Immo

    2014-01-01

    Innate lymphocytes have recently received a lot of attention. However, there are different ideas about the definition of what is “innate” in lymphocytes. Lymphocytes without V(D)J-rearranged antigen receptors are now termed innate lymphoid cells (ILCs) and include cells formerly known as natural killer (NK) cells. Also, lymphocytes that are innate should be able to recognize microbial or stress-induced patterns and react rapidly without prior sensitization, as opposed to adaptive immune responses. Formally, genuine innate lymphocytes would be present before or at birth. Here, we review the ontogeny of human and mouse innate T lymphocyte populations. We focus on γδ T cells, which are prototype lymphocytes that often use their V(D)J rearrangement machinery to generate genetically encoded predetermined recombinations of antigen receptors. We make parallels between the development of γδ T cells with that of innate αβ T cells [invariant (i)NKT and mucosa-associated invariant T cells] and compare this with the ontogeny of innate B cells and ILCs (including NK cells). We conclude that some subsets are more innate than others, i.e., innate lymphocytes that are made primarily early in utero during gestation while others are made after birth. In practice, a ranking of innateness by ontogeny has implications for the reconstitution of innate lymphocyte subsets after hematopoietic stem cell transplantation. PMID:25346734

  1. Assessment of in vitro genotoxic and cytotoxic effects of flurbiprofen on human cultured lymphocytes.

    PubMed

    Timocin, Taygun; Ila, Hasan Basri; Dordu, Tuba; Husunet, Mehmet Tahir; Tazehkand, Mostafa Norizadeh; Valipour, Ebrahim; Topaktas, Mehmet

    2016-07-01

    Flurbiprofen is non-steroidal anti-inflammatory drug which is commonly used for its analgesic, antipyretic, and anti-inflammatory effects. The purpose of the study was to explore the genotoxic and cytotoxic effects of flurbiprofen in human cultured lymphocytes by sister chromatid exchange, chromosome aberration, and cytokinesis-blocked micronucleus tests. 10, 20, 30, and 40 μg/mL concentrations of flurbiprofen (solvent is DMSO) were used to treatment of human cultured lymphocytes at two different treatment periods (24 and 48 h). Flurbiprofen had no significant genotoxic effect in any of these tests. But exposing to flurbiprofen for 24 and 48 h led to significant decrease on proliferation index, mitotic index, and nuclear division index (NDI). Also, all decreases were concentration-dependent (except NDI at 24 h treatment period). Consequently, the findings of this research showed that flurbiprofen had cytotoxic effects in human blood lymphocytes. PMID:26738809

  2. Protection of human myeloid dendritic cell subsets against influenza A virus infection is differentially regulated upon TLR stimulation.

    PubMed

    Baharom, Faezzah; Thomas, Saskia; Bieder, Andrea; Hellmér, Maria; Volz, Julia; Sandgren, Kerrie J; McInerney, Gerald M; Karlsson Hedestam, Gunilla B; Mellman, Ira; Smed-Sörensen, Anna

    2015-05-01

    The proinflammatory microenvironment in the respiratory airway induces maturation of both resident and infiltrating dendritic cells (DCs) upon influenza A virus (IAV) infection. This results in upregulation of antiviral pathways as well as modulation of endocytic processes, which affect the susceptibility of DCs to IAV infection. Therefore, it is highly relevant to understand how IAV interacts with and infects mature DCs. To investigate how different subsets of human myeloid DCs (MDCs) involved in tissue inflammation are affected by inflammatory stimulation during IAV infection, we stimulated primary blood MDCs and inflammatory monocyte-derived DCs (MDDCs) with TLR ligands, resulting in maturation. Interestingly, MDDCs but not MDCs were protected against IAV infection after LPS (TLR4) stimulation. In contrast, stimulation with TLR7/8 ligand protected MDCs but not MDDCs from IAV infection. The reduced susceptibility to IAV infection correlated with induction of type I IFNs. We found that differential expression of TLR4, TRIF, and MyD88 in the two MDC subsets regulated the ability of the cells to enter an antiviral state upon maturation. This difference was functionally confirmed using small interfering RNA and inhibitors. Our data show that different human MDC subsets may play distinct roles during IAV infection, as their capacity to induce type I IFNs is dependent on TLR-specific maturation, resulting in differential susceptibility to IAV infection. PMID:25801434

  3. Identifying developmental toxicity pathways for a subset of ToxCast chemicals using human embryonic stem cells and metabolomics.

    PubMed

    Kleinstreuer, N C; Smith, A M; West, P R; Conard, K R; Fontaine, B R; Weir-Hauptman, A M; Palmer, J A; Knudsen, T B; Dix, D J; Donley, E L R; Cezar, G G

    2011-11-15

    Metabolomics analysis was performed on the supernatant of human embryonic stem (hES) cell cultures exposed to a blinded subset of 11 chemicals selected from the chemical library of EPA's ToxCast™ chemical screening and prioritization research project. Metabolites from hES cultures were evaluated for known and novel signatures that may be indicative of developmental toxicity. Significant fold changes in endogenous metabolites were detected for 83 putatively annotated mass features in response to the subset of ToxCast chemicals. The annotations were mapped to specific human metabolic pathways. This revealed strong effects on pathways for nicotinate and nicotinamide metabolism, pantothenate and CoA biosynthesis, glutathione metabolism, and arginine and proline metabolism pathways. Predictivity for adverse outcomes in mammalian prenatal developmental toxicity studies used ToxRefDB and other sources of information, including Stemina Biomarker Discovery's predictive DevTox® model trained on 23 pharmaceutical agents of known developmental toxicity and differing potency. The model initially predicted developmental toxicity from the blinded ToxCast compounds in concordance with animal data with 73% accuracy. Retraining the model with data from the unblinded test compounds at one concentration level increased the predictive accuracy for the remaining concentrations to 83%. These preliminary results on a 11-chemical subset of the ToxCast chemical library indicate that metabolomics analysis of the hES secretome provides information valuable for predictive modeling and mechanistic understanding of mammalian developmental toxicity. PMID:21925528

  4. S15176 and S16950 interaction with Cyclosporin A antiproliferative effect on cultured human lymphocytes.

    PubMed

    Albengres, E; Le Louët, H; d'Athis, P; Tillement, J P

    2001-02-01

    S15176 and S16950 are trimetazidine derivatives that antagonize more strongly than the parent drug mitochondrial toxicity, which leads to cellular hypoxia and nephrotoxicity in kidneys experimentally exposed to cyclosporin A. We have investigated whether every derivative might interact or not with the inhibitory effect of Cyclosporin A on the proliferation of cultured human lymphocytes. S15176 significantly increased the antilymphoproliferative effect of Cyclosporin A, whereas S15176 by itself neither displayed any antilymphoproliferative effect, nor did it induce any apoptotic process in cultured human lymphocytes. The effect of S16950 was not significant. PMID:11468012

  5. Radioprotective effect of chicory seeds against genotoxicity induced by ionizing radiation in human normal lymphocytes.

    PubMed

    Hosseinimehr, S J; Ghaffari-Rad, V; Rostamnezhad, M; Ghasemi, A; Allahverdi Pourfallah, T; Shahani, S

    2015-01-01

    The search for less-toxic radioprotective agents has led to a growing trend towards natural products. Protective effect of the methanolic extract of chicory seeds (MCS) was investigated against genotoxicity induced by ionizing radiation in human lymphocytes. Human peripheral blood samples were collected and incubated with MCS at different concentrations (10, 50, 100, and 200 μg/mL) for two hours. The whole blood samples were exposed in vitro to X-ray at dose 2.5 Gy. Then, the lymphocytes were cultured with mitogenic stimulation to determine the micronucleus in cytokinesis blocked binucleated cell. The methanolic extract at all doses significantly reduced the frequency of micronuclei in binucleated lymphocytes, as compared with similarly irradiated lymphocytes without any extract treatment. The maximum protection was observed at 200 μg/mL of MCS, it completely protected genotoxicity induced by ionizing radiation in human lymphocytes. The extract exhibited a concentration-dependent radical scavenging activity on 1,1-diphenyl-2-picryl hydrazyl free radicals. HPLC analysis of MCS showed this extract is containing chlorogenic acid as a phenolic compound. These data suggest that the radioprotective effect of methanolic extract of chicory seeds can be attributed to the presence of phenolic compounds such as chlorogenic acid which act as antioxidant agents. PMID:26278267

  6. Radioprotective Effect of Achillea millefolium L Against Genotoxicity Induced by Ionizing Radiation in Human Normal Lymphocytes

    PubMed Central

    Shahani, Somayeh; Rostamnezhad, Mostafa; Ghaffari-rad, Vahid; Ghasemi, Arash; Allahverdi Pourfallah, Tayyeb

    2015-01-01

    The radioprotective effect of Achillea millefolium L (ACM) extract was investigated against genotoxicity induced by ionizing radiation (IR) in human lymphocytes. Peripheral blood samples were collected from human volunteers and incubated with the methanolic extract of ACM at different concentrations (10, 50, 100, and 200 μg/mL) for 2 hours. At each dose point, the whole blood was exposed in vitro to 2.5 Gy of X-ray and then the lymphocytes were cultured with mitogenic stimulation to determine the micronuclei in cytokinesis-blocked binucleated cell. Antioxidant capacity of the extract was determined using free radical-scavenging method. The treatment of lymphocytes with the extract showed a significant decrease in the incidence of micronuclei binucleated cells, as compared with similarly irradiated lymphocytes without any extract treatment. The maximum protection and decrease in frequency of micronuclei were observed at 200 μg/mL of ACM extract which completely protected genotoxicity induced by IR in human lymphocytes. Achillea millefolium extract exhibited concentration-dependent radical-scavenging activity on 1,1-diphenyl-2-picryl hydrazyl free radicals. These data suggest that the methanolic extract of ACM may play an important role in the protection of normal tissues against genetic damage induced by IR. PMID:26675116

  7. Genetic modification of human T lymphocytes for the treatment of hematologic malignancies

    PubMed Central

    Hoyos, Valentina; Savoldo, Barbara; Dotti, Gianpietro

    2012-01-01

    Modern chemotherapy regimens and supportive care have produced remarkable improvements in the overall survival of patients with hematologic malignancies. However, the development of targeted small molecules, monoclonal antibodies, and biological therapies that demonstrate greater efficacy and lower toxicity remains highly desirable in hematology, and oncology in general. In the context of biological therapies, T-lymphocyte based treatments have enormous potential. Donor lymphocyte infusion in patients relapsed after allogeneic hematopoietic stem cell transplant pioneered the concept that T lymphocytes can effectively control tumor growth, and this was then followed by the development of cell culture strategies to generate T lymphocytes with selective activity against tumor cells. Over the past decade, it has become clear that the adoptive transfer of ex vivo expanded antigen-specific cytotoxic T lymphocytes promotes sustained antitumor effects in patients with virus-associated lymphomas, such as Epstein-Barr virus related post-transplant lymphomas and Hodgkin's lymphomas. Because of this compelling clinical evidence and the concomitant development of methodologies for robust gene transfer to human T lymphocytes, the field has rapidly evolved, offering new opportunities to extend T-cell based therapies. This review summarizes the most recent biological and clinical developments using genetically manipulated T cells for the treatment of hematologic malignancies. PMID:22929977

  8. Lymphocyte-conditioned medium protects human monocyte-macrophages from cholesteryl ester accumulation.

    PubMed Central

    Fogelman, A M; Seager, J; Haberland, M E; Hokom, M; Tanaka, R; Edwards, P A

    1982-01-01

    Exposure of human monocyte-macrophages to as little as 50 microliters of cultured medium from lymphocytes stimulated by concanavalin A (Con A) resulted in a dramatic decrease in the activities of the low density lipoprotein (LDL) receptor pathway, the LDL-dextran sulfate pathway, and the scavenger receptor pathway. This effect was not seen when the monocyte-macrophages were exposed to culture medium from lymphocytes cultured without Con A or with Con A together with alpha-methyl mannoside or control medium without lymphocytes. The activity of 3-hydroxy-3-methyglutaryl-coenzyme A reductase also decreased in monocyte-macrophages exposed to culture medium from stimulated lymphocytes. Acyl-CoA:cholesterol O-acyltransferase activity, protein synthesis, protein content, phagocytosis of heat-killed yeast, and non-receptor-mediated endocytosis were not inhibited. Monocyte-macrophages exposed to malondialdehyde altered-LDL in the presence of stimulated lymphocyte culture medium accumulated substantially less cholesteryl esters than did cells in control medium. We propose that substances produced by stimulated lymphocytes may be useful in protecting macrophages from cholesteryl ester accumulation. Images PMID:6278500

  9. Choline deficiency increases lymphocyte apoptosis and DNA damage in humans2,3

    PubMed Central

    da Costa, Kerry-Ann; Niculescu, Mihai D; Craciunescu, Corneliu N; Fischer, Leslie M; Zeisel, Steven H

    2008-01-01

    Background: Whereas deficiency of the essential nutrient choline is associated with DNA damage and apoptosis in cell and rodent models, it has not been shown in humans. Objective: The objective was to ascertain whether lymphocytes from choline-deficient humans had greater DNA damage and apoptosis than did those from choline-sufficient humans. Design: Fifty-one men and women aged 18–70 y were fed a diet containing the recommended adequate intake of choline (control) for 10 d. They then were fed a choline-deficient diet for up to 42 d before repletion with 138–550 mg choline/d. Blood was collected at the end of each phase, and peripheral lymphocytes were isolated. DNA damage and apoptosis were then assessed by activation of caspase-3, terminal deoxynucleotide transferase–mediated dUTP nick end-labeling, and single-cell gel electrophoresis (COMET) assays. Results: All subjects fed the choline-deficient diet had lymphocyte DNA damage, as assessed by COMET assay, twice that found when they were fed the control diet. The subjects who developed organ dysfunction (liver or muscle) when fed the choline-deficient diet had significantly more apoptotic lymphocytes, as assessed by the activated caspase-3 assay, than when fed the control diet. Conclusions: A choline-deficient diet increased DNA damage in humans. Subjects in whom these diets induced liver or muscle dys-function also had higher rates of apoptosis in their peripheral lymphocytes than did subjects who did not develop organ dysfunction. Assessment of DNA damage and apoptosis in lymphocytes appears to be a clinically useful measure in humans (such as those receiving parenteral nutrition) in whom choline deficiency is suspected. PMID:16825685

  10. Dietary nucleotides increase the proportion of a TCR gammadelta+ subset of intraepithelial lymphocytes (IEL) and IL-7 production by intestinal epithelial cells (IEC); implications for modification of cellular and molecular cross-talk between IEL and IEC by dietary nucleotides.

    PubMed

    Nagafuchi, S; Totsuka, M; Hachimura, S; Goto, M; Takahashi, T; Yajima, T; Kuwata, T; Kaminogawa, S

    2000-07-01

    We have investigated the effects of dietary nucleotides on intraepithelial lymphocytes (IEL) and intestinal epithelial cells (IEC) in weanling mice. The proportion of T-cell receptor (TCR) gammadelta+ IEL in BALB/c mice fed a diet supplemented with nucleotides (NT(+) diet) was significantly higher than that in mice fed the nucleotide-free diet, while the proportion of TCR alphabeta+ IEL in NT(+) diet-fed mice was significantly decreased. The change of the TCR alphabeta+/TCR gammadelta+ ratio was mainly observed in a CD8 alphaalpha+ subset of IEL. IEC from NT(+) diet-fed mice produced a higher level of IL-7, which is important in the development of TCR gammadelta+ IEL, than those from control diet-fed mice. The expression levels of IL-7 and IL-2 receptors on IEL were not different between the two dietary groups. Our findings suggest that the increased population of a TCR gammadelta+ IEL subset by feeding nucleotides may be caused by the increased production of IL-7 by IEC. PMID:10945264

  11. Specific subsets of immune cells in human decidua differ between normal pregnancy and preeclampsia - a prospective observational study

    PubMed Central

    2009-01-01

    Background Changes in the balance of decidual leucocyte populations may lead to an unfavourable uterine microenvironment which may be associated with the development of preeclampsia (PE). In this study, we therefore investigated the leucocyte subpopulations in decidual tissues of 33 women with preeclampsia and 66 control patients. Methods Decidua was either obtained via curettage during cesarean section or dissected from the surface of the basal plate of the placenta after spontaneous delivery. We used FACS analysis to quantify decidual leukocytes (CD45), NK cells (CD56+/CD16+ and CD56++/CD16-), antigen presenting cells (HLA-DR, DC-Sign, CD14) and T/B cells (CD8, CD4, alpha-beta-T-cell receptor, gamma-delta-T-cell receptor, CD25, CD19). Results The number of decidual cytotoxic CD8+T-lymphocytes (P < 0.02), alpha-beta -T-cell receptor positive T cells (P < 0.03) and of CD56+/CD16+ NK cells (P < 0.03) was lower in decidua from women with PE than in decidua from control patients. Conclusion The observed reduction of specific leucocyte subsets could create a microenvironment which is unfavourable for an appropriate placentation and could thereby be involved in the development of preeclamptic symptoms. PMID:19930648

  12. Single strand dna breaks in human lymphocytes exposed to para-phenylenediamine and its derivatives.

    PubMed

    Chye, Soi Mei; Hseu, You Cheng; Liang, Shih-Hsiung; Chen, Chin-Hui; Chen, Ssu Ching

    2008-01-01

    Para-Phenylenediamine (PPD), the main aromatic amines used in the hair dye formation, and its four derivatives (2-chloro-p-phenylenediamine, 4-chloro-o-phenylenediamine, 2-nitro-p-phenylenediamine, and 4-nitro-o-phenylenediamine) were examined for their potential to produce single strand DNA breaks in human lymphocytes using the alkaline comet assay. Results revealed that all the tested chemicals within the range of doses from 100 microM to 500 microM showed the genotoxicity in a dose-dependent manner after the incubation of lymphocytes with these chemicals for 2 h. In this study, we first reported that PPD and its four derivatives can elicit the type of single strand breaks in human lymphocytes. PMID:18058049

  13. Methionine synthetase activity of human lymphocytes both replete in and depleted of vitamin B12.

    PubMed

    Hall, C A; Begley, J A; Chu, R C

    1986-10-01

    The activity of the enzyme methionine synthetase (MS) (methyltetrahydrofolate:homocysteine methyltransferase) (EC 2.1.1.13) was measured in human lymphocytes of various types and cobalamin (vitamin B12) status. Total and holo MS activity was low in unstimulated peripheral blood lymphocytes from persons with tissue deficiency of cobalamin, but not in cells from those with low serum cobalamin levels for other reasons. The MS activity of the lymphocyte was increased by treatment of the patients with vitamin B12. The number of lymphocytes was often low or low normal in the circulation of those deficient in cobalamin. Holo MS activity was low in an established line of human B cells, RPMI 6410 cells, depleted of cobalamin. The total and holo MS activity of both RPMI 6410 cells, replete or depleted, and lymphocytes stimulated in culture was increased by cobalamin in vitro; 222 nmol/L free cobalamin was roughly the equivalent of 0.22 nmol/L cobalamin bound to transcobalamin II. Both lymphocytes and RPMI 6410 cells required folate for growth and could meet these needs via methylfolate, homocysteine, and the cobalamin-dependent MS reaction. Depleted RPMI 6410 cells, however, used cobalamin in some way in addition to the provision of available folate from methylfolate. The consequences of the reduced MS activity in deficient cells could include a reduction in available folate with diminished capacity for clonal expansion of lymphocytes in reaction to infection and impairment of essential methylations including those of protein synthesis. The prompt induction of MS activity by cobalamin, especially in the in vitro model, suggests an effect of therapeutic vitamin B12 well in advance of the numerical increase in cells of the blood. PMID:3760673

  14. Expression and regulation of normal and polymorphic epithelial sodium channel by human lymphocytes.

    PubMed

    Bubien, J K; Watson, B; Khan, M A; Langloh, A L; Fuller, C M; Berdiev, B; Tousson, A; Benos, D J

    2001-03-16

    Gene expression, protein expression, and function of amiloride-sensitive sodium channels were examined in human lymphocytes from normal individuals and individuals with Liddle's disease. Using reverse transcriptase polymerase chain reactions, expression of all three cloned epithelial sodium channel (ENaC) subunits was detected in lymphocytes. Polyclonal antibodies to bovine alpha-ENaC bound to the plasma membrane of normal and Liddle's lymphocytes. A quantitative analysis of fluorescence-tagged ENaC antibodies indicated a 2.5-fold greater surface binding of the antibodies to Liddle's lymphocytes compared with normal lymphocytes. The relative binding intensity increased significantly (25%; p < 0.001) for both normal and Liddle's cells after treatment with 40 microM 8-CPT-cAMP. Amiloride-sensitive whole cell currents were recorded under basal and cAMP-treated conditions for both cell types. Liddle's cells had a 4.5-fold larger inward sodium conductance compared with normal cells. A specific 25% increase in the inward sodium current was observed in normal cells in response to cAMP treatment. Outside-out patches from both cell types under both treatment conditions revealed no obvious differences in the single channel conductance. The P(open) was 4.2 +/- 3.9% for patches from non-Liddle's cells, and 27.7 +/- 5.4% in patches from Liddle's lymphocytes. Biochemical purification of a protein complex, using the same antibodies used for the immunohistochemistry, yielded a functional sodium channel complex that was inhibited by amiloride when reconstituted into lipid vesicles and incorporated into planar lipid bilayers. These four independent methodologies yielded findings consistent with the hypotheses that human lymphocytes express functional, regulatable ENaC and that the mutation responsible for Liddle's disease induces excessive channel expression. PMID:11113130

  15. Antibacterial activity of neem nanoemulsion and its toxicity assessment on human lymphocytes in vitro.

    PubMed

    Jerobin, Jayakumar; Makwana, Pooja; Suresh Kumar, R S; Sundaramoorthy, Rajiv; Mukherjee, Amitava; Chandrasekaran, Natarajan

    2015-01-01

    Neem (Azadirachta indica) is recognized as a medicinal plant well known for its antibacterial, antimalarial, antiviral, and antifungal properties. Neem nanoemulsion (NE) (O/W) is formulated using neem oil, Tween 20, and water by high-energy ultrasonication. The formulated neem NE showed antibacterial activity against the bacterial pathogen Vibrio vulnificus by disrupting the integrity of the bacterial cell membrane. Despite the use of neem NE in various biomedical applications, the toxicity studies on human cells are still lacking. The neem NE showed a decrease in cellular viability in human lymphocytes after 24 hours of exposure. The neem NE at lower concentration (0.7-1 mg/mL) is found to be nontoxic while it is toxic at higher concentrations (1.2-2 mg/mL). The oxidative stress induced by the neem NE is evidenced by the depletion of catalase, SOD, and GSH levels in human lymphocytes. Neem NE showed a significant increase in DNA damage when compared to control in human lymphocytes (P<0.05). The NE is an effective antibacterial agent against the bacterial pathogen V. vulnificus, and it was found to be nontoxic at lower concentrations to human lymphocytes. PMID:26491309

  16. Antibacterial activity of neem nanoemulsion and its toxicity assessment on human lymphocytes in vitro

    PubMed Central

    Jerobin, Jayakumar; Makwana, Pooja; Suresh Kumar, RS; Sundaramoorthy, Rajiv; Mukherjee, Amitava; Chandrasekaran, Natarajan

    2015-01-01

    Neem (Azadirachta indica) is recognized as a medicinal plant well known for its antibacterial, antimalarial, antiviral, and antifungal properties. Neem nanoemulsion (NE) (O/W) is formulated using neem oil, Tween 20, and water by high-energy ultrasonication. The formulated neem NE showed antibacterial activity against the bacterial pathogen Vibrio vulnificus by disrupting the integrity of the bacterial cell membrane. Despite the use of neem NE in various biomedical applications, the toxicity studies on human cells are still lacking. The neem NE showed a decrease in cellular viability in human lymphocytes after 24 hours of exposure. The neem NE at lower concentration (0.7–1 mg/mL) is found to be nontoxic while it is toxic at higher concentrations (1.2–2 mg/mL). The oxidative stress induced by the neem NE is evidenced by the depletion of catalase, SOD, and GSH levels in human lymphocytes. Neem NE showed a significant increase in DNA damage when compared to control in human lymphocytes (P<0.05). The NE is an effective antibacterial agent against the bacterial pathogen V. vulnificus, and it was found to be nontoxic at lower concentrations to human lymphocytes. PMID:26491309

  17. Improved four-color flow cytometry method using fluo-3 and triple immunofluorescence for analysis of intracellular calcium ion ([Ca2+]i) fluxes among mouse lymph node B- and T-lymphocyte subsets.

    PubMed

    Greimers, R; Trebak, M; Moutschen, M; Jacobs, N; Boniver, J

    1996-03-01

    A visible-light, dual-laser, flow cytometric method was developed for the simultaneous analysis of intracellular ionized calcium concentration ([Ca2+]i) and three cell-surface markers (CD4, CD8, and Thy-1.2 antigens) by using the calcium probe fluo-3 and using R-phycoerythrin (PE), peridinin chlorophyll-alpha protein (PerCP), and allophycocyanin (APC) conjugated monoclonal antibodies (MoAbs). This improved method was used in the analysis of [Ca2+]i mobilization upon in vitro stimulation with mitogenic lectins [phytohaemagglutinin (PHA) or concanavalin A (ConA)], anti-CD3 MoAbs, or A23187 calcium ionophore in the heterogeneous lymph node cell populations from healthy C57BL/Ka mice. The present results show that the calcium responses were heterogeneous and dependent on the cellular immunophenotype, not only on lectins or anti-CD3 MoAbs stimulation, but also on the receptor-independent A23187 ionophore stimulation. An in situ fluo-3 calibration method (using A23187 and metabolic poisons in Ca2+ /EGTA buffers with known free calcium concentrations) indicated a resting [Ca2+]i in lymphocytes of 103 +/- 23 nM (mean +/- S.D.) but with significant differences between the [Ca2+]i in B cells and in all of the T-cell subsets (CD4+Thy-1+, CD4+Thy-1-, and CD8+T cells). Both the B cells and the T-cell subsets showed an increase of fluo-3 fluorescence upon in vitro stimulation with ConA or PHA, but the calcium mobilization following lectin stimulation was time delayed in all T-cell subsets. Only the T cells, including the CD4+Thy-1- subset, responded to anti-CD3 MoAbs. The percentage of responding cells upon stimulation with ConA was higher in T cells than in B cells. By contrast, PHA gave a higher response in B cells. After stimulation with different mitogens, [Ca2+]i increased in both CD4+ and CD8+ T-cell subsets. However, the percentage of responding cells was far higher in the CD4+Thy-1+ subset than in the CD4+Thy-1- or the CD8+T-cell subsets. The stimulation with A23187

  18. Diphtheria toxin resistance in human lymphocytes and lymphoblasts in the in vivo somatic cell mutation test

    SciTech Connect

    Tomkins, D.J.; Wei, L.; Laurie, K.E.

    1985-01-01

    It has been shown that circulating peripheral blood lymphocytes can be used for the enumeration of 6-thioguanine-resistant cells that presumably arise by mutation in vivo. This somatic cell mutation test has been studied in lymphocytes from human populations exposed to known mutagens and/or carcinogens. The sensitivity of the test could be further enhanced by including other gene markers, since there is evidence for locus-specific differences in response to mutagens. Resistance to diphtheria toxin (Dip/sup r/) seemed like a potential marker to incorporate into the test because the mutation acts codominantly, can readily be selected in human diploid fibroblasts and Chinese hamster cells with no evidence for cell density or cross-feeding effects, and can be assayed for in nondividing cells by measuring protein synthesis inhibition. Blood samples were collected from seven individuals, and fresh, cryopreserved, or Epstein-Barr virus (EBV)-transformed lymphocytes were tested for continued DNA synthesis (TH-thymidine, autoradiography) or protein synthesis (TVS-methionine, scintillation counting). Both fresh and cryopreserved lymphocytes, stimulated to divide with phytohemagglutinin (PHA), continued to synthesize DNA in the presence of high doses of diphtheria toxin (DT). Similarly, both dividing (PHA-stimulated) and nondividing fresh lymphocytes carried on significant levels of protein synthesis even 68 hr after exposure to 100 flocculating units (LF)/ml DT. The results suggest that human T and B lymphocytes may not be as sensitive to DT protein synthesis inhibition as human fibroblast and Chinese hamster cells. For this reason, Dip/sup r/ may not be a suitable marker for the somatic cell mutation test.

  19. Influence of macrophages on HSV-1 induced IL 2 production by human lymphocytes

    SciTech Connect

    Clouse, K.A.; Orosz, C.G.; Sheridan, J.F.

    1986-03-05

    Previous work has demonstrated that human peripheral blood mononuclear cells (PBMC) from HSV-1 seropositive individuals produce interleukin 2 (IL 2) following stimulation in vitro with uv-inactivated herpes simplex virus type 1 (HSV antigen). This study investigated the accessory macrophage (MO) and monokine requirements for IL 2 production by enriched T lymphocytes from HSV-1 seropositive individuals. Following removal of accessory MO populations, enriched T lymphocytes did not secrete IL 2 in response to HSV antigen. However, IL 2 production was restored by the addition of autologous, ..gamma..-irradiated (5000R) MO. HSV antigen-pulsed MO also induced IL 2 production by enriched T lymphocytes. Furthermore, when HSV-pulsed macrophages were treated with paraformaldehyde they no longer caused T lymphocytes to produce IL 2 unless exogenous monokines were provided. Neither exogenous monokines nor purified human IL 1 could support HSV antigen induced IL 2 production in the absence of MO. These studies demonstrated that MO are required for HSV-induced IL 2 production by T lymphocytes from HSV-1 seropositive individuals. Furthermore, these MO appear to provide two functions required for IL 2 production: viral antigen display and monokine production.

  20. Chemopreventive effect and lack of genotoxicity and mutagenicity of the exopolysaccharide botryosphaeran on human lymphocytes.

    PubMed

    Malini, M; Camargo, M S; Hernandes, L C; Vargas-Rechia, C G; Varanda, E A; Barbosa, A M; Dekker, R F H; Matsumoto, S T; Antunes, L M G; Cólus, I M S

    2016-10-01

    Carbohydrate biopolymers of fungal-origin are an important natural resource in the search for new bioagents with therapeutic and nutraceutical potential. In this study the mutagenic, genotoxic, antigenotoxic and antioxidant properties of the fungal exopolysaccharide botryosphaeran, a (1→3)(1→6)-β-D-glucan, from Botryosphaeria rhodina MAMB-05, was evaluated. The mutagenicity was assessed at five concentrations in Salmonella typhimurium by the Ames test. Normal and tumor (Jurkat cells) human T lymphocyte cultures were used to evaluate the genotoxicity and antigenotoxicity (Comet assay) of botryosphaeran alone and in combination with the mutagen methyl methanesulfonate (MMS). The ability of botryosphaeran to reduce the production of reactive oxygen and nitrogen species (RONS) generated by hydrogen peroxide was assessed using the CM-H2DCFDA probe in lymphocyte cultures under different treatment times. None of the evaluated botryosphaeran concentrations were mutagenic in bacteria, nor induced genotoxicity in normal and tumor lymphocytes. Botryosphaeran protected lymphocyte DNA against damage caused by MMS under simultaneous treatment and post-treatment conditions. However, botryosphaeran was not able to reduce the RONS generated by H2O2. Besides the absence of genotoxicity, botryosphaeran exerted a protective effect on human lymphocytes against genotoxic damage caused by MMS. These results are important in the validation of botryosphaeran as a therapeutic agent targeting health promotion. PMID:27387458

  1. Tissue-resident and memory properties of human T-cell and NK-cell subsets.

    PubMed

    Lugli, Enrico; Hudspeth, Kelly; Roberto, Alessandra; Mavilio, Domenico

    2016-08-01

    Efficient immune responses to invading pathogens are the result of the complex but coordinated synergy between a variety of cell types from both the innate and adaptive arms of the immune system. While adaptive and innate immune responses are highly complementary, some cells types within these two systems perform similar functions, underscoring the need for redundancy and increased flexibility. In this review, we will discuss the striking shared features of immunological memory and tissue residency recently discovered between T cells, a component of the adaptive immune system, and natural killer (NK) cells, members generally assigned to the innate compartment. Specifically, we will focus on the T-cell and NK-cell diversity at the single-cell level, on the discrete function of specific subsets, and on their anatomical location. Finally, we will discuss the implication of such diversity in the generation of long-term memory. PMID:27431095

  2. Long-term study of the impact of methotrexate on serum cytokines and lymphocyte subsets in patients with active rheumatoid arthritis: correlation with pharmacokinetic measures

    PubMed Central

    Kremer, Joel M; Lawrence, David A; Hamilton, Robert; McInnes, Iain B

    2016-01-01

    Objective To describe changes in immune parameters observed during long-term methotrexate (MTX) therapy in patients with active rheumatoid arthritis (RA) and explore correlations with simultaneously measured MTX pharmacokinetic (PKC) parameters. Design Prospective, open-label, long-term mechanism of action study. Setting University clinic. Methods MTX was initiated at a single weekly oral dose of 7.5 mg and dose adjusted for efficacy and toxicity for the duration of the study. Standard measures of disease activity were performed at baseline and every 6–36 months. Serum cytokine measurements in blood together with lymphocyte surface immunophenotypes and stimulated peripheral blood mononuclear cell (PBMC) cytokine production were assessed at each clinical evaluation. Results Cytokine concentrations exhibited multiple significant correlations with disease activity measures over time. The strongest correlations observed were for interleukin (IL)-6 (r=0.45, p<0.0001 for swollen joints and r=0.32, p=0.002 for tender joints) and IL-8 (r=0.25, p=0.01 for swollen joints). Significant decreases from baseline were observed in serum IL-1B, IL-6 and IL-8 concentrations. The most significant changes were observed for IL-6 (p<0.001). Significant increases from baseline were observed in IL-2 release from PBMCs ex vivo (p<0.01). In parallel, multiple statistically significant correlations were observed between MTX PKC measures and immune parameters. The change in swollen joint count correlated inversely with the change in area under the curve (AUC) for MTX (r=−0.63, p=0.007). Conclusions MTX therapy of patients with RA is accompanied by a variety of changes in serum cytokine expression, which in turn correlate strongly with clinical disease activity and MTX pharmacokinetics (PKCs). These data strongly support the notion that MTX mediates profound and functionally relevant effects on the immunological hierarchy in the RA lesion. PMID:27335660

  3. CD28 costimulation and CD28 expression in T lymphocyte subsets in HIV-1 infection with and without progression to AIDS

    PubMed Central

    Choremi-Papadopoulou, H; Panagiotou, N; Samouilidou, E; Kontopidou, F; Viglis, V; Antoniadou, A; Kosmidis, J; Kordossis, T

    2000-01-01

    In a prospective study of 152 HIV-1 patients (with and without progression to AIDS) we examined CD28 MoAb costimulation and CD3 MoAb response using whole blood culture at baseline and up to either the time of AIDS diagnosis or the end of the observation period. CD28 antigen expression on both CD4+ and CD8+ T lymphocytes was also studied in both groups of patients. In patients who progressed to AIDS, CD28 MoAb costimulation was found to be decreased. Univariate time-dependent analysis showed that decreases in (i) absolute numbers of either CD4+, CD4+CD28+, CD8+CD28+ T cells, (ii) CD28 MoAb costimulation, and (iii) CD3 MoAb response, and an increase in CD8+CD28− %, are significant predictors for progression to AIDS. In addition, multivariate time-dependent analysis demonstrated that a decrease in CD28 MoAb costimulation (but not a decrease in CD3 MoAb response) was predictive for progression to AIDS, as were decreases in the percentage of CD4+ T cells and the absolute number of CD4+CD28+ T cells. Thus, CD28 MoAb costimulation can be considered a useful assay for monitoring HIV-1 infection. Furthermore, apart from the early increase in the percentage of CD8+CD28− T cells and an increase in the percentage of CD28− on CD8+ T cells in both groups of patients at baseline compared with normal controls, a negative correlation was found to exist between the percentages of CD4+ or CD4+CD28+ T cells and the percentage of CD8+CD28− T cells; this suggests that these cells are probably mutually regulated. PMID:10691923

  4. Inorganic arsenic represses interleukin-17A expression in human activated Th17 lymphocytes

    SciTech Connect

    Morzadec, Claudie; Macoch, Mélinda; Robineau, Marc; Sparfel, Lydie; Fardel, Olivier; Vernhet, Laurent

    2012-08-01

    Trivalent inorganic arsenic [As(III)] is an efficient anticancer agent used to treat patients suffering from acute promyelocytic leukemia. Recently, experimental studies have clearly demonstrated that this metalloid can also cure lymphoproliferative and/or pro-inflammatory syndromes in different murine models of chronic immune-mediated diseases. T helper (Th) 1 and Th17 lymphocytes play a central role in development of these diseases, in mice and humans, especially by secreting the potent pro-inflammatory cytokine interferon-γ and IL-17A, respectively. As(III) impairs basic functions of human T cells but its ability to modulate secretion of pro-inflammatory cytokines by differentiated Th lymphocytes is unknown. In the present study, we demonstrate that As(III), used at concentrations clinically achievable in plasma of patients, has no effect on the secretion of interferon-γ from Th1 cells but almost totally blocks the expression and the release of IL-17A from human Th17 lymphocytes co-stimulated for five days with anti-CD3 and anti-CD28 antibodies, in the presence of differentiating cytokines. In addition, As(III) specifically reduces mRNA levels of the retinoic-related orphan receptor (ROR)C gene which encodes RORγt, a key transcription factor controlling optimal IL-17 expression in fully differentiated Th17 cells. The metalloid also blocks initial expression of IL-17 gene induced by the co-stimulation, probably in part by impairing activation of the JNK/c-Jun pathway. In conclusion, our results demonstrate that As(III) represses expression of the major pro-inflammatory cytokine IL-17A produced by human Th17 lymphocytes, thus strengthening the idea that As(III) may be useful to treat inflammatory immune-mediated diseases in humans. -- Highlights: ► Arsenic inhibits secretion of IL-17A from human naïve and memory Th17 lymphocytes. ► Arsenic represses early expression of IL-17A gene in human activated T lymphocytes. ► Arsenic interferes with activation of

  5. Human tumor-derived exosomes selectively impair lymphocyte responses to interleukin-2.

    PubMed

    Clayton, Aled; Mitchell, J Paul; Court, Jacquelyn; Mason, Malcolm D; Tabi, Zsuzsanna

    2007-08-01

    Exosomes are nanometer-sized vesicles, secreted by normal and neoplastic cells. The outcome following interaction between the cellular immune system and cancer-derived exosomes is not well understood. Interleukin-2 (IL-2) is a key factor supporting expansion and differentiation of CTL and natural killer (NK) cells but can also support regulatory T cells and their suppressive functions. Our study examined whether tumor-derived exosomes could modify lymphocyte IL-2 responses. Proliferation of healthy donor peripheral blood lymphocytes in response to IL-2 was inhibited by tumor exosomes. In unfractionated lymphocytes, this effect was seen in all cell subsets. Separating CD4(+) T cells, CD8(+) T cells, and NK cells revealed that CD8(+) T-cell proliferation was not inhibited in the absence of CD4(+) T cells and that NK cell proliferation was only slightly impaired. Other exosome effects included selective impairment of IL-2-mediated CD25 up-regulation, affecting all but the CD3(+)CD8(-) T-cell subset. IL-2-induced Foxp3 expression by CD4(+)CD25(+) cells was not inhibited by tumor exosomes, and the suppressive function of CD4(+)CD25(+) T cells was enhanced by exosomes. In contrast, exosomes directly inhibited NK cell killing function in a T-cell-independent manner. Analysis of tumor exosomes revealed membrane-associated transforming growth factor beta(1) (TGFbeta(1)), which contributed to the antiproliferative effects, shown by using neutralizing TGFbeta(1)-specific antibody. The data show an exosome-mediated mechanism of skewing IL-2 responsiveness in favor of regulatory T cells and away from cytotoxic cells. This coordinated "double hit" to cellular immunity strongly implicates the role of exosomes in tumor immune evasion. PMID:17671216

  6. Prolactin-immunoglobulin G complexes from human serum act as costimulatory ligands causing proliferation of malignant B lymphocytes.

    PubMed Central

    Walker, A M; Montgomery, D W; Saraiya, S; Ho, T W; Garewal, H S; Wilson, J; Lorand, L

    1995-01-01

    Several lines of evidence indicate that immunoglobulin-bound prolactin found in human serum is not a conventional complex between an anti-prolactin antibody and prolactin but a different type of association of prolactin with the Fab portion of IgG heavy chains. The complex of prolactin with IgG was purified from serum by anti-human prolactin affinity chromatography and was shown to contain close to 1 mole of N epsilon-(gamma-glutamyl)lysine crosslinks per mole of complex, a characteristic feature in structures crosslinked by transglutaminase. Interestingly, the complex caused a proliferation of cells from a subset of patients with chronic lymphocytic leukemia, while it was inactive in a cell proliferation prolactin bioassay. By contrast, human prolactin stimulated the proliferation of cells in the bioassay but had no effect on the complex-responsive cells from the patients. Competition studies with prolactin and free Fc fragment of IgG demonstrated a necessity for engaging both the prolactin and the immunoglobulin receptors for proliferation. More importantly, competition for the growth response by free prolactin and IgG suggests both possible reasons for the slow growth of this neoplasm as well as avenues for control of the disease. Images Fig. 1 Fig. 2 Fig. 4 PMID:7724552

  7. INDUCTION OF MICRONUCLEI BY X-RADIATION IN HUMAN, MOUSE, AND RAT PERIPHERAL BLOOD LYMPHOCYTES

    EPA Science Inventory

    We compared the radiosensitivity of human, rat, and mouse peripheral blood lymphocytes (PBLs) by analyzing micronuclei (MN) in cytochalasin B-induced binucleated (BN) cells. or each species and dose, 4 ml aliquots of whole blood were X-irradiated to obtain doses of 38, 75, 150, o...

  8. Transfection of an immunoglobulin kappa gene into mature human B lymphocytes

    SciTech Connect

    Bich-Thuy, L.T.; Queen, C.

    1988-01-01

    The authors show in this report that the transcription induced by interleukin-2 or pokeweed mitogens of the kappa MOPC 41 immunoglobulin light-chain gene transfected into primary human or murine B lymphocytes initiates from a previously unobserved start site about 26 base pairs upstream of the start site used in myeloma cell lines.

  9. The Effect of a Grape Seed Extract on Radiation-Induced DNA Damage in Human Lymphocytes

    NASA Astrophysics Data System (ADS)

    Dicu, Tiberius; Postescu, Ion D.; Foriş, Vasile; Brie, Ioana; Fischer-Fodor, Eva; Cernea, Valentin; Moldovan, Mircea; Cosma, Constantin

    2009-05-01

    Plant-derived antioxidants due to their phenolic compounds content are reported as potential candidates for reducing the levels of oxidative stress in living organisms. Grape seed extracts are very potent antioxidants and exhibit numerous interesting pharmacologic activities. Hydroethanolic (50/50, v/v) standardized extract was obtained from red grape seed (Vitis vinifera, variety Burgund Mare—BM). The total polyphenols content was evaluated by Folin-Ciocalteu procedure and expressed as μEq Gallic Acid/ml. The aim of this study was to evaluate the potential antioxidant effects of different concentrations of BM extract against 60Co γ-rays induced DNA damage in human lymphocytes. Samples of human lymphocytes were incubated with BM extract (12.5, 25.0 and 37.5 μEq GA/ml, respectively) administered at 30 minutes before in vitro irradiation with γ-rays (2 Gy). The DNA damage and repair in lymphocytes were evaluated using alkaline comet assay. Using the lesion score, the radiation-induced DNA damage was found to be significantly different (p<0.05) from control, both in the absence and presence of BM extract (except the lymphocytes treated with 37.5 μEq GA/ml BM extract). DNA repair analyzed by incubating the irradiated cells at 37° C and 5% CO2 atmosphere for 2 h, indicated a significant difference (p<0.05) in the lymphocytes group treated with 25.0 μEq GA/ml BM extract, immediately and two hours after irradiation. These results suggest radioprotective effects after treatment with BM extract in human lymphocytes.

  10. Inhibition of the lymphocyte metabolic switch by the oxidative burst of human neutrophils.

    PubMed

    Kramer, Philip A; Prichard, Lynn; Chacko, Balu; Ravi, Saranya; Overton, E Turner; Heath, Sonya L; Darley-Usmar, Victor

    2015-09-01

    Activation of the phagocytic NADPH oxidase-2 (NOX-2) in neutrophils is a critical process in the innate immune system and is associated with elevated local concentrations of superoxide, hydrogen peroxide (H2O2) and hypochlorous acid. Under pathological conditions, NOX-2 activity has been implicated in the development of autoimmunity, indicating a role in modulating lymphocyte effector function. Notably, T-cell clonal expansion and subsequent cytokine production requires a metabolic switch from mitochondrial respiration to aerobic glycolysis. Previous studies demonstrate that H2O2 generated from activated neutrophils suppresses lymphocyte activation but the mechanism is unknown. We hypothesized that activated neutrophils would prevent the metabolic switch and suppress the effector functions of T-cells through a H2O2-dependent mechanism. To test this, we developed a model co-culture system using freshly isolated neutrophils and lymphocytes from healthy human donors. Extracellular flux analysis was used to assess mitochondrial and glycolytic activity and FACS analysis to assess immune function. The neutrophil oxidative burst significantly inhibited the induction of lymphocyte aerobic glycolysis, caused inhibition of oxidative phosphorylation and suppressed lymphocyte activation through a H2O2-dependent mechanism. Hydrogen peroxide and a redox cycling agent, DMNQ, were used to confirm the impact of H2O2 on lymphocyte bioenergetics. In summary, we have shown that the lymphocyte metabolic switch from mitochondrial respiration to glycolysis is prevented by the oxidative burst of neutrophils. This direct inhibition of the metabolic switch is then a likely mechanism underlying the neutrophil-dependent suppression of T-cell effector function. PMID:25951298

  11. Loss of telomeric DNA during aging of normal and trisomy 21 human lymphocytes

    SciTech Connect

    Vaziri, H.; Uchida, I.; Lan Wei; Harley, C.B. ); Schaechter, F.; Cohen, D. ); Xiaoming Zhu; Effros, R. )

    1993-04-01

    The telomere hypothesis of cellular aging proposes that loss of telomeric DNA (TTAGGG) from human chromosomes may ultimately cause cell-cycle exit during replicative senescence. Since lymphocytes have a limited replicative capacity and since blood cells were previously shown to lose telomeric DNA during aging in vivo, the authors wished to determine (a) whether accelerated telomere loss is associated with the premature immunosenescence of lymphocytes in individuals with Down syndrome (DS) and (b) whether telomeric DNA is also lost during aging of lymphocytes in vitro. To investigate the effects of aging and trisomy 21 on telomere loss in vivo, genomic DNA was isolated from peripheral blood lymphocytes of 140 individuals (age 0--107 years), including 21 DS patients (age 0--45 years). Digestion with restriction enzymes HinfI and RsaI generated terminal restriction fragments (TRFs), which were detected by Southern analysis using a telomere-specific probe ([sup 32]P-(C[sub 3]TA[sub 2])[sub 3]). The rate of telomere loss was calculated from the decrease in mean TRF length, as a function of donor age. DS patients showed a significantly higher rate of telomere loss with donor age (133 [+-] 15 bp/year) compared with age-matched controls (41 [+-] 7.7 bp/year) (P < .0005), suggesting that accelerated telomere loss is a biomarker of premature immunosenescence of DS patients and that it may play a role in this process. Telomere loss during aging in vitro was calculated for lymphocytes from four normal individuals, grown in culture for 10--30 population doublings. The rate of telomere loss was [approximately]120 bp/cell doubling, comparable to that seen in other somatic cells. Moreover, telomere lengths of lymphocytes from centenarians and from older DS patients were similar to those of senescent lymphocytes in culture, which suggests that replicative senescence could partially account for aging of the immune system in DS patients and in elderly individuals. 31 refs., 3 figs.

  12. Identifying developmental toxicity pathways for a subset of ToxCast chemicals using human embryonic stem cells and metabolomics

    SciTech Connect

    Kleinstreuer, N.C.; Smith, A.M.; West, P.R.; Conard, K.R.; Fontaine, B.R.; Weir-Hauptman, A.M.; Palmer, J.A.; Knudsen, T.B.; Dix, D.J.; Donley, E.L.R.; Cezar, G.G.

    2011-11-15

    Metabolomics analysis was performed on the supernatant of human embryonic stem (hES) cell cultures exposed to a blinded subset of 11 chemicals selected from the chemical library of EPA's ToxCast Trade-Mark-Sign chemical screening and prioritization research project. Metabolites from hES cultures were evaluated for known and novel signatures that may be indicative of developmental toxicity. Significant fold changes in endogenous metabolites were detected for 83 putatively annotated mass features in response to the subset of ToxCast chemicals. The annotations were mapped to specific human metabolic pathways. This revealed strong effects on pathways for nicotinate and nicotinamide metabolism, pantothenate and CoA biosynthesis, glutathione metabolism, and arginine and proline metabolism pathways. Predictivity for adverse outcomes in mammalian prenatal developmental toxicity studies used ToxRefDB and other sources of information, including Stemina Biomarker Discovery's predictive DevTox Registered-Sign model trained on 23 pharmaceutical agents of known developmental toxicity and differing potency. The model initially predicted developmental toxicity from the blinded ToxCast compounds in concordance with animal data with 73% accuracy. Retraining the model with data from the unblinded test compounds at one concentration level increased the predictive accuracy for the remaining concentrations to 83%. These preliminary results on a 11-chemical subset of the ToxCast chemical library indicate that metabolomics analysis of the hES secretome provides information valuable for predictive modeling and mechanistic understanding of mammalian developmental toxicity. -- Highlights: Black-Right-Pointing-Pointer We tested 11 environmental compounds in a hESC metabolomics platform. Black-Right-Pointing-Pointer Significant changes in secreted small molecule metabolites were observed. Black-Right-Pointing-Pointer Perturbed mass features map to pathways critical for normal development and

  13. In vitro modeling of the interaction between human epithelial cells and lymphocytes upon influenza infection.

    PubMed

    Ilyushina, Natalia A; Wright, Peter F

    2016-09-01

    Influenza viruses are a continuous threat to humans because of their ability to cross species barriers and adapt to new hosts. Data from murine studies, along with limited human data, suggest that CD8(+) cytotoxic T lymphocytes (CTL) that recognize conserved epitopes of structural influenza proteins are the main mediators of influenza virus clearance. Additionally, the fact that many CTLs recognize epitopes shared between different influenza strains offers the potential for broad cross-strain immunity. However, the mechanisms of cellular immunity against influenza viruses are poorly defined in humans, where the CTL response has been hard to measure and interpret. We developed a novel CTL assay that utilizes fully differentiated nasal human epithelial cells taken from volunteers as permissive targets for autologous peripheral blood-derived influenza virus-specific cytotoxic T lymphocytes. This in vitro system of human lymphocyte-epithelial cell co-cultures can be considered as the closest approximation to events in vivo and can be employed for studying the interactions between the pathogen and human host. Modeling of the natural interaction process between the primary cell type that supports the productive replication of influenza and immune cells may allow us to put in perspective CTLs as a correlate of immunity to influenza in humans. PMID:27102577

  14. On-line studies of activation events in primary human T lymphocytes.

    PubMed

    Bental, M; Deutsch, C

    1994-04-01

    In this paper, we review our NMR studies of human peripheral blood T lymphocytes. These studies focus on the physiological and biochemical alterations accompanying cell cycle progression. In particular, we have characterized phosphorus metabolism, glucose utilization and lactate production, and pH regulation using 31P, 13C, and 19F NMR, respectively. These studies required developing new methods for monitoring on-line stimulation of quiescent T cells under sterile, physiological conditions (i.e., CO2/HCO3- buffer, 37 degrees C) for prolonged periods of time. A perfusion system optimized for T lymphocytes inside agarose beads is described. In addition, custom-designed 19F NMR pH indicators were synthesized, characterized, and used to determine intracellular pH in quiescent lymphocytes, stimulated lymphocytes, and lymphocytes undergoing the G0-G1 transition. These unique molecular probes are described in detail. Finally, the physiological relevance of our findings regarding carbon metabolism and pH regulation is considered in the context of mitogenesis. PMID:8069534

  15. Toxicological Implications and Inflammatory Response in Human Lymphocytes Challenged with Oxytetracycline.

    PubMed

    Di Cerbo, A; Palatucci, A T; Rubino, V; Centenaro, S; Giovazzino, A; Fraccaroli, E; Cortese, L; Ruggiero, G; Guidetti, G; Canello, S; Terrazzano, G

    2016-04-01

    Antibiotics are widely used in zoo technical and veterinary practices as feed supplementation to ensure wellness of farmed animals and livestock. Several evidences have been suggesting both the toxic role for tetracyclines, particularly for oxytetracycline (OTC). This potential toxicity appears of great relevance for human nutrition and for domestic animals. This study aimed to extend the evaluation of such toxicity. The biologic impact of the drug was assessed by evaluating the proinflammatory effect of OTC and their bone residues on cytokine secretion by in vitro human peripheral blood lymphocytes. Our results showed that both OTC and OTC-bone residues significantly induced the T lymphocyte and non-T cell secretion of interferon (IFN)-γ, as cytokine involved in inflammatory responses in humans as well as in animals. These results may suggest a possible implication for new potential human and animal health risks depending on the entry of tetracyclines in the food-processing chain. PMID:26537863

  16. Early effects of low dose 12C6+ ion or X-ray irradiation on human peripheral blood lymphocytes

    NASA Astrophysics Data System (ADS)

    Chen, Yingtai; Li, Yumin; Zhang, Hong; Xie, Yi; Chen, Xuezhong; Ren, Jinyu; Zhang, Xiaowei; Zhu, Zijiang; Liu, Hongliang; Zhang, Yawei

    2010-04-01

    The aim of this study was to estimate the acute effects of low dose 12C6+ ions or X-ray radiation on human immune function. The human peripheral blood lymphocytes (HPBL) of seven healthy donors were exposed to 0.05 Gy 12C6+ ions or X-ray radiation and cell responses were measured at 24 h after exposure. The cytotoxic activities of HPBL were determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT); the percentages of T and NK cells subsets were detected by flow cytometry; mRNA expression of interleukin (IL)-2, tumor necrosis factor (TNF)-α and interferon (IFN)-γ were examined by real time quantitative RT-PCR (qRT-PCR); and these cytokines protein levels in supernatant of cultured cells were assayed by enzyme-linked immunosorbent assays (ELISA). The results showed that the cytotoxic activity of HPBL, mRNA expression of IL-2, IFN-γ and TNF-α in HPBL and their protein levels in supernatant were significantly increased at 24 h after exposure to 0.05 Gy 12C6+ ions radiation and the effects were stronger than observed for X-ray exposure. However, there was no significant change in the percentage of T and NK cells subsets of HPBL. These results suggested that 0.05 Gy high linear energy transfer (LET) 12C6+ radiation was a more effective approach to host immune enhancement than that of low LET X-ray. We conclude that cytokines production might be used as sensitive indicators of acute response to LDI.

  17. Differentiation of human monocytes and derived subsets of macrophages and dendritic cells by the HLDA10 monoclonal antibody panel

    PubMed Central

    Ohradanova-Repic, Anna; Machacek, Christian; Fischer, Michael B; Stockinger, Hannes

    2016-01-01

    The mononuclear phagocyte system, consisting of monocytes, macrophages and dendritic cells (DCs), has an important role in tissue homeostasis as well as in eliciting immune responses against invading pathogens. Blood monocytes have been viewed for decades as precursors of tissue macrophages. Although the newest data show that in the steady state resident macrophages of many organs are monocyte independent, blood monocytes critically contribute to tissue macrophage and DC pools upon inflammation. To better understand the relationship between these populations and their phenotype, we isolated and differentiated human blood CD14+ monocytes in vitro into immature and mature monocyte-derived dendritic cells (MoDCs) as well as into seven different monocyte-derived macrophage subsets. We used the panel of 70 monoclonal antibodies (mAbs) submitted to the 10th Human Leukocyte Differentiation Antigen Workshop to determine the expression profiles of these 10 populations by flow cytometry. We now can compile subpanels of mAbs to differentiate the 10 monocyte/macrophage/MoDC subsets, providing the basis for novel diagnostic and therapeutic tools. PMID:26900469

  18. Subsets of Human Dendritic Cell Precursors Express Different Toll-like Receptors and Respond to Different Microbial Antigens

    PubMed Central

    Kadowaki, Norimitsu; Ho, Stephen; Antonenko, Svetlana; de Waal Malefyt, Rene; Kastelein, Robert A.; Bazan, Fernando; Liu, Yong-Jun

    2001-01-01

    Toll-like receptors (TLRs) are ancient microbial pattern recognition receptors highly conserved from Drosophila to humans. To investigate if subsets of human dendritic cell precursors (pre-DC), including monocytes (pre-DC1), plasmacytoid DC precursors (pre-DC2), and CD11c+ immature DCs (imDCs) are developed to recognize different microbes or microbial antigens, we studied their TLR expression and responses to microbial antigens. We demonstrate that whereas monocytes preferentially express TLR 1, 2, 4, 5, and 8, plasmacytoid pre-DC strongly express TLR 7 and 9. In accordance with these TLR expression profiles, monocytes respond to the known microbial ligands for TLR2 (peptidoglycan [PGN], lipoteichoic acid) and TLR4 (lipopolysaccharide), by producing tumor necrosis factor (TNF)-α and interleukin (IL)-6. In contrast, plasmacytoid pre-DCs only respond to the microbial TLR9-ligand, CpG-ODNs (oligodeoxynucleotides [ODNs] containing unmethylated CpG motifs), by producing IFN-α. CD11c+ imDCs preferentially express TLR 1, 2, and 3 and respond to TLR 2-ligand PGN by producing large amounts of TNF-α, and to viral double-stranded RNA-like molecule poly I:C, by producing IFN-α and IL-12. The expression of distinct sets of TLRs and the corresponding difference in reactivity to microbial molecules among subsets of pre-DCs and imDCs support the concept that they have developed through distinct evolutionary pathways to recognize different microbial antigens. PMID:11561001

  19. Subsets of human dendritic cell precursors express different toll-like receptors and respond to different microbial antigens.

    PubMed

    Kadowaki, N; Ho, S; Antonenko, S; Malefyt, R W; Kastelein, R A; Bazan, F; Liu, Y J

    2001-09-17

    Toll-like receptors (TLRs) are ancient microbial pattern recognition receptors highly conserved from Drosophila to humans. To investigate if subsets of human dendritic cell precursors (pre-DC), including monocytes (pre-DC1), plasmacytoid DC precursors (pre-DC2), and CD11c(+) immature DCs (imDCs) are developed to recognize different microbes or microbial antigens, we studied their TLR expression and responses to microbial antigens. We demonstrate that whereas monocytes preferentially express TLR 1, 2, 4, 5, and 8, plasmacytoid pre-DC strongly express TLR 7 and 9. In accordance with these TLR expression profiles, monocytes respond to the known microbial ligands for TLR2 (peptidoglycan [PGN], lipoteichoic acid) and TLR4 (lipopolysaccharide), by producing tumor necrosis factor (TNF)-alpha and interleukin (IL)-6. In contrast, plasmacytoid pre-DCs only respond to the microbial TLR9-ligand, CpG-ODNs (oligodeoxynucleotides [ODNs] containing unmethylated CpG motifs), by producing IFN-alpha. CD11c(+) imDCs preferentially express TLR 1, 2, and 3 and respond to TLR 2-ligand PGN by producing large amounts of TNF-alpha, and to viral double-stranded RNA-like molecule poly I:C, by producing IFN-alpha and IL-12. The expression of distinct sets of TLRs and the corresponding difference in reactivity to microbial molecules among subsets of pre-DCs and imDCs support the concept that they have developed through distinct evolutionary pathways to recognize different microbial antigens. PMID:11561001

  20. Potassium currents inhibition by gambierol analogs prevents human T lymphocyte activation.

    PubMed

    Rubiolo, J A; Vale, C; Martín, V; Fuwa, H; Sasaki, M; Botana, L M

    2015-07-01

    Gambierol is a marine polycyclic ether toxin, produced along with ciguatoxin congeners by the dinoflagellate Gambierdiscus toxicus. We have recently reported that two truncated skeletal analogs of gambierol comprising the EFGH- and BCDEFGH-rings of the parent compound showed similar potency to gambierol on voltage-gated potassium channels (Kv) inhibition in neurons. Gambierol and its truncated analogs share the main crucial elements for biological activity, which are the C28=C29 double bond within the H-ring and the unsaturated side chain. Since Kv channels are critical for the regulation of calcium signaling, proliferation, secretion and migration in human T lymphocytes, we evaluated the activity of both the tetracyclic and heptacyclic analogs of gambierol on potassium currents in resting T lymphocyte and their effects on interleukin-2 (IL-2) release and gene expression in activated T lymphocytes. The results presented in this work clearly demonstrate that both truncated analogs of gambierol inhibit Kv channels present in resting T lymphocytes (Kv1.3) and prevented lymphocyte activation by concanavalin A. The main effects of the heptacyclic and tetracyclic analogs of gambierol in human T cells are: (1) inhibition of potassium channels in resting and concanavalin-activated T cells in the nanomolar range, (2) inhibition of IL-2 release from concanavalin-activated T cells and (3) negatively affect the expression of genes involved in cell proliferation and immune response observed in concanavalin-activated lymphocytes. These results together with the lack of toxicity in this cellular model, indicates that both analogs of gambierol have additional potential for the development of therapeutic tools in autoimmune diseases. PMID:25155189

  1. Patient-Derived Tumor Xenografts Are Susceptible to Formation of Human Lymphocytic Tumors1

    PubMed Central

    Bondarenko, Gennadiy; Ugolkov, Andrey; Rohan, Stephen; Kulesza, Piotr; Dubrovskyi, Oleksii; Gursel, Demirkan; Mathews, Jeremy; O’Halloran, Thomas V.; Wei, Jian J.; Mazar, Andrew P.

    2015-01-01

    Patient-derived xenograft (PDX) tumor models have emerged as a new approach to evaluate the effects of cancer drugs on patients’ personalized tumor grafts enabling to select the best treatment for the cancer patient and providing a new tool for oncology drug developers. Here, we report that human tumors engrafted in immunodeficient mice are susceptible to formation of B-and T-cell PDX tumors. We xenografted human primary and metastatic tumor samples into immunodeficient mice and found that a fraction of PDX tumors generated from patients’ samples of breast, colon, pancreatic, bladder and renal cancer were histologically similar to lymphocytic neoplasms. Moreover, we found that the first passage of breast and pancreatic cancer PDX tumors after initial transplantation of the tumor pieces from the same human tumor graft could grow as a lymphocytic tumor in one mouse and as an adenocarcinoma in another mouse. Whereas subcutaneous PDX tumors resembling human adenocarcinoma histology were slow growing and non-metastatic, we found that subcutaneous PDX lymphocytic tumors were fast growing and formed large metastatic lesions in mouse lymph nodes, liver, lungs, and spleen. PDX lymphocytic tumors were comprised of B-cells which were Epstein-Barr virus positive and expressed CD45 and CD20. Because B-cells are typically present in malignant solid tumors, formation of B-cell tumor may evolve in a wide range of PDX tumor models. Although PDX tumor models show great promise in the development of personalized therapy for cancer patients, our results suggest that confidence in any given PDX tumor model requires careful screening of lymphocytic markers. PMID:26476081

  2. Epstein-Barr virus transformation induces B lymphocytes to produce human interleukin 10.

    PubMed

    Burdin, N; Péronne, C; Banchereau, J; Rousset, F

    1993-02-01

    Interleukin 10 (IL-10) is a pleiotropic factor that enhances proliferation of activated human B lymphocytes and induces them to secrete high amounts of immunoglobulins. Here we show that several human B cell lines were able to constitutively secrete human (h)IL-10. Whereas none of the pre-B nor the plasmocytic cell lines tested produced hIL-10, 25 of the 36 tested mature B cell lines (lymphoblastoid and Burkitt lymphoma cell lines) secreted hIL-10. Moreover, 24 of these 25 hIL-10-producing B cell lines contained the Epstein-Barr virus (EBV) genome, suggesting a relationship between hIL-10 production by human B cell lines and EBV expression. Accordingly, whereas polyclonal activation via triggering of surface immunoglobulins or CD40 antigen induced highly purified normal human B lymphocytes to produce only low (0.3-0.4 ng/ml) but significant amounts of hIL-10, EBV infection induced them to secrete high amounts of hIL-10 (4-9 ng/ml). Furthermore, addition of exogenous hIL-10, simultaneously to EBV infection, potentiated cell proliferation, whereas a blocking anti-IL-10 antiserum inhibited it. Thus, hIL-10 produced by infected human B lymphocytes appears to be involved in the mechanisms of EBV-induced B cell proliferation. PMID:8381152

  3. Human CTLA-4 is expressed in situ on T lymphocytes in germinal centers, in cutaneous graft-versus-host disease, and in Hodgkin's disease.

    PubMed

    Vandenborre, K; Delabie, J; Boogaerts, M A; De Vos, R; Lorré, K; De Wolf-Peeters, C; Vandenberghe, P

    1998-04-01

    Cytotoxic T-lymphocyte-associated antigen-4 (CTLA-4, CD152) is a molecule expressed on in vitro activated T cells. CTLA-4 shares important sequence homology with CD28 and binds to the same ligands, CD80 (B7-1) and CD86 (B7-2). CTLA-4 probably functions as a negative regulator of T lymphocyte activation in the mouse, although this remains to be proven for human T lymphocytes. We have developed new monoclonal antibodies against human CTLA-4 and have investigated the in situ expression of CTLA-4 in a wide variety of normal and pathological human tissues expressing CD80 and CD86. As revealed in this study, CTLA-4 is expressed on thymocytes in thymic medulla, on a subset of CD4+ T lymphocytes in germinal centers of follicular hyperplasia, on T cells, mainly CD8+, infiltrating skin affected by graft-versus-host disease, and on T cells, mainly CD4+, infiltrating Hodgkin's disease lesions. In immunoelectron microscopy, CTLA-4 was found on the plasma membrane as well as in the hyaloplasm and cytoplasmic vesicles, in agreement with its pattern of expression on in vitro activated T cells. Interestingly, no or at most scarce expression of CTLA-4 was found in granulomatous lymph nodes, T-cell-mediated inflammatory diseases, or non-Hodgkin's lymphomas, regardless of their expression of CD80 or CD86. Thus, expression of CTLA-4 appears to be induced in selective pathological conditions in vivo. The pathways leading to selective induction of CTLA-4 and its role in the pathophysiology of these conditions need to be further investigated. PMID:9546357

  4. Only a subset of C. canimorsus strains is dangerous for humans.

    PubMed

    Renzi, Francesco; Dol, Melanie; Raymackers, Alice; Manfredi, Pablo; Cornelis, Guy Richard

    2015-08-01

    Capnocytophaga canimorsus are gram-negative bacteria living as commensals in the mouth of dogs and cats. C. canimorsus cause rare but life-threatening generalized infections in humans that have been in contact with a dog or a cat. Over the last years we collected 105 C. canimorsus strains from different geographical origins and from severe human infections or healthy dogs. All these strains were analyzed by 16S rDNA sequencing and a phylogenetic tree revealed two main groups of bacteria instead of one with no relation to the geographical origin. This branching was confirmed by the whole-genome sequencing of 10 strains, supporting the evidence of a new Capnocytophaga species in dogs. Interestingly, 19 out of 19 C. canimorsus strains isolated from human infections belonged to the same species. Furthermore, most strains from this species could grow in heat-inactivated human serum (HIHS) (40/46 tested), deglycosylate IgM (48/66) and were cytochrome-oxidase positive (60/66) while most strains from the other species could not grow in HIHS (22/23 tested), could not deglycosylate IgM (33/34) and were cytochrome-oxidase negative (33/34). Here, we propose to call Capnocytophaga canis (Latin: dog) the novel, presumably less virulent dog-hosted Capnocytophaga species and to keep the name C. canimorsus for the species including human pathogens. PMID:26421271

  5. Proteome profiling of human neutrophil granule subsets, secretory vesicles, and cell membrane: correlation with transcriptome profiling of neutrophil precursors.

    PubMed

    Rørvig, Sara; Østergaard, Ole; Heegaard, Niels H H; Borregaard, Niels

    2013-10-01

    Neutrophils are indispensable in the innate immune defense against invading microorganisms. Neutrophils contain SVs and several subsets of granules that are essential for their function. Proteins present in neutrophil SVs and granules are synthesized during terminal granulopoiesis in the bone marrow. The heterogeneity of granules, as determined by marker proteins characteristic of each granule subset, is thought to result from differences in the biosynthetic windows of major classes of granule proteins, a process referred to as targeting by timing. Qualitative proteomic analysis of neutrophil granules, SVs, and plasma membrane has been performed before. Here, we performed subcellular fractionation on freshly isolated human neutrophils by nitrogen cavitation and density centrifugation on a four-layer Percoll gradient. Granule subsets were pooled and subjected to SDS-PAGE, and gel pieces were in-gel-digested with trypsin. The resulting peptides were analyzed using LTQ Orbitrap XL tandem MS. A total of 1292 unique proteins were identified and grouped, according to the neutrophil fraction, in which they displayed maximal expression. In addition to various known neutrophil proteins, several uncharacterized proteins were found, as well as proteins not described previously in neutrophils. To study the correlation between mRNA expression in neutrophil precursors and the localization of their cognate proteins, the distribution of 126 identified proteins was compared with their mRNA expression profiles. The neutrophil subcellular proteome profiles presented here may be used as a database in combination with the mRNA array database to predict and test the presence and localization of proteins in neutrophil granules and membranes. PMID:23650620

  6. CD8 sup + T lymphocytes of patients with AIDS maintain normal broad cytolytic function despite the loss of human immunodeficiency virus-specific cytotoxicity

    SciTech Connect

    Pantaleo, G.; De Maria, A.; Koenig, S.; Butini, L.; Moss, B.; Lane, H.C.; Fauci, A.S. ); Baseler, M. )

    1990-06-01

    In this study, the authors have investigated the potential mechanisms responsible for the loss of human immunodeficiency virus type 1 (HIV-1)-specific cytolytic activity in the advanced stages of HIV-1 infection. They have demonstrated that HIV-1-specific cytotoxic T lymphocytes are predominantly contained within the CD8{sup +}DR{sup +} subset. Furthermore, they have shown by a redirected killing assay that there is a dichotomy between HIV-1-specific cytolytic activity and broad cytolytic potential since the cytolytic machinery of CD8{sup +}DR{sup +} cells is still functioning even in patients with AIDS who have lost their HIV-1-specific cytolytic activity. In addition, by comparative analysis of these two types of cytolytic activity over time they have demonstrated a progressive loss of HIV-1-specific cytolytic activity in the advanced stages of the disease, whereas the cytolytic potential remained unchanged regardless of the clinical stage. On the basis of these results, they propose that the loss of HIV-1-specific cytolytic activity in HIV-1-infected individuals may result at least in part from a progressive decrease in the pool of HIV-1-specific cytotoxic T lymphocytes belonging to the CD8{sup +}DR{sup +} subset whose ability to expand has been impaired.

  7. Inhibitory effects of extracellular products from oral bacteria on human fibroblasts and stimulated lymphocytes.

    PubMed Central

    Higerd, T B; Vesole, D H; Goust, J M

    1978-01-01

    Extracellular products of 12 strains of Streptococcus mutans and 5 additional species of oral bacteria were analyzed for their ability to inhibit proliferation of fibroblastoid cells (HeLa and AV3) and blast transformation of human peripheral blood lymphocytes obtained from normal individuals. Products from S. mutans strains AHT and BHT, Streptococcus intermedius, and Actinomyces viscosus inhibited [3H]thymidine uptake by fibroblastoid cells and phytohemagglutinin-stimulated lymphocytes. Products from S. mutans E49, Streptococcus salivarius, and Actinomyces naeslundii inhibited blast transformation of human lymphocytes but did not significantly inhibit the growth of fibroblastoid cells. Preparations from S. intermedius gave the greatest inhibitory activity against both target cell types; initial characterization of this preparation suggested a single factor active in both assays, in that the heat lability and Sephadex G-200 elution profile were similar for the inhibitory activity seen with the two cell types. The molecular weight of the inhibitor, estimated by gel filtration on Sephadex G-200 and Ultragel AcA34, was approximately 160,000. The results strongly suggest that oral bacteria produce heat-labile substances that interfere with fibroblast proliferation and alter the lymphocytic immunological response. Images PMID:689736

  8. Inhibitory effects of extracellular products from oral bacteria on human fibroblasts and stimulated lymphocytes.

    PubMed

    Higerd, T B; Vesole, D H; Goust, J M

    1978-08-01

    Extracellular products of 12 strains of Streptococcus mutans and 5 additional species of oral bacteria were analyzed for their ability to inhibit proliferation of fibroblastoid cells (HeLa and AV3) and blast transformation of human peripheral blood lymphocytes obtained from normal individuals. Products from S. mutans strains AHT and BHT, Streptococcus intermedius, and Actinomyces viscosus inhibited [3H]thymidine uptake by fibroblastoid cells and phytohemagglutinin-stimulated lymphocytes. Products from S. mutans E49, Streptococcus salivarius, and Actinomyces naeslundii inhibited blast transformation of human lymphocytes but did not significantly inhibit the growth of fibroblastoid cells. Preparations from S. intermedius gave the greatest inhibitory activity against both target cell types; initial characterization of this preparation suggested a single factor active in both assays, in that the heat lability and Sephadex G-200 elution profile were similar for the inhibitory activity seen with the two cell types. The molecular weight of the inhibitor, estimated by gel filtration on Sephadex G-200 and Ultragel AcA34, was approximately 160,000. The results strongly suggest that oral bacteria produce heat-labile substances that interfere with fibroblast proliferation and alter the lymphocytic immunological response. PMID:689736

  9. Protective Effect of Onion Extract on Bleomycin-Induced Cytotoxicity and Genotoxicity in Human Lymphocytes

    PubMed Central

    Cho, Yoon Hee; Lee, Joong Won; Woo, Hae Dong; Lee, Sunyeong; Kim, Yang Jee; Lee, Younghyun; Shin, Sangah; Joung, Hyojee; Chung, Hai Won

    2016-01-01

    Following one of the world’s largest nuclear accidents, occured at Fukushima, Japan in 2011, a significant scientific effort has focused on minimizing the potential adverse health effects due to radiation exposure. The use of natural dietary antioxidants to reduce the risk of radiation-induced oxidative DNA damage is a simple strategy for minimizing radiation-related cancer rates and improving overall health. The onion is among the richest sources of dietary flavonoids and is an important food for increasing their overall intake. Therefore, we examined the effect of an onion extract on cyto- and geno-toxicity in human lymphocytes treated with bleomycin (BLM), a radiomimetic agent. In addition, we measured the frequency of micronuclei (MN) and DNA damage following treatment with BLM using a cytokinesis-blocked micronucleus assay and a single cell gel electrophoresis assay. We observed a significant increase in cell viability in lymphocytes treated with onion extract then exposed to BLM compared to cells treated with BLM alone. The frequency of BLM induced MN and DNA damage increased in a dose-dependent manner; however, when lymphocytes were pretreated with onion extract (10 and 20 μL/mL), the frequency of BLM-induced MN was decreased at all doses of BLM and DNA damage was decreased at 3 μg/mL of BLM. These results suggest that onion extract may have protective effects against BLM-induced cyto- and genotoxicity in human lymphocytes. PMID:26907305

  10. Protective Effect of Onion Extract on Bleomycin-Induced Cytotoxicity and Genotoxicity in Human Lymphocytes.

    PubMed

    Cho, Yoon Hee; Lee, Joong Won; Woo, Hae Dong; Lee, Sunyeong; Kim, Yang Jee; Lee, Younghyun; Shin, Sangah; Joung, Hyojee; Chung, Hai Won

    2016-02-01

    Following one of the world's largest nuclear accidents, occured at Fukushima, Japan in 2011, a significant scientific effort has focused on minimizing the potential adverse health effects due to radiation exposure. The use of natural dietary antioxidants to reduce the risk of radiation-induced oxidative DNA damage is a simple strategy for minimizing radiation-related cancer rates and improving overall health. The onion is among the richest sources of dietary flavonoids and is an important food for increasing their overall intake. Therefore, we examined the effect of an onion extract on cyto- and geno-toxicity in human lymphocytes treated with bleomycin (BLM), a radiomimetic agent. In addition, we measured the frequency of micronuclei (MN) and DNA damage following treatment with BLM using a cytokinesis-blocked micronucleus assay and a single cell gel electrophoresis assay. We observed a significant increase in cell viability in lymphocytes treated with onion extract then exposed to BLM compared to cells treated with BLM alone. The frequency of BLM induced MN and DNA damage increased in a dose-dependent manner; however, when lymphocytes were pretreated with onion extract (10 and 20 μL/mL), the frequency of BLM-induced MN was decreased at all doses of BLM and DNA damage was decreased at 3 μg/mL of BLM. These results suggest that onion extract may have protective effects against BLM-induced cyto- and genotoxicity in human lymphocytes. PMID:26907305

  11. The tax gene of human T-cell leukemia virus type 2 is essential for transformation of human T lymphocytes.

    PubMed Central

    Ross, T M; Pettiford, S M; Green, P L

    1996-01-01

    The mechanism of human T-cell leukemia virus (HTLV)-mediated transformation and induction of malignancy is unknown; however, several studies have implicated the viral gene product, Tax. Conclusive evidence for the role of Tax in the HTLV malignant process has been impeded by the inability to mutate tax in the context of an infectious virus and dissociate viral replication from cellular transformation. To circumvent this problem we constructed a mutant of HTLV type 2 (HTLV-2) that replicates by a Tax-independent mechanism. For these studies, the Tax response element in the viral long terminal repeat was replaced with the cytomegalovirus immediate-early promoter enhancer (C-enh). Transcription of the chimeric HTLV-2 (HTLVC-enh) was efficiently directed by this heterologous promoter. Also, the chimeric virus transformed primary human T lymphocytes with an efficiency similar to that of wild-type HTLV-2. A tax-knockout virus, termed HTLVC-enhDeltaTax, was constructed to directly assess the importance of Tax in cellular transformation. Transfection and infection studies indicated that HTLVC-enhDeltaTax was replication competent; however, HTLVC-enhDeltaTax failed to transform primary human T lymphocytes. We conclude that Tax is essential for HTLV-mediated transformation of human T lymphocytes. Furthermore, this chimeric HTLV, that replicates in the absence of Tax, should facilitate studies to determine the precise mechanism of T-lymphocyte transformation by HTLV. PMID:8764028

  12. B lymphocyte reconstitution after human bone marrow transplantation. Leu-1 antigen defines a distinct population of B lymphocytes.

    PubMed Central

    Antin, J H; Ault, K A; Rappeport, J M; Smith, B R

    1987-01-01

    Differences in the expression of Leu-1 (CD5) define two populations of recovering B cells after human marrow transplantation, Leu-1+ and Leu-1- B cells. The Leu-1+ B cells were polyclonal, of donor origin, and did not express detectable interleukin 2 receptor. Leu-1+ B cells generally appeared 2-4 wk after marrow grafting and often preceded the recovery of Leu-1- B cells. Acute and chronic graft vs. host disease (GvHD) resulted in the recovery of significantly fewer Leu-1+ B cells, whereas Leu-1- B cells were only decreased in acute GvHD. Multivariate analysis showed no significant effect of age, disease, prednisone or azathioprine, or ex vivo treatment of the marrow with anti-Leu-1 and complement on recovery of Leu-1+ and Leu-1- B cells, independent of the effects of GvHD. Leu-1+ B cells are a major lymphocyte population posttransplant. They may reflect a stage of differentiation of normal B cells or a separate B cell lineage. PMID:3112184

  13. A subset of human plasmacytoid dendritic cells expresses CD8α upon exposure to herpes simplex virus type 1

    PubMed Central

    Schuster, Philipp; Thomann, Sabrina; Werner, Maren; Vollmer, Jörg; Schmidt, Barbara

    2015-01-01

    Classical and plasmacytoid dendritic cells (DC) play important roles in the defense against murine and human infections with herpes simplex virus (HSV). So far, CD8α expression has only been reported for murine DC. CD8α+ DC have prominent cross-presenting activities, which are enhanced by murine CD8α+ PDC. The human orthologue of murine CD8α+ DC, the CD141 (BDCA3)+ DC, mainly cross-present after TLR3 ligation. We report here the serendipitous finding that a subset of human PDC upregulates CD8α upon HSV-1 stimulation, as shown by gene array and flow cytometry analyses. CD8α, not CD8ß, was expressed upon exposure. Markers of activation, migration, and costimulation were upregulated on CD8α-expressing human PDC. In these cells, increased cytokine and chemokine levels were detected that enhance development and function of T, B, and NK cells, and recruit immature DC, monocytes, and Th1 cells, respectively. Altogether, human CD8α+ PDC exhibit a highly activated phenotype and appear to recruit other immune cells to the site of inflammation. Further studies will show whether CD8α-expressing PDC contribute to antigen cross-presentation, which may be important for immune defenses against HSV infections in vitro and in vivo. PMID:26082771

  14. Studies of Two Subpopulations of Human Lymphocytes Differing in Responsiveness to Concanavalin A

    PubMed Central

    Boldt, David; Skinner, Sister Ann Marie; Kornfeld, Stuart

    1972-01-01

    We have identified two populations of human lymphocytes differing in responsiveness to the plant mitogen concanavalin A (Con-A). When peripheral blood lymphocytes are passed through a nylon column a population of lymphocytes highly responsive to Con-A adheres to the fibers while a second population of cells relatively unresponsive to Con-A emerges from the column. The untreated peripheral blood lymphocytes are termed “unfiltered” cells while the lymphocytes which pass through the column are termed “filtered” cells. Under standard assay conditions the Con-A-stimulated DNA synthesis is 6.5-fold greater, and the percentage blast formation is four-to fivefold greater in the unfiltered than in the filtered population. Mixing unfiltered with filtered cells fails to induce responsiveness in the latter indicating that a “helper” cell is not involved. The failure of filtered cells to respond to Con-A is specific for that mitogen since both populations respond nearly equally to erythroagglutinating phytohemagglutinin (E-PHA) and the poke weed mitogen (PWM). Binding studies with Con-A-131I demonstrate that the unfiltered population possesses approximately three times as many Con-A receptor sites per cell as the filtered cells, although both cell populations bind the mitogen with the same affinity (apparent association constant [K] of 1.67 × 106m−1). The relationship between Con-A binding and lymphocyte activation was determined by measuring the effect on DNA synthesis of incubating the two lymphocyte populations with increasing amounts of Con-A. The concentration of Con-A required for half-maximal stimulation of DNA synthesis was 5-14 times greater for the filtered cells. However in the presence of very high Con-A concentrations the filtered cells achieved a maximal rate of DNA synthesis approaching that of the unfiltered population. These data implicate the decreased number of Con-A receptor sites on the filtered cells in their failure to respond to low

  15. The Identification of Mitogen Responding Subpopulations of Human Lymphocytes by Flow Polarimeter Fluorescence Measurements.

    NASA Astrophysics Data System (ADS)

    Chan, Sandra Lynn

    I have developed a method to identify the mitogen responding subpopulation of human peripheral blood lymphocytes. This method employs a flow polarimeter to measure the distribution of the intensity and the polarization of intracellular fluorescein fluorescence in suspensions of mononuclear cells isolated on density gradients from the peripheral blood of donors. I have used the change in the fluorescence of cells exposed to the mitogens PHA and Con A to identify the responding cells and to quantitate this number. I have found that for most donors, the responding cells constitute about 20-40% of the lymphocyte population. The percent of responding cells decreases to zero in patients with acute lymphocytic leukemia (2 patients) and chronic lymphocyte leukemia (10 patients). For a variety of patients with other types of cancer, the responding fraction was not significantly different from healthy controls. Moreover, the number of responding cells does not appear to be age dependent in the age range of 20-80 years. I also found that the change in fluorescence polarization correlated strongly with changes in fluorescence intensity induced by mitogens--the number of responding cells, therefore can be estimated either from the intensity or polarization distributions. The shapes of fluorescence distributions depend strongly on a number of variables including the composition and density of the lymphocyte isolating medium, the mitogen and dye concentrations, the length of incubation with mitogen or dye, and the potassium, calcium, and magnesium concentrations in the medium. In the case of fluorescein, I have worked out a methodology that allows a consistent estimate of the responding lymphocyte number. I have also investigated the use of the dye carbocyanine for the same purpose. This dye presumably identifies the mitogen responding lymphocytes on the basis of changes in membrane potential. The results with carbocyanine were found to depend on a number of variables and I could

  16. Expansion of human tumor infiltrating lymphocytes for use in immunotherapy trials.

    PubMed

    Topalian, S L; Muul, L M; Solomon, D; Rosenberg, S A

    1987-08-24

    The potential utility of tumor-infiltrating lymphocytes (TIL) in the adoptive immunotherapy of human tumors has been suggested by murine experiments showing these cells to be 50-100 times more powerful than LAK cells in treating advanced metastatic disease. A method for the large-scale expansion of human TIL for the use of these cells in clinical trials is described in this report. TIL were successfully expanded on an experimental scale from 24 of 25 consecutive human tumors, including six melanomas, ten sarcomas, and eight adenocarcinomas. Tumors were digested enzymatically to yield single cell suspensions which were cultured in RPMI 1640 medium with 10% human serum and 1000 U/ml recombinant interleukin-2. Lymphocytes constituted from 3% to 74% of single cell tumor suspensions, and expanded from 2.9-fold to 9.1 X 10(8)-fold over a culture period ranging from 14 to 100 days. Nine of 24 TIL cultures lysed fresh autologous tumor targets in 4 h chromium release assays. Cell surface phenotyping identified cultured TIL as activated cytotoxic/suppressor T cells. Subsequently, large-scale expansion of TIL was successful in generating more than 10(10) lymphocytes in five of eight consecutive cases. Clinical trials employing the adoptive transfer of expanded TIL to patients with metastatic disease have begun. PMID:3305708

  17. Activation of Ca2+-dependent K+ channels in human B lymphocytes by anti-immunoglobulin.

    PubMed Central

    MacDougall, S L; Grinstein, S; Gelfand, E W

    1988-01-01

    Many mammalian cell types exhibit Ca2+-dependent K+ channels, and activation of these channels by increasing intracellular calcium generally leads to a hyperpolarization of the plasma membrane. Their presence in B lymphocytes is as yet uncertain. Crosslinking Ig on the surface of B lymphocytes is known to increase the level of free cytoplasmic calcium ([Ca2+]i). However, rather than hyperpolarization, a depolarization has been reported to occur after treatment of B lymphocytes with anti-Ig. To determine if Ca2+-dependent K+ channels are present in B lymphocytes, and to examine the relationship between intracellular free calcium and membrane potential, we monitored [Ca2+]i by means of indo-1 and transmembrane potential using bis(1,3-diethylthiobarbituric)trimethine oxonol in human tonsillar B cells activated by anti-IgM. Treatment with anti-IgM induced a biphasic increase in [Ca2+]i and a simultaneous hyperpolarization. A similar hyperpolarization was induced by ionomycin, a Ca2+ ionophore. Delaying the development of the [Ca2+]i response by increasing the cytoplasmic Ca2+-buffering power delayed the hyperpolarization. Conversely, eliminating the sustained phase of the [Ca2+]i response by omission of external Ca2+ abolished the prolonged hyperpolarization. In fact, a sizable Na+-dependent depolarization was unmasked. This study demonstrates that in human B lymphocytes, Ca2+-dependent K+ channels can be activated by crosslinking of surface IgM. Moreover, it is likely that, by analogy with voltage-sensitive Ca2+ channels, Na+ can permeate through these ligand-gated Ca2+ "channels" in the absence of extracellular Ca2+. PMID:2448342

  18. PGC1α Expression Defines a Subset of Human Melanoma Tumors with Increased Mitochondrial Capacity and Resistance to Oxidative Stress

    PubMed Central

    Vazquez, Francisca; Lim, Ji-Hong; Chim, Helen; Bhalla, Kavita; Girnun, Geoff; Pierce, Kerry; Clish, Clary B.; Granter, Scott R.; Widlund, Hans R.; Spiegelman, Bruce M.; Puigserver, Pere

    2013-01-01

    SUMMARY Cancer cells reprogram their metabolism using different strategies to meet energy and anabolic demands to maintain growth and survival. Understanding the molecular and genetic determinants of these metabolic programs is critical to successfully exploit them for therapy. Here, we report that the oncogenic melanocyte lineage-specification transcription factor MITF drives PGC1α (PPARGC1A) overexpression in a subset of human melanomas and derived cell lines. Functionally, PGC1α positive melanoma cells exhibit increased mitochondrial energy metabolism and ROS detoxification capacities that enables survival under oxidative stress conditions. Conversely, PGC1α negative melanoma cells are more glycolytic and sensitive to ROS-inducing drugs. These results demonstrate that differences in PGC1α levels in melanoma tumors have a profound impact in their metabolism, biology and drug sensitivity. PMID:23416000

  19. Blister fluid T lymphocytes during toxic epidermal necrolysis are functional cytotoxic cells which express human natural killer (NK) inhibitory receptors

    PubMed Central

    Le Cleach, L; Delaire, S; Boumsell, L; Bagot, M; Bourgault-Villada, I; Bensussan, A; Roujeau, J C

    2000-01-01

    Toxic epidermal necrolysis (TEN) is a rare life-threatening adverse drug reaction characterized by a massive destruction of the epidermis. Immunohistological studies of skin biopsies of TEN showed infiltrates of predominantly CD8+ T lymphocytes even though other authors reported a prominent involvement of cells of the monocyte-macrophage lineage. The aim of this study was to characterize phenotypically and functionally the cells present in the cutaneous blister fluid of four patients with TEN. We first determined that lymphocytes were predominant in blister fluid obtained early, while monocytes/macrophages later became the most important population. We then showed that this lymphocyte population, mainly CD3+CD8+, corresponded to a peculiar cell subset as they expressed cutaneous leucocyte antigen, killer inhibitory receptors KIR/KAR and failed to express CD28 molecule. Functionally, we determined that blister T lymphocytes had a cytotoxic T lymphocyte (CTL)- and NK-like cytotoxicity. The role of this cytotoxic lymphocyte population present at the site of lesions during TEN remains to be understood. PMID:10606987

  20. Study of apoptosis in human lymphocytes by toxic substances implicated in toxic oil syndrome.

    PubMed

    Gallardo, S; Cárdaba, B; del Pozo, V; de Andrés, B; Cortegano, I; Jurado, A; Tramón, P; Palomino, P; Lahoz, C

    1997-03-14

    Toxic Oil Syndrome is a multisystemic disease that occurred in epidemic proportions in Spain in 1981 caused by the ingestion of rapeseed oil denatured with aniline. Several data implicate T cells in the pathogenesis of the disease. We evaluated the mechanisms of cytotoxicity in human lymphocytes of TOS-related products: aniline, 3-(N-phenylamino)-1,2-propanediol and its mono- and di-oleyl esters and eosinophilia myalgia-related product such as 3-(phenylamino)-L-alanine, which is chemically similar to 3-(N-phenylamino)-1,2-propanediol, and has been found in manufactured L-tryptophan. Our results show that only di-oleyl ester of 3-(N-phenylamino)-1,2-propanediol induces apoptosis in human lymphocytes, in a concentration and time-dependent way, confirmed by morphology, expression of phosphatidylserine in membrane and analysis of DNA degradation. PMID:9074655

  1. Protective effects of black tea extract against oxidative DNA damage in human lymphocytes.

    PubMed

    Ježovičová, Miriam; Koňariková, Katarína; Ďuračková, Zdeňka; Keresteš, Ján; Králik, Gabriel; Žitňanová, Ingrid

    2016-02-01

    The aim of the present study was to examine the genoprotective and radioprotective effects of black tea extract (BTE) against the induction of single strand DNA breaks in human lymphocytes subjected to hydrogen peroxide (H2O2) or gamma-rays (2 Gy dose). Lymphocytes were incubated with or without different concentrations of BTE (0.005-500 µg/ml) for 30 min, followed by treatment with or without H2O2 (0.088 µmol/l) for 5 min. To examine the radioprotective effect of BTE, the lymphocytes were incubated with or without BTE for 30 and 60 min prior to and following in vitro irradiation. Oxidative damage to DNA was monitored using a comet assay. BTE at lower concentrations prevented H2O2-induced DNA damage. An increase in BTE concentrations resulted in increased formation of single strand DNA breaks. BTE also exerted significant protective effects against gamma radiation-induced total DNA damage in healthy lymphocytes during their 30 or 60 min incubation with BTE prior to or following irradiation. Therefore, the protective effect of BTE against irradiation was time-dependent. The results contribute to the research on potential beneficial effects of natural compounds, such as BTE, in cancer and its protective effects of normal tissue during radiation therapy. PMID:26718244

  2. In vitro protection of human lymphocytes from toxic effects of chlorpyrifos by selenium-enriched medicines

    PubMed Central

    Navaei-Nigjeh, Mona; Asadi, Hamidreza; Baeeri, Maryam; Pedram, Sahar; Rezvanfar, Mohammad Amin; Mohammadirad, Azadeh; Abdollahi, Mohammad

    2015-01-01

    Objective(s): Chlorpyrifos (CP) is a broad-spectrum organophosphorus pesticide used extensively in agricultural and domestic pest control, accounting for 50% of the global insecticidal use. In the present study, protective effects of two selenium-enriched strong antioxidative medicines IMOD and Angipars were examined in human lymphocytes treated with CP in vitro. Materials and Methods: Isolated lymphocytes were exposed to 12 µg/ml CP either alone or in combination with effective doses (ED50) of IMOD (0.2 µg/ml) and Angipars (1 µg/ml). After 3 days incubation, the viability and oxidative stress markers including cellular lipid peroxidation (LPO), myeloperoxidase (MPO), total thiol molecules (TTM), and total antioxidant power (TAP) were evaluated. Also, the levels of tumor necrosis factor-α (TNF-α), as inflammatory index along with acetylcholinesterase (AChE) activity and cell apoptosis were assessed by flow cytometry. Results: Results indicated that effective doses of IMOD and Angipars reduced CP-exposed lymphocyte mortality rate along with oxidative stress. Both agents restored CP-induced elevation of TNF-α and protected the lymphocytes from CP-induced apoptosis and necrosis. Conclusion: Overall, results confirm that IMOD and Angipars reduce the toxic effects associated with CP through free radical scavenging and protection from apoptosis and necrosis. PMID:25945242

  3. In human alloreactive CD4+ T-cells, dichloroacetate inhibits aerobic glycolysis, induces apoptosis and favors differentiation towards the regulatory T-cell subset instead of effector T-cell subsets.

    PubMed

    Eleftheriadis, Theodoros; Sounidaki, Maria; Pissas, Georgios; Antoniadi, Georgia; Liakopoulos, Vassilios; Stefanidis, Ioannis

    2016-04-01

    Although kidney transplantation is the best therapy for end-stage renal disease, rejection remains a concern, and currently available immunosuppressive agents contribute to morbidity and mortality. Thus, novel immunosuppressive drugs are required. Dichloroacetate (DCA) is already used in the treatment of congenital lactic acidosis and characterized by limited toxicity. As DCA inhibits aerobic glycolysis, which is a prerequisite for CD4+ T-cell proliferation and differentiation into effector T-cells, its possible immunosuppressive role in mixed lymphocyte reaction (MLR), a model of alloreactivity, was investigated. Glucose and lactate concentrations were measured in the supernatants, and cell proliferation was assessed immunoenzymatically. CD4+ T‑cells were then isolated from the MLRs and the expression of cleaved caspase‑3, various enzymes involved in glycolysis, and the signature transcription factors of CD4+ T‑cell subsets were evaluated by western blotting. In MLRs, DCA decreased glucose consumption and aerobic glycolysis, while it exerted a negligible effect on cell proliferation. In CD4+ T‑cells, DCA induced apoptosis, and decreased the expression of glucose trasporter‑1, hexokinase II, lactate dehydrogenase‑A and phosphorylated pyruvate dehydrogenase, while it increased total pyruvate dehydrogenase. In addition, DCA increased the expression of transcription factor forkhead box P3, whereas it decreased the expression of T‑box transcription factor TBX21, trans‑acting T-cell-specific transcription factor GATA‑3 and retinoic acid receptor related orphan receptor‑γt. In conclusion, in alloreactive CD4+ T‑cells, DCA inhibits aerobic glycolysis, induces apoptosis and favors differentiation towards the regulatory T‑cell subset. These characteristics render it a promising immunosuppressive agent in the field of transplantation. PMID:26935268

  4. CD8+ T lymphocytes of patients with AIDS maintain normal broad cytolytic function despite the loss of human immunodeficiency virus-specific cytotoxicity.

    PubMed Central

    Pantaleo, G; De Maria, A; Koenig, S; Butini, L; Moss, B; Baseler, M; Lane, H C; Fauci, A S

    1990-01-01

    In this study, we have investigated the potential mechanisms responsible for the loss of human immunodeficiency virus type 1 (HIV-1)-specific cytolytic activity in the advanced stages of HIV-1 infection. We have demonstrated that HIV-1-specific cytotoxic T lymphocytes are predominantly contained within the CD8+DR+ subset. Furthermore, we have shown by a redirected killing assay that there is a dichotomy between HIV-1-specific cytolytic activity and broad cytolytic potential since the cytolytic machinery of CD8+DR+ cells is still functioning even in patients with AIDS who have lost their HIV-1-specific cytolytic activity. In addition, by comparative analysis of these two types of cytolytic activity over time we have demonstrated a progressive loss of HIV-1-specific cytolytic activity in the advanced stages of the disease, whereas the cytolytic potential remained unchanged regardless of the clinical stage. As previously shown in patients with AIDS, even in asymptomatic HIV-1-seropositive patients, CD8+DR+ cells from the same patient, compared to CD8+DR- lymphocytes, showed a substantial reduction in their ability to proliferate in vitro in response to different stimuli, such as mitogens (phytohemagglutinin and phorbol 12-myristate 13-acetate) and monoclonal antibodies directed against CD3, CD2, and CD28 molecules, and displayed a defective clonogenic potential. Thus, on the basis of these results we propose that the loss of HIV-1-specific cytolytic activity in HIV-1-infected individuals may result at least in part from a progressive decrease in the pool of HIV-1-specific cytotoxic T lymphocytes belonging to the CD8+DR+ subset whose ability to expand has been impaired. Images PMID:2112749

  5. Microgravity simulations with human lymphocytes in the free fall machine and in the random positioning machine

    NASA Technical Reports Server (NTRS)

    Schwarzenberg, M.; Pippia, P.; Meloni, M. A.; Cossu, G.; Cogoli-Greuter, M.; Cogoli, A.

    1998-01-01

    The purpose of this paper is to present the results obtained in our laboratory with both instruments, the FFM [free fall machine] and the RPM [random positioning machine], to compare them with the data from earlier experiments with human lymphocytes conducted in the FRC [fast rotating clinostat] and in space. Furthermore, the suitability of the FFM and RPM for research in gravitational cell biology is discussed.

  6. Ectopic lymphokine gene expression in human peripheral blood lymphocytes in vivo

    SciTech Connect

    Chambers, C.A.; Kang, Joonsoo; Hozumi, Nobumichi Mount Sinai Hospital, Toronto, Ontario )

    1992-02-01

    An animal model to study the effects of ectopic expression of cytokines involved in cell growth and differentiation has been established. Retrovirus vectors containing the human interleukin 6 cDNA were used to produce high titer virus-producing lines. Human peripheral blood lymphocytes (hPBLs) were successfully infected with the retrovirus and engrafted into severe combined immunodeficient mice. The majority of the animals were engrafted with hPBLs, as determined by the presence of human glucose phosphate isomerase. Furthermore, six of seven mice engrafted with hPBLs infected with high titer virus and detectable hPBLs present in the spleen expressed the retroviral human interleukin 6 gene. Importantly, human interleukin 6 protein was expressed at physiologically significant levels in these mice. These results demonstrate that models for human disease and immunotherapy involving retrovirus-mediated gene transfer into human cells can be developed in mice.

  7. The role of the enzymatic antioxidant system in cylindrospermopsin-induced toxicity in human lymphocytes.

    PubMed

    Poniedziałek, Barbara; Rzymski, Piotr; Karczewski, Jacek

    2015-08-01

    Cylindrospermopsin (CYN) is known to induce cytotoxic effects in eukaryotic cells although the exact mechanism underlying these alterations is not fully explained. Given that CYN was previously found to decrease the proliferation of human lymphocytes through DNA damage and cell cycle arrest followed by an increase in the apoptotic rate, the present study evaluated the possible involvement of reactive oxygen species (ROS) and oxidative stress in these cytopathic responses. The status of enzymatic antioxidants: superoxide dismutase (SOD), glutathione peroxidase (GPx) and catalase (CAT) as well as level of lipid peroxidation (LO) under CYN influence in human lymphocytes were also studied. It was found that CYN exposure (0.01-1.0 μg/ml) induces a concentration-dependent increase in H2O2 content within a time as short as 0.5h, reaching its maximum level after 3 and 6h. The highest H2O2 content was accompanied by a significant decrease of SOD and CAT activity and an elevated level of GPx. Moreover, CYN treatment resulted in a detectable increase in LO. We conclude that ROS and the products of LO play an essential role in CYN-induced toxicity in human lymphocytes. Our study helps to elucidate the sequence of events caused by CYN in eukaryotic cells and explain the background for previously observed cytopathic responses. PMID:25863213

  8. Deoxynivalenol induced oxidative stress and genotoxicity in human peripheral blood lymphocytes.

    PubMed

    Yang, Wei; Yu, Miao; Fu, Juan; Bao, Wei; Wang, Di; Hao, Liping; Yao, Ping; Nüssler, Andreas K; Yan, Hong; Liu, Liegang

    2014-02-01

    Deoxynivalenol (DON) is one of the most common mycotoxins. The aim of this study consists in using diverse cellular and molecular assays to evaluate cytotoxicity, genotoxicity as well as oxidative damage and to investigate their mechanisms in human peripheral blood lymphocytes. The human lymphocytes were cultured in eight different doses of DON (0, 6.25, 12.5, 25, 50, 100, 250 and 500 ng/mL) during 6, 12 and 24 h. DON was able to decrease cell viability and cause damage to the membrane, the chromosomes or the DNA at all times of culture. It was also able to induce lipid peroxidation and raise the levels of 8-OHdG and ROS in 6, 12 and 24 h. The results of the RT-PCR and the Western Blot indicated that DON is able to enhance mRNA or protein expressions of DNA repair genes and HO-1 in 6 h and to inhibit these expressions in 24 h. DON potentially triggers genotoxicity in human lymphocytes. This mechanism is probably related to depletion of antioxidase and oxidative damage to the DNA that reduced expression of HO-1, thereby inhibiting the ability of DNA repair. PMID:24355168

  9. Physical association and functional interaction between beta1 integrin and CD98 on human T lymphocytes

    NASA Technical Reports Server (NTRS)

    Miyamoto, Yuko J.; Mitchell, Jason S.; McIntyre, Bradley W.

    2003-01-01

    CD98 is a cell surface protein previously characterized as a cell activation marker, an amino acid transporter, and has recently been implicated in integrin-related functions. Integrins are cell surface proteins, important for homotypic cell aggregation, cell adhesion, and coactivation of T lymphocytes. We have previously shown that the anti-CD98 mAb 80A10, when coimmobilized with anti-CD3 mAb OKT3, is able to mediate human T cell coactivation that is inhibited by anti-beta1 integrin specific mAb 18D3. These results indicated a functional association of CD98 and beta1 integrin signaling but left open the question of a physical association. We now show the induction of homotypic aggregation through CD98 among human T cells and this aggregation was inhibited by anti-beta1 integrin mAb. Therefore, CD98-dependent lymphocyte proliferation and adhesion may involve integrins. Competitive binding assays and fluorescence colocalization analysis suggested that CD98 and beta1 integrin were physically associated. Differential extraction techniques and immunoprecipitations provided the first evidence that the alpha4beta1 integrin and CD98 are specifically associated on human T lymphocytes.

  10. Enhanced lymphocyte longevity and absence of proliferation and lymphocyte apoptosis in Quilty effects of human heart allografts.

    PubMed Central

    Dong, C.; Winters, G. L.; Wilson, J. E.; McManus, B. M.

    1997-01-01

    "Quilty effect" (QE) is a common and problematic observation in endomyocardial biopsy specimens from patients after cardiac transplantation. The origin, fate, and significance of QE cellular elements are unknown. Twenty-six paraffin-embedded endomyocardial biopsy specimens with QE (five QE As and twenty-one QE Bs) from twenty-two cardiac allografts were studied by immunohistochemistry for expression of Bcl-2, Fas antigen, proliferating cell nuclear antigen (PCNA), perforin, T cells (UCHL-1), macrophages (CD68), and apoptosis by in situ terminal deoxyribonucleotide transferase (TdT)-mediated dUTP nick end labeling (TUNEL). Approximately 50% of the lymphocytes present, mainly in the deeper region of 20 of 21 QE Bs and all 5 QE As, expressed Bcl-2 in a pseudo-nodular pattern surrounding high endothelial venules. Fas expression was detected in lymphocytes in 20 of 21 QE Bs and 5 QE As in a similar pattern to Bcl-2. However, endothelial cells and macrophages were Bcl-2 negative, whereas both cell types were Fas positive. Perforin was negative in nearly all lymphocytes. TUNEL staining revealed that lymphocytes in QEs did not undergo apoptosis; however, TUNEL positivity was observed in approximately 70% of endothelial cells and macrophages and certain adjacent cardiac myocytes in 20 of 21 QE Bs and 5 QE As. One large QE B with a germinal center was noted. Germinal center cells expressed PCNA intensely but were negative for Bcl-2, Fas, and TUNEL. Cells surrounding the germinal center expressed abundant Bcl-2. The following conclusions were drawn. 1) Apoptosis does not occur in lymphocytes in QE where enhanced Bcl-2 (apoptosis inhibitor) and Fas antigen (apoptosis inducer) are expressed. 2) PCNA negativity indicates that QE lymphocytes may not proliferate, and perforin negativity indicates that they may not exhibit perforin-based cytotoxicity. We propose that there may be a relationship between the longevity of lymphocytes in QE and the absence of apoptosis. Images Figure 1

  11. Novel Observations From Next-Generation RNA Sequencing of Highly Purified Human Adult and Fetal Islet Cell Subsets.

    PubMed

    Blodgett, David M; Nowosielska, Anetta; Afik, Shaked; Pechhold, Susanne; Cura, Anthony J; Kennedy, Norman J; Kim, Soyoung; Kucukural, Alper; Davis, Roger J; Kent, Sally C; Greiner, Dale L; Garber, Manuel G; Harlan, David M; diIorio, Philip

    2015-09-01

    Understanding distinct gene expression patterns of normal adult and developing fetal human pancreatic α- and β-cells is crucial for developing stem cell therapies, islet regeneration strategies, and therapies designed to increase β-cell function in patients with diabetes (type 1 or 2). Toward that end, we have developed methods to highly purify α-, β-, and δ-cells from human fetal and adult pancreata by intracellular staining for the cell-specific hormone content, sorting the subpopulations by flow cytometry, and, using next-generation RNA sequencing, we report the detailed transcriptomes of fetal and adult α- and β-cells. We observed that human islet composition was not influenced by age, sex, or BMI, and transcripts for inflammatory gene products were noted in fetal β-cells. In addition, within highly purified adult glucagon-expressing α-cells, we observed surprisingly high insulin mRNA expression, but not insulin protein expression. This transcriptome analysis from highly purified islet α- and β-cell subsets from fetal and adult pancreata offers clear implications for strategies that seek to increase insulin expression in type 1 and type 2 diabetes. PMID:25931473

  12. Unique and shared signaling pathways cooperate to regulate the differentiation of human CD4+ T cells into distinct effector subsets.

    PubMed

    Ma, Cindy S; Wong, Natalie; Rao, Geetha; Nguyen, Akira; Avery, Danielle T; Payne, Kathryn; Torpy, James; O'Young, Patrick; Deenick, Elissa; Bustamante, Jacinta; Puel, Anne; Okada, Satoshi; Kobayashi, Masao; Martinez-Barricarte, Ruben; Elliott, Michael; Sebnem Kilic, Sara; El Baghdadi, Jamila; Minegishi, Yoshiyuki; Bousfiha, Aziz; Robertson, Nic; Hambleton, Sophie; Arkwright, Peter D; French, Martyn; Blincoe, Annaliesse K; Hsu, Peter; Campbell, Dianne E; Stormon, Michael O; Wong, Melanie; Adelstein, Stephen; Fulcher, David A; Cook, Matthew C; Stepensky, Polina; Boztug, Kaan; Beier, Rita; Ikincioğullari, Aydan; Ziegler, John B; Gray, Paul; Picard, Capucine; Boisson-Dupuis, Stéphanie; Phan, Tri Giang; Grimbacher, Bodo; Warnatz, Klaus; Holland, Steven M; Uzel, Gulbu; Casanova, Jean-Laurent; Tangye, Stuart G

    2016-07-25

    Naive CD4(+) T cells differentiate into specific effector subsets-Th1, Th2, Th17, and T follicular helper (Tfh)-that provide immunity against pathogen infection. The signaling pathways involved in generating these effector cells are partially known. However, the effects of mutations underlying human primary immunodeficiencies on these processes, and how they compromise specific immune responses, remain unresolved. By studying individuals with mutations in key signaling pathways, we identified nonredundant pathways regulating human CD4(+) T cell differentiation in vitro. IL12Rβ1/TYK2 and IFN-γR/STAT1 function in a feed-forward loop to induce Th1 cells, whereas IL-21/IL-21R/STAT3 signaling is required for Th17, Tfh, and IL-10-secreting cells. IL12Rβ1/TYK2 and NEMO are also required for Th17 induction. Strikingly, gain-of-function STAT1 mutations recapitulated the impact of dominant-negative STAT3 mutations on Tfh and Th17 cells, revealing a putative inhibitory effect of hypermorphic STAT1 over STAT3. These findings provide mechanistic insight into the requirements for human T cell effector function, and explain clinical manifestations of these immunodeficient conditions. Furthermore, they identify molecules that could be targeted to modulate CD4(+) T cell effector function in the settings of infection, vaccination, or immune dysregulation. PMID:27401342

  13. Modeled microgravity-induced protein kinase C isoform expression in human lymphocytes

    NASA Technical Reports Server (NTRS)

    Sundaresan, A.; Risin, D.; Pellis, N. R.

    2004-01-01

    In long-term space travel, the crew is exposed to microgravity and radiation that invoke potential hazards to the immune system. T cell activation is a critical step in the immune response. Receptor-mediated signaling is inhibited in both microgravity and modeled microgravity (MMG) as reflected by diminished DNA synthesis in peripheral blood lymphocytes and their locomotion through gelled type I collagen. Direct activation of protein kinase C (PKC) bypassing cell surface events using the phorbol ester PMA rescues MMG-inhibited lymphocyte activation and locomotion, whereas the calcium ionophore ionomycin had no rescue effect. Thus calcium-independent PKC isoforms may be affected in MMG-induced locomotion inhibition and rescue. Both calcium-dependent isoforms and calcium-independent PKC isoforms were investigated to assess their expression in lymphocytes in 1 g and MMG culture. Human lymphocytes were cultured and harvested at 24, 48, 72, and 96 h, and serial samples were assessed for locomotion by using type I collagen and expression of PKC isoforms. Expression of PKC-alpha, -delta, and -epsilon was assessed by RT-PCR, flow cytometry, and immunoblotting. Results indicated that PKC isoforms delta and epsilon were downregulated by >50% at the transcriptional and translational levels in MMG-cultured lymphocytes compared with 1-g controls. Events upstream of PKC, such as phosphorylation of phospholipase Cgamma in MMG, revealed accumulation of inactive enzyme. Depressed calcium-independent PKC isoforms may be a consequence of an upstream lesion in the signal transduction pathway. The differential response among calcium-dependent and calcium-independent isoforms may actually result from MMG intrusion events earlier than PKC, but after ligand-receptor interaction.

  14. Modeled Microgravity-Induced Protein Kinase C Isoform Expression in Human Lymphocytes

    NASA Technical Reports Server (NTRS)

    Sundaresan, A.; Risin, D.; Pellis, N. R.

    2003-01-01

    In long-term space travel, the crew is exposed to microgravity and radiation that invoke potential hazards to the immune system. T cell activation is a critical step in the immune response. Receptor-mediated signaling is inhibited both in microgravity and modeled microgravity (MMG) as reflected in diminished DNA synthess in peripheral blood lymphocytes and their locomotion through gelled type 1 collagen. Direct activation of Protein Kinase C (PKC) bypassing cell surface events using the phorbol ester PMA rescues MMG-inhibited lymphocyte activation and locomotion, whereas calcium ionophore ionomycin had no rescue effect. Thus calcium-independent PKC isoforms may be affected in MMG-induced locomotion inhibition and rescue. Both calcium-dependent isoforms and calcium-independent PKC isoforms were investigated to assess their expression in lymphocytes in 19 and MMG-culture. Human lymphocytes were cultured and harvested at 24, 48, 72 and 96 hours and serial samples assessed for locomotion using type I collagen and expression of PKC isoforms. Expression of PKC-alpha, -delta and -epsilon was assessed by RT-PCR, flow cytometry and immunoblotting. Results indicated that PKC isoforms delta and epsilon were down-regulated by more than 50% at the transcriptional and translational levels in MMG-cultured lymphocytes compared with 19 controls. Events upstream of PKC such as phosphorylation of Phospholipase C(gamma) (PLC-gamma) in MMG, revealed accumulation of inactive enzyme. Depressed Ca++ -independent PKC isoforms may be a consequence of an upstream lesion in the signal transduction pathway. The differential response among calcium-dependent and calcium-independent isoforms may actually result from MMG intrusion events earlier than, but after ligand-receptor interaction. Keywords: Signal transduction, locomotion, immunity

  15. Studies on human blood lymphocytes with iC3b (type 3) complement receptors: III. Abnormalities in patients with active systemic lupus erythematosus.

    PubMed Central

    Gray, J D; Lash, A; Bakke, A C; Kitridou, R C; Horwitz, D A

    1987-01-01

    Lymphocytes displaying iC3b (Type 3) complement receptors (CR3) were quantified by flow cytometry in patients with systemic lupus erythematosus. The percentages and absolute numbers were compared to age and sex matched controls. Total CR3+ lymphocytes identified by the monoclonal antibodies OKM1 or Leu 15 were significantly decreased in patients with symptomatic arthritis, serositis or vasculitis and those with lupus nephritis, whereas values for CR3+ lymphocytes in patients with inactive disease were similar to normal donors. The phenotype of CR3+ lymphocytes was markedly different in patients with active SLE. In normals granular lymphocytes bearing Fc receptors for IgG (L cells) comprised two-thirds of CR3+ lymphocytes. However, in SLE this subset was reduced to 20% and there was a corresponding increase in CR3+ lymphocytes co-expressing the T3 marker. Percentages of CR3 T4+ but not CR3+ T8+ lymphocytes were significantly increased in SLE. Although patients with active disease were lymphopenic, absolute numbers of CR3+ lymphocytes co-expressing T cell markers were similar to normal controls. Since L cells are non-specific suppressors of Ig production, the reduction of this subset along with the increase in CR3 T4+ cells could contribute to unregulated antibody production characteristic of SLE. PMID:2955974

  16. Fas-mediated apoptosis of melanoma cells and infiltrating lymphocytes in human malignant melanomas.

    PubMed

    Shukuwa, Tetsuo; Katayama, Ichiro; Koji, Takehiko

    2002-04-01

    In a rodent system, melanoma cells expressing Fas ligand (FasL) could kill Fas-positive lymphocytes, suggesting that FasL expression was an essential factor for melanoma cell survival in vivo. These findings led us to investigate apoptosis, and to histochemically analyze involvement of Fas and FasL in the induction of apoptosis, in human malignant melanoma tissues. The percentages of terminal deoxynucleotidyl transferase-mediated biotin-dUTP nick end-labeling (TUNEL)-positive melanoma cells and of proliferating cell nuclear antigen (PCNA)-positive melanoma cells in melanoma tissues (n = 22) were greater than those in melanocytes in uninvolved skin (n = 6) and nevus cells in nevi tissues (n = 9). The infiltrating lymphocytes around melanomas were also TUNEL positive. Immunohistochemistry revealed expression of Fas and FasL in melanoma cells and lymphocytes, whereas no Fas or FasL expression was detected in normal skin melanocytes and nevus cells. There was significant correlation between Fas-positive indices and TUNEL indices in melanoma tissues. Moreover, TUNEL-, Fas-, and FasL-positive indices of melanoma cells from patients with Stage 3 melanomas were significantly lower than those with Stage 2 melanomas. The PCNA index of Stage 1 melanoma was significantly lower than that of the other stages, although the difference of PCNA index was insignificant among Stages 2 to 4. Among Stages 1 to 4, there was no difference in the PCNA, TUNEL-, and Fas-positive indices of lymphocytes, although the FasL-positive index of lymphocytes from Stage 3 melanomas was significantly lower than in that from Stage 2. These data reveal that melanoma cells and infiltrating lymphocytes have the potential to induce their own apoptosis regulated by Fas and FasL in an autocrine and/or paracrine fashion and that the decline of Fas-mediated apoptosis of melanoma cells, rather than the apoptosis of infiltrating lymphocytes, may affect the prognosis of melanoma patients, possibly through the

  17. Comparison of Extraintestinal Pathogenic Escherichia coli Strains from Human and Avian Sources Reveals a Mixed Subset Representing Potential Zoonotic Pathogens▿

    PubMed Central

    Johnson, Timothy J.; Wannemuehler, Yvonne; Johnson, Sara J.; Stell, Adam L.; Doetkott, Curt; Johnson, James R.; Kim, Kwang S.; Spanjaard, Lodewijk; Nolan, Lisa K.

    2008-01-01

    Since extraintestinal pathogenic Escherichia coli (ExPEC) strains from human and avian hosts encounter similar challenges in establishing infection in extraintestinal locations, they may share similar contents of virulence genes and capacities to cause disease. In the present study, 1,074 ExPEC isolates were classified by phylogenetic group and possession of 67 other traits, including virulence-associated genes and plasmid replicon types. These ExPEC isolates included 452 avian pathogenic E. coli strains from avian colibacillosis, 91 neonatal meningitis E. coli (NMEC) strains causing human neonatal meningitis, and 531 uropathogenic E. coli strains from human urinary tract infections. Cluster analysis of the data revealed that most members of each subpathotype represent a genetically distinct group and have distinguishing characteristics. However, a genotyping cluster containing 108 ExPEC isolates was identified, heavily mixed with regard to subpathotype, in which there was substantial trait overlap. Many of the isolates within this cluster belonged to the O1, O2, or O18 serogroup. Also, 58% belonged to the ST95 multilocus sequence typing group, and over 90% of them were assigned to the B2 phylogenetic group typical of human ExPEC strains. This cluster contained strains with a high number of both chromosome- and plasmid-associated ExPEC genes. Further characterization of this ExPEC subset with zoonotic potential urges future studies exploring the potential for the transmission of certain ExPEC strains between humans and animals. Also, the widespread occurrence of plasmids among NMEC strains and members of the mixed cluster suggests that plasmid-mediated virulence in these pathotypes warrants further attention. PMID:18820066

  18. Cardiomyocyte marker expression in a human lymphocyte cell line using mouse cardiomyocyte extract.

    PubMed

    Vojdani, Zahra; Tavakolinejad, Sima; Talaei-Khozani, Tahereh; Esmaeilpour, Tahereh; Rasooli, Manuchehr

    2011-03-01

    Cell transplantation shows potential for the treatment of cardiac diseases. Embryonic stem cells, cord blood and mesenchymal stem cells have been suggested as sources for transplantation therapy. Because of some technical limitations with the use of stem cells, transdifferentiation of fully differentiated cells is a potentially useful alternative. We investigated whether human peripheral blood cells could transdifferentiate into cardiomyocyte. Transdifferentiation was induced in a human B lymphocyte cell line (Raji). Cardiomyocyte extract was prepared from adult mouse cardiomyocytes. The cells were treated with 5-aza-2-deoxycytidine and trichostatin A, permeabilized with streptolysin O, and exposed to the mouse cardiomyocyte extract. They were cultured for 10 days, 3 weeks and 4 weeks. Cardiomyocyte markers were detected with immunohistochemistry and flow cytometry. Immunocytochemistry revealed that some cells expressed myosin heavy chain, α-actinin and cardiac troponin T after 3 and 4 weeks. Flow cytometry confirmed these data. In cells exposed to trichostatin A and 5-aza-2-deoxycytidine and permeabilized in the presence of the cardiomyocyte extract, troponin T expression was seen in 3.53% of the cells and 3.11% of them expressed α-actinin. After exposure to the cardiomyocyte extract, some permeabilized cells adhered to the plate loosely; however, the morphology did not change significantly, and they continued to show a rounded shape after 4 weeks. Our treated lymphocytes expressed cardiomyocyte markers. Our results suggest that lymphocytes may be useful in future research as a source of cells for reprogramming procedures. PMID:21547694

  19. A secreted form of the human lymphocyte cell surface molecule CD8 arises from alternative splicing

    SciTech Connect

    Giblin, P.; Kavathas, P. ); Ledbetter, J.A. )

    1989-02-01

    The human lymphocyte differentiation antigen CD8 is encoded by a single gene that gives rise to a 33- to 34-kDa glycoprotein expressed on the cell surface as a dimer and in higher molecular mass forms. The authors demonstrate that the mRNA is alternatively spliced so that an exon encoding a transmembrane domain is deleted. This gives rise to a 30-kDa molecule that is secreted and exists primarily as a monomer. mRNA corresponding to both forms is present in peripheral blood lymphocytes, Con A-activated peripheral blood lymphocytes, and three CD8{sup +} T-cell lines, with the membrane form being the major species. However, differences in the ratio of mRNA for membrane CD8 and secreted CD8 exist. In addition, the splicing pattern observed differs from the pattern found for the mouse CD8 gene. This mRNA is also alternatively spliced, but an exon encoding a cytoplasmic region is deleted, giving rise to a cell surface molecule that differs in its cytoplasmic tail from the protein encoded by the longer mRNA. Neither protein is secreted. This is one of the first examples of a different splicing pattern between two homologous mouse and human genes giving rise to very different proteins. This represents one mechanism of generating diversity during speciation.

  20. Metabolic and cytoskeletal modulation of transferrin receptor mobility in mitogen-activated human lymphocytes.

    PubMed Central

    Galbraith, G M; Galbraith, R M

    1980-01-01

    The transferrin receptors which appear on mitogen-activated human peripheral blood lymphocytes were found by the use of immunofluorescence techniques to display temperature-dependent patching and capping reactions upon binding of transferrin. Lateral mobility of ligand-occupied membrane sites was accompanied by both shedding and endocytosis of receptor-transferrin complexes. In the presence of sodium azide or the microfilament inhibitor cytochalasin B, cap formation and shedding were markedly inhibited. In contrast, endocytosis of patched receptor-ligand complexes was inhibited by azide and microtubule inhibitors, including colchicine, vinblastine and vincristine. Co-capping experiments performed to elucidate further the alterations in membrane configuration involved in these reactions failed to reveal any topographical relationship between transferrin receptors and lectin-binding sites in these cells. These studied indicate that temperature-dependent mobility of transferrin receptors upon mitogen-activated peripheral blood lymphocytes is dependent upon the integrity of the cytoskeletal system and metabolic function of the cell. PMID:6258830

  1. Seasonal variations of DNA damage in human lymphocytes: correlation with different environmental variables.

    PubMed

    Giovannelli, Lisa; Pitozzi, Vanessa; Moretti, Silvia; Boddi, Vieri; Dolara, Piero

    2006-01-29

    Several types of DNA damage, including DNA breaks and DNA base oxidation, display a seasonal trend. In the present work, a sample of 79 healthy subjects living in the city of Florence, Italy, was used to analyse this effect. Three possible causative agents were taken into consideration: solar radiation, air temperature and air ozone level. DNA damage was measured in isolated human lymphocytes at different times during the year and the observed damage was correlated with the levels of these three agents in the days preceding blood sampling. Three time windows were chosen: 3, 7 and 30 days before blood sampling. DNA strand breaks and the oxidized purinic bases cleaved by the formamidopyrimidine glycosylase (FPG sites) were measured by means of the comet assay. The results of multivariate regression analysis showed a positive correlation between lymphocyte DNA damage and air temperature, and a less strong correlation with global solar radiation and air ozone levels. PMID:16095632

  2. Human EBV-transformed lymphocytes of patients with Schistosoma japonicum infection secrete idiotypically related immunoregulatory antibodies.

    PubMed

    Kresina, T F; Cheever, L W; Chireau, M; Johnson, J; Ramirez, B; Peters, P; Olds, G R

    1992-12-01

    Lymphocytes derived from the peripheral blood of individuals infected with Schistosoma japonica were transformed in vitro with Ebstein-Barr virus (EBV). Serological characterization of antibody molecules revealed both antigen reactive (idiotypic) and anti-idiotypic transformants. One idiotypic EBV transformant, LO2C2, describes a major cross-reactive idiotype associated with anti-antigen binding molecules. Other antibody populations expressing idiotypic cross-reactivity were derived from separate individuals showing shared idiotypy in S. japonicum field study populations in the Republic of Philippines. Both idiotypic and anti-idiotypic molecules suppressed parasite antigen-driven blastogenesis of heterologous human peripheral blood lymphocytes. The data show a serologically related immunoregulatory immune network in patients in the Republic of the Philippines which is serologically distinct from idiotypy expressed in other selected S. japonicum endemic areas in the Far East. PMID:1333380

  3. Assessment of genomic damage and repair on human lymphocytes by paint thinner in vitro.

    PubMed

    Londoño-Velasco, Elizabeth; Hidalgo-Cerón, Victor; Escobar-Hoyos, Luisa F; Hoyos-Giraldo, Luz Stella

    2014-05-01

    Paint thinners are organic-solvent complex mixtures frequently used by car painters around the world in industries and shops. Some studies have revealed the oxidative effect induced by thinner inhalation; however, its genotoxic effect is poorly studied. The aim of this study was to assess the cytotoxicity, genomic damage and DNA repair in vitro induced by commercial paint thinner 0.14 in human lymphocytes. Cytotoxicity was determined by cell-viability analysis with trypan blue after 4 h treatment with different thinner concentrations (0.025 to 1.2 µL/mL). Genomic damage was evaluated by means of the alkaline single-cell gel electrophoresis (SCGE; pH > 13) in treated cultures after 1 h with three low-cytotoxic thinner concentrations (0.05, 0.1 and 0.2 µL/mL). In order to evaluate the genomic DNA repair, one set of SCGE slides was prepared immediately after treatment, and another one was prepared after 4 h of liquid-holding recovery. A significant level of cytotoxicity was observed over the entire concentration range of paint thinner in lymphocytes (F = 175.98; p ≤ 0.001). In the SCGE % tail DNA assessment, a significant increase of lymphocyte genomic damage was evidenced (F = 72.32; p < 0.001). In addition, we found a significant decrease in the % tail DNA in thinner-treated cells after liquid-holding recovery period (all p < 0.05), demonstrating that DNA primary lesions induced by low-cytotoxic thinner concentrations are efficiently repaired. In conclusion, thinner components induce cytotoxicity and genomic damage in human lymphocytes under the study conditions, possibly by oxidative and alkylative DNA damage. PMID:24236478

  4. An ATP-activated channel is involved in mitogenic stimulation of human T lymphocytes.

    PubMed

    Baricordi, O R; Ferrari, D; Melchiorri, L; Chiozzi, P; Hanau, S; Chiari, E; Rubini, M; Di Virgilio, F

    1996-01-15

    We investigated the effect of pharmacologic modulation of the ATP receptor on intracellular ion changes and proliferative response of human peripheral blood lymphocytes (PBLs) and purified T lymphocytes. Extracellular ATP (ATPe) triggered in these cells an increase in the cytoplasmic Ca2+ concentration ([Ca2+]i) and plasma membrane depolarization. Whereas both Ca2+ release from intracellular stores and influx across the plasma membrane were detected in the whole PBL population, only Ca2+ influx was observed in T cells. In the presence of near physiologic extracellular Na+ concentrations (125 mmol/L), Ca2+ permeability through the ATPe-gated channel was very low, suggesting a higher selectivity for monovalent over divalent cations. The selective P2Z agonist benzoylbenzoic ATP (BzATP) increased [Ca2+]i in the presence but not the absence of extracellular Ca2+ and also caused plasma membrane depolarization. The covalent blocker oxidized ATP (oATP), an inhibitor of P2X and P2Z receptors, prevented Ca2+ influx and plasma membrane depolarization, but had no effect on Ca2+ release from stores. Stimulation with ATPe alone had no significant effects on PBL 3H-thymidine incorporation. On the contrary, ATPe or BzATP had a synergistic effect on DNA synthesis stimulated by selective T-cell mitogens such as phytohemagglutinin, anti-CD3 monoclonal antibody, or allogenic PBLs (mixed lymphocyte cultures). Treatment with oATP inhibited mitogenic stimulation by these receptor-directed agents but not by the combined application of the Ca2+ ionophore ionomycin and phorbol myristate acetate. Interleukin-2 partially relieved inhibition by oATP. These results suggest that human T lymphocytes express a plasma membrane channel gated by ATPe that is involved in mitogenic stimulation. PMID:8555491

  5. Lymphocyte trafficking and HIV infection of human lymphoid tissue in a rotating wall vessel bioreactor

    NASA Technical Reports Server (NTRS)

    Margolis, L. B.; Fitzgerald, W.; Glushakova, S.; Hatfill, S.; Amichay, N.; Baibakov, B.; Zimmerberg, J.

    1997-01-01

    The pathogenesis of HIV infection involves a complex interplay between both the infected and noninfected cells of human lymphoid tissue, the release of free viral particles, the de novo infection of cells, and the recirculatory trafficking of peripheral blood lymphocytes. To develop an in vitro model for studying these various aspects of HIV pathogenesis we have utilized blocks of surgically excised human tonsils and a rotating wall vessel (RWV) cell culture system. Here we show that (1) fragments of the surgically excised human lymphoid tissue remain viable and retain their gross cytoarchitecture for at least 3 weeks when cultured in the RWV system; (2) such lymphoid tissue gradually shows a loss of both T and B cells to the surrounding growth medium; however, this cellular migration is reversible as demonstrated by repopulation of the tissue by labeled cells from the growth medium; (3) this cellular migration may be partially or completely inhibited by embedding the blocks of lymphoid tissue in either a collagen or agarose gel matrix; these embedded tissue blocks retain most of the basic elements of a normal lymphoid cytoarchitecture; and (4) both embedded and nonembedded RWV-cultured blocks of human lymphoid tissue are capable of productive infection by HIV-1 of at least three various strains of different tropism and phenotype, as shown by an increase in both p24 antigen levels and free virus in the culture medium, and by the demonstration of HIV-1 RNA-positive cells inside the tissue identified by in situ hybridization. It is therefore reasonable to suggest that gel-embedded and nonembedded blocks of human lymphoid tissue, cocultured with a suspension of tonsillar lymphocytes in an RWV culture system, constitute a useful model for simulating normal lymphocyte recirculatory traffic and provide a new tool for testing the various aspects of HIV pathogenesis.

  6. Fc Receptor-like 5 Expression Distinguishes Two Distinct Subsets of Human Circulating Tissue-like Memory B Cells.

    PubMed

    Li, Huifang; Borrego, Francisco; Nagata, Satoshi; Tolnay, Mate

    2016-05-15

    Fc receptor-like (FCRL) 5 is a novel IgG binding protein expressed on B cells, with the capacity to regulate Ag receptor signaling. We assessed FCRL5 expression on circulating B cells from healthy donors and found that FCRL5(+) cells are most enriched among atypical CD21(-/lo)/CD27(-) tissue-like memory (TLM) B cells, which are abnormally expanded in several autoimmune and infectious diseases. Using multicolor flow cytometry, FCRL5(+) TLM cells were found to express more CD11c and several inhibitory receptors than did the FCRL5(-) TLM subset. The homing receptor profiles of the two TLM subsets shared features consistent with migration away from lymphoid tissues, but they also displayed distinct differences. Analysis of IgH V regions in single cells indicated that although both subsets are diverse, the FCRL5(+) subset accumulated significantly more somatic mutations. Furthermore, the FCRL5(+) subset had more switched isotype expression and more extensive proliferative history. Microarray analysis and quantitative RT-PCR demonstrated that the two TLM subsets possess distinct gene expression profiles, characterized by markedly different CD11c, SOX5, T-bet, and RTN4R expression, as well as differences in expression of inhibitory receptors. Functional analysis revealed that the FCRL5(+) TLM subset responds poorly to multiple stimuli compared with the FCRL5(-) subset, as reflected by reduced calcium mobilization and blunted cell proliferation. We propose that the FCRL5(+) TLM subset, but not the FCRL5(-) TLM subset, underwent Ag-driven development and is severely dysfunctional. The present study elucidates the heterogeneity of TLM B cells and provides the basis to dissect their roles in the pathogenesis of inflammatory and infectious diseases. PMID:27076679

  7. Human Lymphocytes

    NASA Technical Reports Server (NTRS)

    2004-01-01

    Biomedical research offers hope for a variety of medical problems, from diabetes to the replacement of damaged bone and tissues. Bioreactors, which are used to grow cells and tissue cultures, play a major role in such research and production efforts. The objective of the research was to define a way to differentiate between effects due to microgravity and those due to possible stress from non-optimal spaceflight conditions.

  8. Assessment of genotoxicity of Lannate-90® and its plant and animal metabolites in human lymphocyte cultures.

    PubMed

    Valencia-Quintana, Rafael; Gómez-Arroyo, Sandra; Sánchez-Alarcón, Juana; Milić, Mirta; Olivares, José Luis Gómez; Waliszewski, Stefan M; Cortés-Eslava, Josefina; Villalobos-Pietrini, Rafael; Calderón-Segura, María Elena

    2016-06-01

    This study evaluated direct and metabolic genotoxic effects caused by Lannate-90®, a methomyl-based formulation (90 % active ingredient), in human lymphocyte cultures using sister chromatid exchange assay (SCE). Two processes were used for the plant promutagens evaluation: in vivo activation, applying the insecticide systemically in plants for 4 h and subsequently adding plant metabolites containing extracts to lymphocyte cultures; and in vitro activation, where the insecticide was incubated with Vicia faba S10 mix plus human lymphocyte culture. Direct treatment with the insecticide significantly increased SCE frequency in human lymphocytes (250-750 mgL-1), with cellular death observed at 1000 mgL-1 concentration. Using the extracts of Vicia faba treated with Lannate-90® to treat human lymphocytes, a dose-response relationship was observed. In lymphocyte cultures treated directly with the insecticide for 2 h, a negative response was obtained. When S10 mix was added, SCE frequency did not change significantly. Meanwhile, a mixture of S9 mammalian metabolic mix and Lannate-90® increased the SCE frequency, with an observed concentration-dependent response. Although Lannate-90® induced cellular death at the highest concentrations, it did not cause a delay in cell proliferation in any of the treatments, confirming its genotoxic action. This study is one of the first to evaluate and compare the direct effect of Lannate-90® in two bioassays, animal and vegetal, and the effect of plant and animal metabolism on its genotoxic potential. PMID:27331299

  9. Differential expression of three T lymphocyte-activating CXC chemokines by human atheroma-associated cells

    PubMed Central

    Mach, François; Sauty, Alain; Iarossi, Albert S.; Sukhova, Galina K.; Neote, Kuldeep; Libby, Peter; Luster, Andrew D.

    1999-01-01

    Activated T lymphocytes accumulate early in atheroma formation and persist at sites of lesion growth and rupture, suggesting that they may play an important role in the pathogenesis of atherosclerosis. Moreover, atherosclerotic lesions contain the Th1-type cytokine IFN-γ, a potentiator of atherosclerosis. The present study demonstrates the differential expression of the 3 IFN-γ–inducible CXC chemokines — IFN-inducible protein 10 (IP-10), monokine induced by IFN-γ (Mig), and IFN-inducible T-cell α chemoattractant (I-TAC) — by atheroma-associated cells, as well as the expression of their receptor, CXCR3, by all T lymphocytes within human atherosclerotic lesions in situ. Atheroma-associated endothelial cells (ECs), smooth muscle cells (SMCs), and macrophages (MØ) all expressed IP-10, whereas Mig and I-TAC were mainly expressed in ECs and MØ, as detected by double immunofluorescence staining. ECs of microvessels within lesions also expressed abundant I-TAC. In vitro experiments supported these results and showed that IL-1β, TNF-α, and CD40 ligand potentiated IP-10 expression from IFN-γ–stimulated ECs. In addition, nitric oxide (NO) treatment decreased IFN-γ induction of IP-10. Our findings suggest that the differential expression of IP-10, Mig, and I-TAC by atheroma-associated cells plays a role in the recruitment and retention of activated T lymphocytes observed within vascular wall lesions during atherogenesis. PMID:10525042

  10. Visualization of antigen-specific human cytotoxic T lymphocytes labeled with superparamagnetic iron-oxide particles.

    PubMed

    Beer, Ambros J; Holzapfel, Konstantin; Neudorfer, Juliana; Piontek, Guido; Settles, Marcus; Krönig, Holger; Peschel, Christian; Schlegel, Jürgen; Rummeny, Ernst J; Bernhard, Helga

    2008-06-01

    New technologies are needed to characterize the migration and survival of antigen-specific T cells in vivo. In this study, we developed a novel technique for the labeling of human cytotoxic T lymphocytes with superparamagnetic iron-oxide particles and the subsequent depiction with a conventional 1.5-T magnetic resonance scanner. Antigen-specific CD8(+) T lymphocytes were labeled with ferucarbotran by lipofection. The uptake of ferucarbotran was confirmed by immunofluorescence microscopy using a dextran-specific antibody, and the intracellular enrichment of iron was measured by atomic absorption spectrometry. The imaging of T cells was performed by magnetic resonance on day 0, 2, 7 and 14 after the labeling procedure. On day 0 and 2 post labeling, a pronounced shortening of T2*-relaxation times was observed, which diminished after 7 days and was not detectable anymore after 14 days, probably due to the retained mitotic activity of the labeled T cells. Of importance, the antigen-specific cytolytic activity of the T cells was preserved following ferucarbotran labeling. Efficient ferucarbotran labeling of functionally active T lymphocytes and their detection by magnetic resonance imaging allows the in vivo monitoring of T cells and, subsequently, will impact the further development of T cell-based therapies. PMID:18286290

  11. Early and Late Damages in Chromosome 3 of Human Lymphocytes After Radiation Exposure

    NASA Technical Reports Server (NTRS)

    Sunagawa, Mayumi; Mangala, Lingegowda; Zhang, Ye; Kahdim, Munira; Wilson, Bobby; Cucinotta, Francis A.; Wu, Honglu

    2011-01-01

    Tumor formation in humans or animals is a multi-step process. An early stage of cancer development is believed to be genomic instability (GI) which accelerates the mutation rate in the descendants of the cells surviving radiation exposure. GI is defined as elevated or persistent genetic damages occurring many generations after the cells are exposed. While early studies have demonstrated radiation-induced GI in several cell types as detected in endpoints such as mutation, apoptosis and damages in chromosomes, the dependence of GI on the quality of radiation remains uncertain. To investigate GI in human lymphocytes induced by both low- and high-LET radiation, we initially exposed white blood cells collected from healthy subjects to gamma rays in vitro, and cultured the cells for multiple generations. Chromosome aberrations were analyzed in cells collected at first mitosis post irradiation and at several intervals during the culture period. Among a number of biological endpoints planned for the project, the multi-color banding fluorescent in situ hybridization (mBAND) allows identification of inversions that were expected to be stable. We present here early and late chromosome aberrations detected with mBAND in chromosome 3 after gamma exposure. Comparison of chromosome damages in between human lymphocytes and human epithelial cells is also discussed

  12. Monocyte chemotactic protein-1 (MCP-1), -2, and -3 are chemotactic for human T lymphocytes.

    PubMed Central

    Taub, D D; Proost, P; Murphy, W J; Anver, M; Longo, D L; van Damme, J; Oppenheim, J J

    1995-01-01

    Monocyte chemotactic protein (MCP)-1, -2, and -3 all have been shown to induce monocyte/macrophage migration in vitro and MCP-1, also known as MCAF, chemoattracts basophils and mast cells. We report here that natural MCP-1 as well as synthetic preparations of MCP-2 and MCP-3 stimulate significant in vitro chemotaxis of human peripheral blood T lymphocytes. This MCP-induced migration was dose-dependent and directional, but not chemokinetic. Phenotypic analysis of the T cell population responsive to MCP-1, MCP-2, and MCP-3 demonstrates that both CD4+ and CD8+ T cells migrated in response to these chemokines. Similar results were observed using human CD4+ and CD8+ T cell clones. Neutralizing antisera to MCAF or MCP-2 abrogated T cell migration in response to MCP-1 and MCP-2, respectively, but not to RANTES. Subcutaneous administration of purified MCP-1 into the hind flanks of SCID mice engrafted with human peripheral blood lymphocytes (PBL) induced significant human CD3+ T cell infiltration into the site of injection at 4 h. These results demonstrate that MCP-1, MCP-2, and MCP-3 are inflammatory mediators that specifically stimulate the directional migration of T cells as well as monocytes and may play an important role in immune cell recruitment into sites of antigenic challenge. Images PMID:7883984

  13. The pyrimidin analogue cyclopentenyl cytosine induces alloantigen-specific non-responsiveness of human T lymphocytes

    PubMed Central

    Nikolaeva, N; Bemelman, F J; Yong, S-L; Verschuur, A; van Lier, R A W; ten Berge, I J M

    2008-01-01

    Cyclopentenyl cytosine (CPEC) has been shown to induce apoptosis in human T lymphoblastic cell lines and T cells from leukaemia patients. In this study we have addressed the question of whether CPEC is able to decrease proliferation and effector functions of human alloresponsive T lymphocytes and induce T cell anergy. The proliferative capacity of human peripheral blood mononuclear cells in response to allogeneic stimulation was measured by 5,6-carboxy-succinimidyl-diacetate-fluorescein-ester staining. Flow cytometric analysis was performed using surface CD4, CD8, CD25, CD103 and intracellular perforin, granzyme A, granzyme B, caspase-3 and forkhead box P3 (FoxP3) markers. The in vivo immunosuppressive capacity was tested in a murine skin graft model. Addition of CPEC at a concentration of 20 nM strongly decreased the expansion and cytotoxicity of alloreactive T cells. Specific restimulation in the absence of CPEC showed that the cells became anergic. The drug induced caspase-dependent apoptosis of alloreactive T lymphocytes. Finally, CPEC increased the percentage of CD25high FoxP3+ CD4+ and CD103+ CD8+ T cells, and potentiated the effect of rapamycin in increasing the numbers of alloreactive regulatory T cells. Treatment with CPEC of CBA/CA mice transplanted with B10/Br skin grafts significantly prolonged graft survival. We conclude that CPEC inhibits proliferation and cytotoxicity of human alloreactive T cells and induces alloantigen non-responsiveness in vitro. PMID:18062797

  14. Impact of Varicella-Zoster Virus on Dendritic Cell Subsets in Human Skin during Natural Infection▿ †

    PubMed Central

    Huch, Jennifer H.; Cunningham, Anthony L.; Arvin, Ann M.; Nasr, Najla; Santegoets, Saskia J. A. M.; Slobedman, Eric; Slobedman, Barry; Abendroth, Allison

    2010-01-01

    Varicella-zoster virus (VZV) causes varicella and herpes zoster, diseases characterized by distinct cutaneous rashes. Dendritic cells (DC) are essential for inducing antiviral immune responses; however, the contribution of DC subsets to immune control during natural cutaneous VZV infection has not been investigated. Immunostaining showed that compared to normal skin, the proportion of cells expressing DC-SIGN (a dermal DC marker) or DC-LAMP and CD83 (mature DC markers) were not significantly altered in infected skin. In contrast, the frequency of Langerhans cells was significantly decreased in VZV-infected skin, whereas there was an influx of plasmacytoid DC, a potent secretor of type I interferon (IFN). Langerhans cells and plasmacytoid DC in infected skin were closely associated with VZV antigen-positive cells, and some Langerhans cells and plasmacytoid DC were VZV antigen positive. To extend these in vivo observations, both plasmacytoid DC (PDC) isolated from human blood and Langerhans cells derived from MUTZ-3 cells were shown to be permissive to VZV infection. In VZV-infected PDC cultures, significant induction of alpha IFN (IFN-α) did not occur, indicating the VZV inhibits the capacity of PDC to induce expression of this host defense cytokine. This study defines changes in the response of DC which occur during cutaneous VZV infection and implicates infection of DC subtypes in VZV pathogenesis. PMID:20130046

  15. Serine leucocyte proteinase inhibitor-treated monocyte inhibits human CD4(+) lymphocyte proliferation.

    PubMed

    Guerrieri, Diego; Tateosian, Nancy L; Maffía, Paulo C; Reiteri, Romina M; Amiano, Nicolás O; Costa, María J; Villalonga, Ximena; Sanchez, Mercedes L; Estein, Silvia M; Garcia, Verónica E; Sallenave, Jean-Michel; Chuluyan, Héctor E

    2011-08-01

    Serine leucocyte proteinase inhibitor (SLPI) is the main serine proteinase inhibitor produced by epithelial cells and has been shown to be a pleiotropic molecule with anti-inflammatory and microbicidal activities. However, the role of SLPI on the adaptive immune response is not well established. Therefore, we evaluated the effect of SLPI on lymphocyte proliferation and cytokine production. Human peripheral blood mononuclear cells (PBMC) were treated with mitogens plus SLPI and proliferation was assessed by [(3) H]thymidine uptake. The SLPI decreased the lymphocyte proliferation induced by interleukin-2 (IL-2) or OKT3 monoclonal antibodies in a dose-dependent manner. Inhibition was not observed when depleting monocytes from the PBMC and it was restored by adding monocytes and SLPI. SLPI-treated monocyte slightly decreased MHC II and increased CD18 expression, and secreted greater amounts of IL-4, IL-6 and IL-10 in the cell culture supernatants. SLPI-treated monocyte culture supernatant inhibited the CD4(+) lymphocyte proliferation but did not affect the proliferation of CD8(+) cells. Moreover, IL-2 increased T-bet expression and the presence of SLPI significantly decreased it. Finally, SLPI-treated monocyte culture supernatant dramatically decreased interferon-γ but increased IL-4, IL-6 and IL-10 in the presence of IL-2-treated T cells. Our results demonstrate that SLPI target monocytes, which in turn inhibit CD4 lymphocyte proliferation and T helper type 1 cytokine secretion. Overall, these results suggest that SLPI is an alarm protein that modulates not only the innate immune response but also the adaptive immune response. PMID:21574992

  16. Serine leucocyte proteinase inhibitor-treated monocyte inhibits human CD4+ lymphocyte proliferation

    PubMed Central

    Guerrieri, Diego; Tateosian, Nancy L; Maffía, Paulo C; Reiteri, Romina M; Amiano, Nicolás O; Costa, María J; Villalonga, Ximena; Sanchez, Mercedes L; Estein, Silvia M; Garcia, Verónica E; Sallenave, Jean-Michel; Chuluyan, Héctor E

    2011-01-01

    Serine leucocyte proteinase inhibitor (SLPI) is the main serine proteinase inhibitor produced by epithelial cells and has been shown to be a pleiotropic molecule with anti-inflammatory and microbicidal activities. However, the role of SLPI on the adaptive immune response is not well established. Therefore, we evaluated the effect of SLPI on lymphocyte proliferation and cytokine production. Human peripheral blood mononuclear cells (PBMC) were treated with mitogens plus SLPI and proliferation was assessed by [3H]thymidine uptake. The SLPI decreased the lymphocyte proliferation induced by interleukin-2 (IL-2) or OKT3 monoclonal antibodies in a dose-dependent manner. Inhibition was not observed when depleting monocytes from the PBMC and it was restored by adding monocytes and SLPI. SLPI-treated monocyte slightly decreased MHC II and increased CD18 expression, and secreted greater amounts of IL-4, IL-6 and IL-10 in the cell culture supernatants. SLPI-treated monocyte culture supernatant inhibited the CD4+ lymphocyte proliferation but did not affect the proliferation of CD8+ cells. Moreover, IL-2 increased T-bet expression and the presence of SLPI significantly decreased it. Finally, SLPI-treated monocyte culture supernatant dramatically decreased interferon-γ but increased IL-4, IL-6 and IL-10 in the presence of IL-2-treated T cells. Our results demonstrate that SLPI target monocytes, which in turn inhibit CD4 lymphocyte proliferation and T helper type 1 cytokine secretion. Overall, these results suggest that SLPI is an alarm protein that modulates not only the innate immune response but also the adaptive immune response. PMID:21574992

  17. Impact of different concentrations of human recombinant growth hormone on T lymphocytes.

    PubMed

    Borrione, P; Grasso, L; Pautasso, M; Parisi, A; Quaranta, F; Ciminelli, E; Di Gianfrancesco, A; Di Luigi, L; Pigozzi, F

    2012-01-01

    The aim of the present study is to evaluate the effects induced by increasing concentrations of human recombinant growth hormone on T lymphocytes. Ten healthy volunteers and twelve subjects with symptomatic allergies were enrolled in the study. Peripheral blood mononuclear cells and purified T lymphocytes were cultured in the presence of graded concentrations of growth hormone. Following appropriate in vitro stimulations, the proportion of apoptotic T cells, the percentage of activated T lymphocyte subpopulations, the phytohemagglutinin responsiveness and the Th2 response were assessed by flow cytometry analysis. Moreover, in order to evaluate the phosphoinositol-3-kinase signaling pathway involvement, cells were also analyzed after treatment with LY294002. The treatment with different concentrations of growth hormone did not influence the activation pattern of un-stimulated T lymphocytes. On the contrary, growth hormone was able to modify the CD38/HLA-DR co-expression of T cells activated with phytohemoagglutinin. A different response was observed when samples obtained from healthy donors and from subjects with symptomatic allergies were analysed. Moreover, growth hormone treatment was able to increase the Th2 response in the samples obtained from healthy donors only. The results of the present study strongly support the hypothesis that growth hormone administration may play an important role in conditions of impaired/activated immune systems. The observation that growth hormone administration at high doses may reverse its effects and that it may promote a Th2-oriented response have significant clinical implications when considering the use of this hormone for artificially enhancing the physical performances of healthy athletes. PMID:22507321

  18. Impact of the New Generation Reconstituted Surfactant CHF5633 on Human CD4+ Lymphocytes

    PubMed Central

    Fehrholz, Markus; Glaser, Kirsten; Seidenspinner, Silvia; Ottensmeier, Barbara; Curstedt, Tore; Speer, Christian P.; Kunzmann, Steffen

    2016-01-01

    Background Natural surfactant preparations, commonly isolated from porcine or bovine lungs, are used to treat respiratory distress syndrome in preterm infants. Besides biophysical effectiveness, several studies have documented additional immunomodulatory properties. Within the near future, synthetic surfactant preparations may be a promising alternative. CHF5633 is a new generation reconstituted synthetic surfactant preparation with defined composition, containing dipalmitoyl-phosphatidylcholine, palmitoyl-oleoyl-phosphatidylglycerol and synthetic analogs of surfactant protein (SP-) B and SP-C. While its biophysical effectiveness has been demonstrated in vitro and in vivo, possible immunomodulatory abilities are currently unknown. Aim The aim of the current study was to define a potential impact of CHF5633 and its single components on pro- and anti-inflammatory cytokine responses in human CD4+ lymphocytes. Methods Purified human CD4+ T cells were activated using anti CD3/CD28 antibodies and exposed to CHF5633, its components, or to the well-known animal-derived surfactant Poractant alfa (Curosurf®). Proliferative response and cell viability were assessed using flow cytometry and a methylthiazolyldiphenyltetrazolium bromide colorimetric assay. The mRNA expression of IFNγ, IL-2, IL-17A, IL-22, IL-4, and IL-10 was measured by quantitative PCR, while intracellular protein expression was assessed by means of flow cytometry. Results Neither CHF5633 nor any of its phospholipid components with or without SP-B or SP-C analogs had any influence on proliferative ability and viability of CD4+ lymphocytes under the given conditions. IFNγ, IL-2, IL-17A, IL-22, IL-4, and IL-10 mRNA as well as IFNγ, IL-2, IL-4 and IL-10 protein levels were unaffected in both non-activated and activated CD4+ lymphocytes after exposure to CHF5633 or its constituents compared to non-exposed controls. However, in comparison to Curosurf®, expression levels of anti-inflammatory IL-4 and IL-10 m

  19. Adenovirus-specific T-cell Subsets in Human Peripheral Blood and After IFN-γ Immunomagnetic Selection.

    PubMed

    Qian, Chongsheng; Wang, Yingying; Cai, Huili; Laroye, Caroline; De Carvalho Bittencourt, Marcelo; Clement, Laurence; Stoltz, Jean-François; Decot, Véronique; Reppel, Loïc; Bensoussan, Danièle

    2016-01-01

    Adoptive antiviral cellular immunotherapy by infusion of virus-specific T cells (VSTs) is becoming an alternative treatment for viral infection after hematopoietic stem cell transplantation. The T memory stem cell (TSCM) subset was recently described as exhibiting self-renewal and multipotency properties which are required for sustained efficacy in vivo. We wondered if such a crucial subset for immunotherapy was present in VSTs. We identified, by flow cytometry, TSCM in adenovirus (ADV)-specific interferon (IFN)-γ+ T cells before and after IFN-γ-based immunomagnetic selection, and analyzed the distribution of the main T-cell subsets in VSTs: naive T cells (TN), TSCM, T central memory cells (TCM), T effector memory cell (TEM), and effector T cells (TEFF). In this study all of the different T-cell subsets were observed in the blood sample from healthy donor ADV-VSTs, both before and after IFN-γ-based immunomagnetic selection. As the IFN-γ-based immunomagnetic selection system sorts mainly the most differentiated T-cell subsets, we observed that TEM was always the major T-cell subset of ADV-specific T cells after immunomagnetic isolation and especially after expansion in vitro. Comparing T-cell subpopulation profiles before and after in vitro expansion, we observed that in vitro cell culture with interleukin-2 resulted in a significant expansion of TN-like, TCM, TEM, and TEFF subsets in CD4IFN-γ T cells and of TCM and TEM subsets only in CD8IFN-γ T cells. We demonstrated the presence of all T-cell subsets in IFN-γ VSTs including the TSCM subpopulation, although this was weakly selected by the IFN-γ-based immunomagnetic selection system. PMID:26641259

  20. Dopamine Receptors in Human Lymphocytes: Radioligand Binding and Quantitative RT-PCR Assays

    PubMed Central

    Kirillova, Galina P.; Hrutkay, Rebecca J.; Shurin, Michael R.; Shurin, Galina V.; Tourkova, Irina L.; Vanyukov, Michael M.

    2008-01-01

    Analysis of dopamine receptors (DR) in lymphocytes of the human peripheral blood mononuclear cell (PBMC) fraction is an attractive tool for evaluation of functional properties of dopaminergic function underlying variation in complex psychological/psychopathological traits. Receptor binding assays (RBA) with selective radioligands, which are widely used in CNS studies, have not produced consistent results when applied to isolated PBMC. We tested the assay conditions that could be essential for detection of DR in human PBMC and their membrane preparations. Using [3H]SCH23390, a dopamine D1-like receptor antagonist, we demonstrated the presence of two binding sites in PBMC-derived membrane fraction. One of them is characterized by the Kd value consistent with that reported for D5 dopamine receptors in human lymphocytes, whereas the other Kd value possibly corresponds to serotonin receptor(s). Although D5 receptor binding sites in PBMC membranes could be characterized by binding assays, the low protein expression and the large volume of blood needed for membrane preparation render the binding method impracticable for individual phenotyping. In contrast, real-time RT-PCR may be used for this purpose, contingent on the relationship between DR expression in the brain and in lymphocytes. The expression of the DRD2-DRD5 genes, as detected by this method, varied widely among samples, whereas the DRD1 expression was not detected. The expression levels were comparable with those in the brain for DRD3 and DRD4, and were significantly lower for DRD2 and DRD5. PMID:18721826

  1. Effects of increasing docosahexaenoic acid intake in human healthy volunteers on lymphocyte activation and monocyte apoptosis

    PubMed Central

    Mebarek, Saïda; Ermak, Natalia; Benzaria, Amal; Vicca, Stéphanie; Dubois, Madeleine; Némoz, Georges; Laville, Martine; Lacour, Bernard; Véricel, Evelyne; Lagarde, Michel; Prigent, Annie-France

    2009-01-01

    Dietary intake of long-chain n-3 polyunsaturated fatty acids (n-3 PUFA) has been reported to decrease several markers of lymphocyte activation and modulate monocyte susceptibility to apoptosis. However most human studies examined the combined effect of docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA) using relatively high daily amounts of n-3 PUFA. The present study investigated the effects of increasing doses of DHA added to the regular diet of human healthy volunteers on lymphocyte response to tetradecanoylphorbol acetate (TPA) plus ionomycin activation, and on monocyte apoptosis induced by oxidized LDL (oxLDL). Eight subjects were supplemented with increasing daily doses of DHA (200, 400, 800 and 1600mg) in a triacylglycerol form containing DHA as the only PUFA, for two weeks each dose. DHA intake dose-dependently increased the proportion of DHA in mononuclear cell phospholipids, the augmentation being significant after 400mg DHA/day. The TPA plus ionomycin-stimulated IL-2 mRNA level started to increase after ingestion of 400mg DHA/day, with a maximum after 800mg intake, and was positively correlated (P<0.003) with DHA enrichment in cell phospholipids. The treatment of monocytes by oxLDL before DHA supplementation drastically reduced mitochondrial membrane potential as compared with native LDL treatment. OxLDL apoptotic effect was significantly attenuated after 400mg DHA/day and the protective effect was maintained throughout the experiment, although to a lesser extent at higher doses. The present results show that supplementation of the human diet with low DHA dosages improves lymphocyte activability. It also increases monocyte resistance to oxLDL-induced apoptosis, which may be beneficial in the prevention of atherosclerosis. PMID:18710607

  2. Cytofluorimetric identification of two populations of double positive (CD4+,CD8+) T lymphocytes in human peripheral blood.

    PubMed

    Ortolani, C; Forti, E; Radin, E; Cibin, R; Cossarizza, A

    1993-03-15

    Two different subsets of CD4+,CD8+ T lymphocytes have been identified in peripheral blood collected from normal subjects and from patients with different diseases. The subpopulations differed in the degree of CD4 and CD8 antigen expression. Hence, it was possible to distinguish by cytofluorimetric analysis cells with a low (dim) or with a high (bright) fluorescence intensity after the staining with anti-CD4 or anti-CD8 mAbs. CD4+dim,CD8+bright lymphocytes were found in patients with EBV-infectious mononucleosis and were present for less than a month. CD4+bright,CD8+dim T cells were observed in neoplastic patients as well as in healthy subjects and were continuously present in similar percentages over a long period of time (at the moment, about 3 years). Both the subpopulations expressed CD2, CD3, CD5 antigens and had an alpha beta-TCR, but did not express CD1a or CD7. Only CD4+dim,CD8+bright cells expressed HLA-DR antigen and the activation marker CD38, while only CD4+bright,CD8+dim lymphocytes expressed CD56 and CD57 molecules. The hypothesis may be put forward that these two subsets represent an effort of the immune system to cope with different requirements, i.e., of viral or neoplastic origin, while it is not clear the meaning of these cells in healthy subjects. PMID:8461016

  3. Brucella fractions behave as nonspecific mitogens and polyclonal B-cell activators for human lymphocytes.

    PubMed Central

    Vendrell, J P; Rabesandratana, H; Huguet, M F; Cannat, A; Serre, A

    1985-01-01

    Two lipid-A-free fractions which were extracted from Brucella melitensis and were designated PI and SF stimulated human unsensitized mononuclear cells to proliferate and to secrete immunoglobulins. Both of these effects were observed in cultures of peripheral blood, tonsils, and cord blood lymphocytes. Neither B cells nor T cells alone proliferated in the presence of these fractions, whereas the proliferative response of T cells plus B cells was largely independent of accessory cells. Polyclonal activation was estimated by counting the cells which secreted immunoglobulins of different isotypes into culture supernatants. This phenomenon was strongly T dependent. PMID:3876286

  4. The effects of lipid A on gamma-irradiated human peripheral blood lymphocytes in vitro

    NASA Astrophysics Data System (ADS)

    Dubničková, M.; Kuzmina, E. A.; Chausov, V. N.; Ravnachka, I.; Boreyko, A. V.; Krasavin, E. A.

    2016-03-01

    The modulatory effects of lipid A (diphosphoryl lipid A (DLA) and monophosphoryl lipid A (MLA)) on apoptosis induction and DNA structure damage (single and double-strand breaks (SSBs and DSBs, respectively)) in peripheral human blood lymphocytes are studied for 60Co gamma-irradiation. It is shown that in the presence of these agents the amount of apoptotic cells increases compared with the irradiated control samples. The effect is most strongly pronounced for DLA. In its presence, a significant increase is observed in the number of radiation-induced DNA SSBs and DSBs. Possible mechanisms are discussed of the modifying influence of the used agents on radiation-induced cell reactions

  5. Suppression of human lymphocyte responses to specific and non-specific stimuli in human onchocerciasis.

    PubMed Central

    Elkhalifa, M Y; Ghalib, H W; Dafa'Alla, T; Williams, J F

    1991-01-01

    Characterization of in vitro lymphocyte responsiveness was performed on selected groups of onchocerciasis patients from Sudan and Sierra Leone. These patients manifested a very broad range of clinical signs and showed widely divergent parasite infection intensities. Lymphocyte proliferative responses to soluble Onchocerca volvulus antigen (sAg) were poor in infected persons; mitogen and PPD responses were maintained in the normal range in one group of patients from southwestern Sudan, but were profoundly depressed in a group from N.E. Sudan. Proliferative responses and interferon-gamma (INF-gamma) secretion were very significantly depressed in the presence of live microfilariae of O. volvulus or secretions/excretions (S/E) from microfilariae (mf) or from female, but not male, adult parasites. Lymphocyte responses were maintained near normal when exogenous IL-2 was added to these cultures. The results indicate that O. volvulus infection and its clinical consequences are not consistently associated with systemic deficits in immune responsiveness. However, suppression of lymphocyte reactivity by mf and S/E in vitro suggests that direct parasite intervention in host cell responses could be taking place in vivo, perhaps at the local microenvironment level; mediated by effects on cytokine production. PMID:1747951

  6. Erythropoietin exerts direct immunomodulatory effects on the cytokine production by activated human T-lymphocytes.

    PubMed

    Todosenko, N M; Shmarov, V A; Malashchenko, V V; Meniailo, M E; Melashchenko, O B; Gazatova, N D; Goncharov, A G; Seledtsov, V I

    2016-07-01

    The effect of erythropoietin-β (Epo-β) on the functional profile of activated human T-lymphocytes remains largely unknown, which hinders clinical application of Epo as an immunomodulatory agent. We studied the direct impact of Epo on the activation status of human T lymphocytes following activation by particles loaded with antibodies (Abs) against human CD2, CD3, and CD28. T cell activation was assessed by the surface expression of CD38 activation marker. Epo did not significantly affect activation status of both CD4(+) and CD4(-) T cells, as well as of naive (CD45RA(+)CD197(+)), central memory (CD45RA(-)CD197(+)), effector memory (CD45RA(-)CD197(-)), and terminally-differentiated (CD45RA(+)CD197(-)) T cells. However, Epo markedly augmented production of IL-2, IL-4 and IL10 by activated T cells with concomitant reduction in IFN-γ secretion. Taken together, our data showed that Epo could directly down-regulate pro-inflammatory T cell responses without affecting T cell activation status. PMID:27208431

  7. Characterization of effector and memory T cell subsets in the immune response to bovine tuberculosis in cattle

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Vaccine-elicited long-term cultured IFN-gamma ELISPOT responses correlate with protection against bovine tuberculosis in cattle. With humans, cultured IFN-gamma ELISPOT assays are primarily a measure of central memory T cell (Tcm) responses; however, this important subset of lymphocytes is poorly ch...

  8. Interaction of Epstein-Barr virus (EBV) with human B-lymphocytes

    SciTech Connect

    Klein, George; Klein, Eva; Kashuba, Elena

    2010-05-21

    Epstein-Barr virus, EBV, and humans have a common history that reaches back to our primate ancestors. The virus co-evolved with man and has established a largely harmless and highly complex co-existence. It is carried as silent infection by almost all human adults. A serendipitous discovery established that it is the causative agent of infectious mononucleosis. Still, EBV became known first in 1964, in a rare, geographically prevalent malignant lymphoma of B-cell origin, Burkitt lymphoma BL. Its association with a malignancy prompted intensive studies and its capacity to immortalize B-lymphocytes in vitro was soon demonstrated. Consequently EBV was classified therefore as a potentially tumorigenic virus. Despite of this property however, the virus carrier state itself does not lead to malignancies because the transformed cells are recognized by the immune response. Consequently the EBV induced proliferation of EBV carrying B-lymphocytes is manifested only under immunosuppressive conditions. The expression of EBV encoded genes is regulated by the cell phenotype. The virus genome can be found in malignancies originating from cell types other than the B-lymphocyte. Even in the EBV infected B-cell, the direct transforming capacity is restricted to a defined window of differentiation. A complex interaction between virally encoded proteins and B-cell specific cellular proteins constitute the proliferation inducing program. In this short review we touch upon aspects which are the subject of our present work. We describe the mechanisms of some of the functional interactions between EBV encoded and cellular proteins that determine the phenotype of latently infected B-cells. The growth promoting EBV encoded genes are not expressed in the virus carrying BL cells. Still, EBV seems to contribute to the etiology of this tumor by modifying events that influence cell survival and proliferation. We describe a possible growth promoting mechanism in the genesis of Burkitt lymphoma

  9. Protective effect of apigenin on radiation-induced chromosomal damage in human lymphocytes

    NASA Technical Reports Server (NTRS)

    Rithidech, Kanokporn Noy; Tungjai, Montree; Whorton, Elbert B.

    2005-01-01

    The potential use of flavonoids as a radioprotector is of increasing interest because of their high antioxidant activity and abundance in the diet. The aim of this study is to examine genotoxic and radioprotective effects of one of the most common flavonoids, apigenin, on radiation-induced chromosome aberrations in human lymphocytes. The cytokinesis-block micronucleus (CBMN) assay was used to evaluate such effects of apigenin. Blood samples were collected from two non-smoking healthy male volunteers who had no history of previous exposure to other clastogenic agents. Isolated lymphocytes were cultured. There were two tubes per concentration for all treatments. To evaluate the genotoxicity of apigenin, cells were first treated with different concentrations of apigenin (0, 2.5, 5, 10 and 25 microg/mL) at 24 h after culture initiation, followed by cytochalasin-B (Cyt-B) treatment (3 microg/mL) and cell harvest at 44 and 72 h, respectively. Secondly, to investigate the radioprotective effect, cell cultures were exposed to different concentrations of apigenin as described above for 30 min before being irradiated to 2 Gy of 137Cs gamma rays (at a dose rate of 0.75 Gy/min). In all instances, the frequency of MN was scored in binucleated (BN) cells. The nuclear proliferation index also was calculated. We did not detect an increase in the frequency of MN in non-irradiated human lymphocyte cultures treated with 2.5, 5.0 or 10 microg/mL apigenin; although, we did observe an increase in cultures treated with 25 microg/mL apigenin (the highest concentration of apigenin used in our study). We also observed a significant increase in the frequency of MN in irradiated cells overall; however, the frequency was decreased as the concentration of apigenin increased, suggesting a radioprotective effect. These findings provide a basis for additional studies to help clarify the potential use and benefit of apigenin as a radioprotector.

  10. Activated human B lymphocytes express cyclooxygenase-2 and cyclooxygenase inhibitors attenuate antibody production.

    PubMed

    Ryan, Elizabeth P; Pollock, Stephen J; Pollack, Stephen J; Murant, Thomas I; Bernstein, Steven H; Felgar, Raymond E; Phipps, Richard P

    2005-03-01

    Nonsteroidal anti-inflammatory drugs (NSAIDs) are widely used for the treatment of inflammatory diseases and target cyclooxygenases 1 and 2 (Cox-1, Cox-2) that are responsible for PG production. Newer Cox-2-selective drugs have been heavily prescribed to quench inflammation. Little is known about whether or not these drugs influence human B lymphocytes and their ability to produce Ab. We report herein that activated human B cells not only highly express Cox-2 and produce PGs, but that the NSAID indomethacin and Cox-2-selective drugs profoundly inhibit the ability of human B cells to produce IgG and IgM in vitro. Human blood B cells highly express Cox-2 mRNA and protein and produce PGs after activation with CD40L, pansorbin, or CD40L plus BCR engagement. Cox-2 is also highly expressed by human tonsil B cells, as shown by immunohistochemistry. Cox-inhibiting drugs modestly affect purified B cell proliferation but profoundly reduce Ab production. The ability of whole blood to produce IgM and IgG following stimulation is also strongly inhibited. In support that Cox-2 plays a seminal role in B lymphocyte Ab production, Cox-2 knockout mice have 64% less IgM and 35% less IgG than normal littermate controls. These findings support that NSAIDs and the new Cox-2-selective drugs have an unsuspected target, the B cell, and attenuate Ab production in humans. Use of NSAIDs may therefore influence autoantibody production in autoimmune diseases and may dampen humoral immunity in response to antigenic challenge/vaccination. PMID:15728468

  11. Abundant expression of myosin heavy-chain IIB RNA in a subset of human masseter muscle fibres

    PubMed Central

    Horton, Michael J.; Brandon, Carla A.; Morris, Terence J.; Braun, Thomas W.; Yaw, Kenneth M.; Sciote, James J.

    2013-01-01

    Type IIB fast fibres are typically demonstrated in human skeletal muscle by histochemical staining for the ATPase activity of myosin heavy-chain (MyHC) isoforms. However, the monoclonal antibody specific for the mammalian IIB isoform does not detect MyHC IIB protein in man and MyHC IIX RNA is found in histochemically identified IIB fibres, suggesting that the IIB protein isoform may not be present in man; if this is not so, jaw-closing muscles, which express a diversity of isoforms, are likely candidates for their presence. ATPase histochemistry, immunohistochemistry polyacrylamide gel electrophoresis and in situ hybridization, which included a MyHC IIB-specific mRNA riboprobe, were used to compare the composition and RNA expression of MyHC isoforms in a human jaw-closing muscle, the masseter, an upper limb muscle, the triceps, an abdominal muscle, the external oblique, and a lower limb muscle, the gastrocnemius. The external oblique contained a mixture of histochemically defined type I, IIA and IIB fibres distributed in a mosaic pattern, while the triceps and gastrocnemius contained only type I and IIA fibres. Typical of limb muscle fibres, the MyHC I-specific mRNA probes hybridized with histochemically defined type I fibres, the IIA-specific probes with type IIA fibres and the IIX-specific probes with type IIB fibres. The MyHC IIB mRNA probe hybridized only with a few histochemically defined type I fibres in the sample from the external oblique; in addition to this IIB message, these fibres also expressed RNAs for MyHC I, IIA and IIX. MyHC IIB RNA was abundantly expressed in histochemical and immunohistochemical type IIA fibres of the masseter, together with transcripts for IIA and in some cases IIX. No MyHC IIB protein was detected in fibres and extracts of either the external oblique or masseter by immunohistochemistry, immunoblotting and electrophoresis. Thus, IIB RNA, but not protein, was found in the fibres of two different human skeletal muscles. It is

  12. STAT3-silenced human dendritic cells have an enhanced ability to prime IFNγ production by both αβ and γδ T lymphocytes.

    PubMed

    Sanseverino, Isabella; Purificato, Cristina; Varano, Barbara; Conti, Lucia; Gessani, Sandra; Gauzzi, M Cristina

    2014-07-01

    Dendritic cells (DC) are an attractive target for therapeutic manipulation of the immune system to enhance insufficient immune responses, such those occurring in cancer, or to dampen dangerous responses in allergic and autoimmune diseases. Main goal of this study was to manipulate human monocyte-derived DC (MDDC) function by silencing STAT3, since this transcription factor plays a key role as a negative regulator of immune surveillance, and is strongly involved in inflammation. STAT3 silencing did not affect the immunophenotype of both immature and toll-like receptor (TLR) ligand-matured DC. However, an altered cytokine secretion profile, characterized by lower IL10 and higher IL12 and TNFα levels, was observed in silenced DC with respect to control cells upon TLR triggering. Accordingly, STAT3 silenced MDDC promoted a higher IFNγ production by CD4(+) naïve T cells. Furthermore, STAT3 silencing in MDDC favored the activation of γδ T lymphocytes, an immune cell population with important antitumor effector activities. This effect was at least in part mediated by the increased IL12 production by silenced cells. STAT3 silencing also increased the levels of CCL4, a CCR5-binding chemokine known to be involved in T helper 1 (Th1) cell recruitment. Altogether these results strengthen the role of STAT3 as a critical check point of the suppression of Th1 responses, unraveling its potential to dampen DC capability to both induce and recruit different IFNγ producing T lymphocyte subsets. PMID:24674241

  13. Development of a chemically defined serum- and protein-free medium for growth of human peripheral lymphocytes.

    PubMed

    Shive, W; Pinkerton, F; Humphreys, J; Johnson, M M; Hamilton, W G; Matthews, K S

    1986-01-01

    A chemically defined, protein-free medium (designated CFBI 1000, where CFBI = Clayton Foundation Biochemical Institute) that supports human peripheral lymphocyte proliferation has been developed. This medium allows exploration of individual metabolic differences by varying the medium composition as well as providing a base to explore further the mechanisms of lymphocyte activation in a system initially free of added macromolecular species other than mitogen. The peripheral blood lymphocyte is an ideal system for metabolic studies because it is easily obtained, is a primary resting cell that can be activated to proliferate, and presumably reflects both the genetic makeup and biochemical environmental history of the individual at the time the cells were formed. Examination of the role of various factors in lymphocyte activation and subsequent events may be simplified by the utilization of a medium that is protein-free and chemically defined. The CFBI 1000 medium supports the growth response of human peripheral lymphocytes to mitogen as measured by [3H]thymidine incorporation to an extent comparable to other media used widely in assessment of lymphocyte proliferation. PMID:3079905

  14. Blackberry seed extracts and isolated polyphenolic compounds showing protective effect on human lymphocytes DNA.

    PubMed

    Gođevac, Dejan; Tešević, Vele; Vajs, Vlatka; Milosavljević, Slobodan; Stanković, Miroslava

    2011-09-01

    The tentative identification of seed extracts from 3 cultivars of blackberry (blackberry seed extracts [BSEs]) constituents was performed by LC/UV/MS technique. The identified compounds belonged to ellagitannins, galic acid derivatives, and ellagic acid derivatives. Two ellagitannins, Lambertianin C and Sanguiniin H-6, and an ellagic acid derivative, 4-α-L-arabinofuranosylellagic acid, were isolated using semipreparative High-performance liquid chromatography. The structure elucidations were based on high resolution-mass spectrometry and nuclear magnetic resonance studies. The BSEs and 3 isolated pure compounds were tested for in vitro protective effect on chromosome aberrations in peripheral human lymphocytes using cytochalasin-B blocked micronucleus (MN) assay. The frequency of MN was scored in binucleated cells, and nuclear proliferation index was calculated. Among the tested extracts, the seeds of cv. Thornfree at concentration of 1 μg/mL exhibit the most prominent effect decreasing the frequency of MN by 62.4%, when compared with the controls cell cultures. Antioxidant potential of pure ellagitannins cannot explain the strong effect of BSEs. The assumption was that better antioxidant effect of BSEs result from synergistic effects of individual compounds contained in the extracts and/or some minor components possessed strong activity. PraCTICAL APPLICATION: Our results provide evidence of protective effects of BSEs and isolated pure compounds on cytogenetic damages in human lymphocytes. Thus, BSEs could exert beneficial effects in quite a few diseases, because many of the biological actions have been attributed to their antioxidant properties. PMID:21824137

  15. Effects of copaene, a tricyclic sesquiterpene, on human lymphocytes cells in vitro.

    PubMed

    Türkez, Hasan; Celik, Kübra; Toğar, Başak

    2014-08-01

    In this study, the cytotoxic, genotoxic/antigenotoxic and antioxidant/oxidant activity of copaene (COP), a plant-derived tricyclic sesquiterpene, on human lymphocyte cultures (n = 5) was investigated. COP was added into culture tubes at various concentrations (0, 10, 25, 50, 100, 200 and 400 mg/L). While the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and lactate dehydrogenase (LDH) assays were used for viability and cytotoxic evaluations, the micronucleus (MN) and sister chromatid exchange (SCE) assays were used for genetic evaluations. Moreover, total antioxidant capacity (TAC) and total oxidative status analysis were used for biochemical evaluations. According to LDH and MTT assays COP significantly reduced cell proliferation at high concentrations (200 and 400 mg/L). In addition, there was no significant increase (P < 0.05) in both SCE and MN frequencies of cultures treated with COP as compared to controls. We have also concluded that concentrations of COP of 50 and 100 mg/L increased TAC level when compared to the controls. In conclusion, in this study it has been reported for the first time that copaene is not genotoxic and it increases the antioxidant capacity in human lymphocyte cultures. PMID:24287609

  16. 50 Hz sinusoidal magnetic fields do not affect human lymphocyte activation and proliferation in vitro

    NASA Astrophysics Data System (ADS)

    Capri, Miriam; Mesirca, Pietro; Remondini, Daniel; Carosella, Simona; Pasi, Sara; Castellani, Gastone; Franceschi, Claudio; Bersani, Ferdinando

    2004-12-01

    In the last 30 years, an increasing public concern about the possible harmful effects of electromagnetic fields generated by power lines and domestic appliances has pushed the scientific community to search for a correct and comprehensive answer to this problem. In this work the effects of exposure to 50 Hz sinusoidal magnetic fields, with a magnetic flux density of 0.05 mT and 2.5 mT (peak values), were studied on human peripheral blood mononuclear cells (PBMCs) collected from healthy young and elderly donors. Cell activation and proliferation were investigated by using flow cytometry techniques and 3H-TdR incorporation assays, respectively. The results obtained indicated that exposure to the fields altered neither DNA synthesis nor the capacity of lymphocytes to enter the activation phase and progress into the cell cycle. Thus, the conclusions are that two important functional phases of human lymphocytes, such as activation and proliferation, are not affected by exposures to 50 Hz magnetic fields similar to those found under power lines.

  17. Evaluation of Possible Genotoxic Activity of Dirithromycin in Cultured Human Lymphocytes

    PubMed Central

    Kayraldız, Ahmet; Dönbak, Lale; Yavuz Kocaman, Ayşe; Köker, Esra; Gökçe, Şule

    2015-01-01

    Dirithromycin antibiotic is a 14-membered lactone ring macrolide and is widely used in medicine to treat many different types of bacterial infections. In the present study, the possible genotoxicity of dirithromycin was evaluated in cultured human lymphocytes by using sister chromatid exchanges (SCEs), chromosome aberration (CA), and micronucleus (MN) tests and also cell proliferation kinetics such as mitotic index (MI), replication index (RI), and nuclear division index (NDI) were analyzed for cytotoxicity. Cell cultures were treated with four different concentrations of dirithromycin (37.75, 67.50, 125, and 250 µg/mL) for 24 and 48 h periods. Dirithromycin significantly induced SCE and MN frequency at all concentrations in both 24 and 48 h treated cells. In addition, CA level has been markedly increased in the cells treated with almost all concentrations of dirithromycin for 24 (except 37.75 µg/mL) and 48 h treatment periods as compared to control. However, MI, RI, and NDI values were not affected by the dirithromycin treatment (p > 0.05). The results of this study indicated that dirithromycin treatment caused genetic damage by increasing the level of cytogenetic endpoints, suggesting its genotoxic and mutagenic action on human lymphocytes in vitro. PMID:26576152

  18. Effects of sotrastaurin, mycophenolic acid and everolimus on human B-lymphocyte function and activation.

    PubMed

    Matz, Mareen; Lehnert, Martin; Lorkowski, Christine; Fabritius, Katharina; Unterwalder, Nadine; Doueiri, Salim; Weber, Ulrike A; Mashreghi, Mir-Farzin; Neumayer, Hans-H; Budde, Klemens

    2012-10-01

    Humoral rejection processes may lead to allograft injury and subsequent dysfunction. Today, only one B-cell-specific agent is in clinical use and the effects of standard and new immunosuppressant substances on B-cell activation and function are not fully clarified. The impact of sotrastaurin, mycophenolic acid and everolimus on human B-lymphocyte function was assessed by analysing proliferation, apoptosis, CD80/CD86 expression and immunoglobulin and IL-10 production in primary stimulated B cells. In addition, B-cell co-cultures with pre-activated T cells were performed to evaluate the effect of the different immunosuppressive agents on T-cell-dependent immunoglobulin production. Sotrastaurin did not inhibit B-cell proliferation, CD80/CD86 expression, and IgG production and had only minor effects on IgM levels at the highest concentration administered. In contrast, mycophenolic acid and everolimus had strong effects on all B-cell functions in a dose-dependent manner. All immunosuppressive agents caused decreased immunoglobulin levels in T-cell-dependent B-cell cultures. The data provided here suggest that mycophenolic acid and everolimus, but not sotrastaurin, are potent inhibitors of human B-lymphocyte function and activation. PMID:22816666

  19. The effect of stem cell from human exfoliated deciduous teeth on T lymphocyte proliferation

    PubMed Central

    Alipour, Razieh; Adib, Minoo; Hashemi-Beni, Batool; Sadeghi, Farzaneh

    2014-01-01

    Background: Mesenchymal stem cells (MSC), a specific type of adult tissue stem cell; have the immunosuppressive effects that make them valuable targets for regenerative medicine and treatment of many human illnesses. Hence, MSC have been the subject of numerous studies. The classical source of MSC is adult bone marrow (BM). Due to many shortcomings of harvesting MSC from BM, finding the alternative sources for MSC is an urgent. Stem cells from human exfoliated deciduous teeth (SHED) are relative new MSC populations that fulfill these criteria but their potential immunosuppressive effect has not been studied enough yet. Thus, in this work the effect of SHED on the proliferation of in vitro activated T lymphocytes were explored. Materials and Methods: In this study, both mitogen and alloantigen activated T cells were cultured in the presence of different numbers of SHED. In some co-cultures, activated T cells were in direct contact to MSCs and in other co-cultures; they were separated from SHED by a permeable membrane. In all co-cultures, the proliferation of T cells was measured by ELISA Bromodeoxyuridine proliferation assay. Results: In general, our results showed that SHED significantly suppress the proliferation of activated T cells in a dose-dependent manner. Moreover, the suppression was slightly stronger when MSCs were in physical contact to activated T cells. Conclusion: This study showed that SHED likewise other MSC populations can suppress the activation of T lymphocytes, which can be used instead of BM derived MSCs in many investigational and clinical applications. PMID:25337532

  20. Lead and cadmium at very low doses affect in vitro immune response of human lymphocytes

    SciTech Connect

    Borella, P.; Giardino, A. )

    1991-08-01

    The effect of lead chloride and cadmium chloride on in vitro immunoglobulin (Ig) production by human lymphocytes was investigated. After 7 days in culture, lead added in the range of human exposure (207-1035 {mu}g/liter) significantly enhanced Ig production either when cells were activated by pokeweed mitogen (PWM) or not. The effect was dose-dependent and was related to the Pb were measured in the extracellular medium and in the cells. Independently of the mitogen addition, about 2% of the Pb added was accumulated in the cells, most being associated with the nuclear fraction. Those findings suggest that the Pb effects could depend on its uptake and distribution in the cells. Cadmium added in the 50-500 nM range exhibited a dose-independent mitogenic activity in unstimulated cells, whereas the Ig secretion was not significantly affected by Cd when cells were PWM-activated. A considerable intraindividual variability, however, was observed when blood donors were separately examined, with both an increase, a decrease, or no variation on Ig production. Furthermore, higher percentages of Cd were accumulated in the nuclear fraction, and lower in the cytosol and precipitate, in PWM-activated compared to resting lymphocytes. Genetic factors could be of importance for the observed variability of the immune response to cadmium, and the authors support the hypothesis that differences in the metallothionein (MT) inducibility could play a role.

  1. Synergistic effect of DHT and IGF-1 hyperstimulation in human peripheral blood lymphocytes.

    PubMed

    Imperlini, Esther; Spaziani, Sara; Mancini, Annamaria; Caterino, Marianna; Buono, Pasqualina; Orrù, Stefania

    2015-06-01

    The abuse of mixed or combined performance-enhancing drugs is widespread among athletes and amateurs, adults and adolescents. Clinical studies demonstrated that misuse of these doping agents is associated with serious adverse effects to many organs in human. Previously, we demonstrated in human peripheral blood lymphocytes that high doses of anabolic androgenic steroids, such as dihydrotestosterone (DHT) and growth factors, such as insulin-like growth factor-1 (IGF-1), have effects at gene and protein levels. Supraphysiological treatments of DHT and IGF-1 affected the expression of genes involved in skeletal muscle disorders as well as in cell-mediated immunological response. At protein level, DHT hyperdosage affects cell motility and apoptosis; IGF-1 hyperstimulation triggers an active cytoskeletal reorganization and an overproduction of immune response- and inflammation-related cytokines. In this study, we investigate the combined effects of DHT and IGF-1 hyperdosage in peripheral blood lymphocytes using a differential proteomic approach. DHT and IGF-1 combined treatment affects cell adhesion, migration, and survival through modulation of expression levels of cytokines and paxillin-signaling-related proteins, and activation of several pathways downstream focal adhesion kinase. Our results indicate a synergistic effect of DHT and IGF-1 which has potential implications for health risk factors. PMID:25669835

  2. Cytogenetic response to coffee in Chinese hamster ovary AUXB1 cells and human peripheral lymphocytes.

    PubMed

    Tucker, J D; Taylor, R T; Christensen, M L; Strout, C L; Hanna, M L

    1989-09-01

    We have investigated the genotoxic effects of three different brands and three types of coffee (freshly brewed regular, instant regular and freshly brewed decaffeinated) in two mammalian systems: the Chinese hamster ovary (CHO) AUXB1 cell line and human peripheral lymphocytes. Sister-chromatid exchanges (SCEs) and endoreduplicated cells (ERCs) were used as the endpoints. Coffee was prepared according to the manufacturer's suggestions, and after cooling, administered to cultured cells at dilutions ranging up to 11% that of full-strength coffee. Each brand and type of coffee induced significant levels of SCEs and ERCs in AUXB1 cells. SCEs, but not ERCs, were induced in human peripheral lymphocytes. Bisulfite, which complexes with carbonyls and reduces their genotoxicity, significantly diminished the number of SCEs and ERCs found after administration of coffee. Catalase and peroxidase, enzymes that destroy hydrogen peroxide activity, had no significant effect upon the SCE and ERC frequencies in AUXB1 cultures treated with freshly brewed regular coffee. These experiments indicate that different brands and types of coffee have sufficient genotoxic activity to increase SCEs and ERCs at levels only a fraction of those normally consumed. 1,2-Dicarbonyls alone and peroxides alone do not appear to be responsible for the majority of SCEs and ERCs that were observed to be induced by dilute coffee. PMID:2687627

  3. Micronucleus assay in human lymphocytes after exposure to alloxydim sodium herbicide in vitro.

    PubMed

    Akyıl, Dilek; Özkara, Arzu; Erdoğmuş, S Feyza; Eren, Yasin; Konuk, Muhsin; Sağlam, Esra

    2015-12-01

    This study evaluates the cytotoxic and genotoxic potential of alloxydim sodium using micronucleus (MN) assay, in human peripheral lymphocytes. MN assay was used to investigate the genotoxic effects of alloxydim sodium in human peripheral lymphocytes treated with 250, 500, 750, 1,000 µg/ml concentrations of alloxydim sodium for 24 and 48 h. Solvent, negative and positive controls were also used in the experiments in parallel. The obtained results were evaluated in statistical analyses by using Dunnett-t test (two sided) and p < 0.05 was accepted as significant. Alloxydim sodium significantly increased the MN formation compared with the negative control, at both 750 and 1,000 µg/ml concentrations and treatment periods. We also evaluated the nuclear division index (NDI) for cytotoxicity of this pesticide in the experiment, and finally observed a significant decrease of the NDI values at all concentrations of alloxydim sodium and at both treatment periods. PMID:25017922

  4. Grapevine fruit extract protects against radiation-induced oxidative stress and apoptosis in human lymphocyte.

    PubMed

    Singha, Indrani; Das, Subir Kumar

    2015-11-01

    Ionizing radiation (IR) causes oxidative stress through overwhelming generation of reactive oxygen species (ROS) in the living cells leading the oxidative damage further to biomolecules. Grapevine (Vitis vinifera L.) posses several bioactive phytochemicals and is the richest source of antioxidants. In this study, we investigated V. vinifera for its phytochemical content, enzymes profile and, ROS- and oxidant-scavenging activities. We have also studied the fruit extract of four different grapevine viz., Thompson seedless, Flame seedless, Kishmish chorni and Red globe for their radioprotective actions in human lymphocytes. The activities of ascorbic acid oxidase and catalase significantly (P < 0.01) differed among extracts within the same cultivar, while that of peroxidase and polyphenol oxidase did not differ significantly. The superoxide radical-scavenging activity was higher in the seed as compared to the skin or pulp of the same cultivar. Pretreatment with grape extracts attenuated the oxidative stress induced by 4 Gy γ-radiation in human lymphocytes in vitro. Further, γ-radiation-induced increase in caspase 3/7 activity was significantly attenuated by grape extracts. These results suggest that grape extract serve as a potential source of natural antioxidants against the IR-induced oxidative stress and also inhibit apoptosis. Furthermore, the protective action of grape depends on the source of extract (seed, skin or pulp) and type of the cultivars. PMID:26669019

  5. The effects of boric acid on sister chromatid exchanges and chromosome aberrations in cultured human lymphocytes

    PubMed Central

    Arslan, Mehmet; Topaktas, Mehmet

    2007-01-01

    The aim of this study was to determine the possible genotoxic effects of boric acid (BA) (E284), which is used as an antimicrobial agent in food, by using sister chromatid exchange (SCEs) and chromosome aberration (CAs) tests in human peripheral lymphocytes. The human lymphocytes were treated with 400, 600, 800, and 1000 μg/mL concentrations of BA dissolved in dimethyl sulfoxide (DMSO), for 24 h and 48 h treatment periods. BA did not increase the SCEs for all the concentrations and treatment periods when compared to control and solvent control (DMSO). BA induced structural and total CAs at all the tested concentrations for 24 and 48 h treatment periods. The induction of the total CAs was dose dependent for the 24 h treatment period. However, BA did not cause numerical CAs. BA showed a cytotoxic effect by decreasing the replication index (RI) and mitotic index (MI). BA decreased the MI in a dose-dependent manner for the 24 h treatment period. PMID:19002846

  6. Evaluation of DNA damage induced by norcantharidin in human cultured lymphocytes.

    PubMed

    Khabour, Omar F; Enaya, Fatima M; Alzoubi, Karem; Al-Azzam, Sayer I

    2016-07-01

    Norcantharidin (NCTD) is currently used in the treatment of several cancers such as leukemia, melanoma and hepatoma. The mechanism of action of NCTD is suggested to involve induction of apoptosis of cancer cells via production of reactive oxygen species. In this study, the genotoxic effect of different concentrations of NCTD (1, 10 and 20 μm) in human lymphocytes was investigated using sister chromatid exchanges (SCEs) and chromosomal aberrations (CAs) assays. The results revealed that NCTD significantly increased the rate of SCEs (p < 0.05) in a dose-dependent manner. In addition, NCTD significantly increased the number of high-frequency cells (SCEs ≥ 8, p < 0.05). However, NCTD did not have any significant effect on the rate of CAs (p > 0.05). In addition, no significant differences were detected in the mitotic index or proliferative index at examined doses (up to 20 μm). In conclusion, NCTD is genotoxic to human cultured lymphocytes as measured by SCE assay. PMID:26599593

  7. Diversified Expression of NG2/CSPG4 Isoforms in Glioblastoma and Human Foetal Brain Identifies Pericyte Subsets

    PubMed Central

    Rizzi, Marco; Errede, Mariella; Wälchli, Thomas; Mucignat, Maria Teresa; Frei, Karl; Roncali, Luisa; Perris, Roberto; Virgintino, Daniela

    2013-01-01

    NG2/CSPG4 is a complex surface-associated proteoglycan (PG) recognized to be a widely expressed membrane component of glioblastoma (WHO grade IV) cells and angiogenic pericytes. To determine the precise expression pattern of NG2/CSPG4 on glioblastoma cells and pericytes, we generated a panel of >60 mouse monoclonal antibodies (mAbs) directed against the ectodomain of human NG2/CSPG4, partially characterized the mAbs, and performed a high-resolution distributional mapping of the PG in human foetal, adult and glioblastoma-affected brains. The reactivity pattern initially observed on reference tumour cell lines indicated that the mAbs recognized 48 immunologically distinct NG2/CSPG4 isoforms, and a total of 14 mAbs was found to identify NG2/CSPG4 isoforms in foetal and neoplastic cerebral sections. These were consistently absent in the adult brain, but exhibited a complementary expression pattern in angiogenic vessels of both tumour and foetal tissues. Considering the extreme pleomorphism of tumour areas, and with the aim of subsequently analysing the distributional pattern of the NG2/CSPG4 isoforms on similar histological vessel typologies, a preliminary study was carried out with endothelial cell and pericyte markers, and with selected vascular basement membrane (VBM) components. On both tumour areas characterized by 'glomeruloid' and 'garland vessels', which showed a remarkably similar cellular and molecular organization, and on developing brain vessels, spatially separated, phenotypically diversified pericyte subsets with a polarized expression of key surface components, including NG2/CSPG4, were disclosed. Interestingly, the majority of the immunolocalized NG2/CSPG4 isoforms present in glioblastoma tissue were present in foetal brain, except for one isoform that seemed to be exclusive of tumour cells, being absent in foetal brain. The results highlight an unprecedented, complex pattern of NG2/CSPG4 isoform expression in foetal and neoplastic CNS, discriminating

  8. Evaluation of toxicity of essential oils palmarosa, citronella, lemongrass and vetiver in human lymphocytes.

    PubMed

    Sinha, Sonali; Jothiramajayam, Manivannan; Ghosh, Manosij; Mukherjee, Anita

    2014-06-01

    The present investigation was undertaken to study the cytotoxic and genotoxic potential of the essential oils (palmarosa, citronella, lemongrass and vetiver) and monoterpenoids (citral and geraniol) in human lymphocytes. Trypan blue dye exclusion and MTT test was used to evaluate cytotoxicity. The genotoxicity studies were carried out by comet and DNA diffusion assays. Apoptosis was confirmed by Annexin/PI double staining. In addition, generation of reactive oxygen species was evaluated by DCFH-DA staining using flow cytometry. The results demonstrated that the four essential oils and citral induced cytotoxicity and genotoxicity at higher concentrations. The essential oils were found to induce oxidative stress evidenced by the generation of reactive oxygen species. With the exception of geraniol, induction of apoptosis was confirmed at higher concentrations of the test substances. Based on the results, the four essential oils are considered safe for human consumption at low concentrations. PMID:24650756

  9. Loss of Telomeres in the Progeny of Human Lymphocytes Exposed to Energetic Heavy Ions

    NASA Technical Reports Server (NTRS)

    Cucinotta, F.A.; George, K.; Durante, M.

    2006-01-01

    We have used cross-species multi-color banding (RxFISH) combined with telomere FISH probes, to measure chromosomal aberrations in the progeny of human peripheral blood lymphocytes exposed to ionizing radiation. Accelerated iron particles (energy 1 GeV/nucleon) induced many more terminal deletions than the same dose of gamma-rays. We found that truncated chromosomes without telomeres could be transmitted for at least three cell cycles following exposure, and represented about 10% of all aberrations observed in the progeny of cells exposed to iron ions. High energy heavy ions generate the most significant health risk for human space exploration and the results suggest that telomere loss may be the leading mechanism for their high efficiency in the induction of late effects.

  10. Beta-casomorphin (BCM) and human colonic lamina propria lymphocyte proliferation.

    PubMed Central

    Elitsur, Y; Luk, G D

    1991-01-01

    BCM is a milk-derived peptide with opiate-like properties which is absorbed through the gastrointestinal mucosa. It has been shown to affect gastrointestinal motility, absorption and secretion. Recently, modulation of the immune system by BCM was also reported. In this study we investigated the in vitro effect of BCM on the human mucosal immune response as represented by lamina propria lymphocyte (LPL) proliferation. Results show that BCM significantly inhibited concanavalin A (ConA) stimulated LPL DNA synthesis. BCM also inhibited ornithine decarboxylase activity (ODC) in ConA-stimulated LPL. Although BCM also inhibited 12-O-tetradecanoyl phorbol-13-acetate (TPA) stimulated LPL DNA synthesis, the degree of inhibition was much lower than in ConA-stimulated LPL. The anti-proliferative effect of BCM was reversed by the opiate receptor antagonist, neloxone. Our results suggest that BCM may affect the human mucosal immune system, possibly via the opiate receptor. PMID:1893631

  11. Origin and Role of a Subset of Tumor-Associated Neutrophils with Antigen-Presenting Cell Features in Early-Stage Human Lung Cancer.

    PubMed

    Singhal, Sunil; Bhojnagarwala, Pratik S; O'Brien, Shaun; Moon, Edmund K; Garfall, Alfred L; Rao, Abhishek S; Quatromoni, Jon G; Stephen, Tom Li; Litzky, Leslie; Deshpande, Charuhas; Feldman, Michael D; Hancock, Wayne W; Conejo-Garcia, Jose R; Albelda, Steven M; Eruslanov, Evgeniy B

    2016-07-11

    Based on studies in mouse tumor models, granulocytes appear to play a tumor-promoting role. However, there are limited data about the phenotype and function of tumor-associated neutrophils (TANs) in humans. Here, we identify a subset of TANs that exhibited characteristics of both neutrophils and antigen-presenting cells (APCs) in early-stage human lung cancer. These APC-like "hybrid neutrophils," which originate from CD11b(+)CD15(hi)CD10(-)CD16(low) immature progenitors, are able to cross-present antigens, as well as trigger and augment anti-tumor T cell responses. Interferon-γ and granulocyte-macrophage colony-stimulating factor are requisite factors in the tumor that, working through the Ikaros transcription factor, synergistically exert their APC-promoting effects on the progenitors. Overall, these data demonstrate the existence of a specialized TAN subset with anti-tumor capabilities in human cancer. PMID:27374224

  12. p53 mutations in human lymphoid malignancies: Association with Burkitt lymphoma and chronic lymphocytic leukemia

    SciTech Connect

    Gaidano, G.; Ballerini, P.; Gong, J.Z.; Inghirami, G.; Knowles, D.M.; Dalla-Favera, R. ); Neri, A, Centro Malattie del Sangue G. Marcora, Milan ); Newcomb, E.W. ); Magrath, I.T. )

    1991-06-15

    The authors have investigated the frequency of p53 mutations in B- and T-cell human lymphoid malignancies, including acute lymphoblastic leukemia, the major subtypes of non-Hodgkin lymphoma, and chronic lymphocytic leukemia. p53 exons 5-9 were studied by using genomic DNA from 197 primary tumors and 27 cell lines by single-strand conformation polymorphism analysis and by direst sequencing of PCR-amplified fragments. Mutations were found associated with (i) Burkitt lymphoma (9/27 biopsoes; 17/27 cell lines) and its leukemic counterpart L{sub 3}-type B-cell acute lymphoblastic leukemia (5/9), both of which also carry activated c-myc oncogenes, and (ii) B-cell chronic lymphocytic leukemia (6/40) and, in particular, its stage of progression known as Richter's transformation (3/7). Mutations were not found at any significant frequency in other types of non-Hodgkin lymphoma or acute lymphoblastic leukemia. In many cases, only the mutated allele was detectable, implying loss of the normal allele. These results suggest that (1) significant differences in the frequency of p53 mutations are present among subtypes of neoplasms derived from the same tissue; (2) p53 may play a role in tumor progression in B-cell chronic lymphocytic leukemia; (3) the presence of both p53 loss/inactivation and c-myc oncogene activation may be important in the pathogenesis of Burkitt lymphoma and its leukemia form L{sub 3}-type B-cell acute lymphoblastic leukemia.

  13. Induction of adaptive response in human blood lymphocytes exposed to radiofrequency radiation.

    PubMed

    Sannino, Anna; Sarti, Maurizio; Reddy, Siddharth B; Prihoda, Thomas J; Vijayalaxmi; Scarfì, Maria Rosaria

    2009-06-01

    The incidence of micronuclei was evaluated to assess the induction of an adaptive response to non-ionizing radiofrequency (RF) radiation in peripheral blood lymphocytes collected from five different human volunteers. After stimulation with phytohemagglutinin for 24 h, the cells were exposed to an adaptive dose of 900 MHz RF radiation used for mobile communications (at a peak specific absorption rate of 10 W/kg) for 20 h and then challenged with a single genotoxic dose of mitomycin C (100 ng/ml) at 48 h. Lymphocytes were collected at 72 h to examine the frequency of micronuclei in cytokinesis-blocked binucleated cells. Cells collected from four donors exhibited the induction of adaptive response (i.e., responders). Lymphocytes that were pre-exposed to 900 MHz RF radiation had a significantly decreased incidence of micronuclei induced by the challenge dose of mitomycin C compared to those that were not pre-exposed to 900 MHz RF radiation. These preliminary results suggested that the adaptive response can be induced in cells exposed to non-ionizing radiation. A similar phenomenon has been reported in cells as well as in animals exposed to ionizing radiation in several earlier studies. However, induction of adaptive response was not observed in the remaining donor (i.e., non-responder). The incidence of micronuclei induced by the challenge dose of mitomycin C was not significantly different between the cells that were pre-exposed and unexposed to 900 MHz RF radiation. Thus the overall data indicated the existence of heterogeneity in the induction of an adaptive response between individuals exposed to RF radiation and showed that the less time-consuming micronucleus assay can be used to determine whether an individual is a responder or non-responder. PMID:19580480

  14. Stability of Radiation Induced Chromosome Damage in Human Peripheral Blood Lymphocytes

    NASA Technical Reports Server (NTRS)

    Cucinotta, F. A.; George, K.; Willingham, V.

    2006-01-01

    Chromosome damage in an individual's peripheral blood lymphocytes can be an indicator of radiation exposure and this data can be used to evaluate dose after accidental or occupational exposure. Evidence suggests that the yield of chromosome damage in lymphocytes is also a relevant biomarker of cancer risk in humans that reflects individual cancer susceptibility. It follows that biomonitoring studies can be used to uncover subjects who are particularly susceptible to radiation damage and therefore at higher risk of cancer. Translocations and other stable aberrations are commonly believed to persist in peripheral blood cells for many years after exposure, and it has been suggested that translocations can be used for assessing retrospective radiation doses or chronic exposures. However, recent investigations suggest that translocations might not always persist indefinitely. We measured chromosome aberrations in the blood lymphocytes of six astronauts before their respective missions of approximately 3 to 6 months onboard the international space station, and again at various intervals up to 5 years after flight. In samples collected a few days after return to earth, the yield of chromosome translocations had significantly increased compared with preflight values, and results indicate that biodosimetry estimates lie within the range expected from physical dosimetry. However, for five of the astronauts, follow up analysis revealed a temporal decline in translocations with half-lives ranging from 10 to 58 months. The yield of exchanges remained unchanged for the sixth astronaut during an observation period of 5 months post-flight. These results may indicate complications with the use of stable aberrations for retrospective dose reconstruction and could affect cancer risk predictions that are estimated from yields of chromosome damage obtained shortly after exposure.

  15. Rapid alterations of cell cycle control proteins in human T lymphocytes in microgravity

    PubMed Central

    2012-01-01

    In our study we aimed to identify rapidly reacting gravity-responsive mechanisms in mammalian cells in order to understand if and how altered gravity is translated into a cellular response. In a combination of experiments using "functional weightlessness" provided by 2D-clinostats and real microgravity provided by several parabolic flight campaigns and compared to in-flight-1g-controls, we identified rapid gravity-responsive reactions inside the cell cycle regulatory machinery of human T lymphocytes. In response to 2D clinorotation, we detected an enhanced expression of p21 Waf1/Cip1 protein within minutes, less cdc25C protein expression and enhanced Ser147-phosphorylation of cyclinB1 after CD3/CD28 stimulation. Additionally, during 2D clinorotation, Tyr-15-phosphorylation occurred later and was shorter than in the 1 g controls. In CD3/CD28-stimulated primary human T cells, mRNA expression of the cell cycle arrest protein p21 increased 4.1-fold after 20s real microgravity in primary CD4+ T cells and 2.9-fold in Jurkat T cells, compared to 1 g in-flight controls after CD3/CD28 stimulation. The histone acetyltransferase (HAT) inhibitor curcumin was able to abrogate microgravity-induced p21 mRNA expression, whereas expression was enhanced by a histone deacetylase (HDAC) inhibitor. Therefore, we suppose that cell cycle progression in human T lymphocytes requires Earth gravity and that the disturbed expression of cell cycle regulatory proteins could contribute to the breakdown of the human immune system in space. PMID:22273506

  16. DNA damage and repair in human peripheral blood lymphocytes following treatment with microcystin-LR.

    PubMed

    Lankoff, Anna; Krzowski, Łukasz; Głab, Joanna; Banasik, Anna; Lisowska, Halina; Kuszewski, Tomasz; Góźdź, Stanisław; Wójcik, Andrzej

    2004-04-11

    The purpose of this study was to find a possible explanation of the inconsistency of data regarding the genotoxicity of microcystin-LR (MC-LR). We compared the results of the comet assay with the results of the analysis of chromosome aberrations and apoptosis. In order to investigate the influence of MC-LR on DNA damage in human lymphocytes, cells were treated with MC-LR at different concentrations (1, 10 and 25 microg/ml) for 6, 12, 18 and 24 h. Analyses of Olive Tail Moment (OTM) as an indicator of DNA damage showed that MC-LR treatment induced DNA damage in a time-dependent manner, reaching its maximum after 18 h. The lowest values of OTM were observed after 24 h. MC-LR had no effect on the frequency of chromosome aberrations in lymphocytes. Since some data available in the literature indicate that apoptosis may lead to overestimated or false positive results regarding the genotoxicity of mutagens in the comet assay, we measured the frequency of late apoptotic cells by use of the comet assay and the frequency of early apoptotic cells with the TUNEL method. The comet assay results revealed that the highest level of apoptosis was observed after 24 h and the lowest after 18 h. The comparison of the frequency of apoptotic cells determined by the comet assay with DNA damage (OTM) examined by the comet assay revealed a statistically significant, negative correlation. The TUNEL results showed that the frequency of apoptotic cells progressively increased in a dose- and time-dependent manner. The comparison of the frequency of apoptotic cells determined by TUNEL method with DNA damage (OTM) examined by the comet assay showed a significant positive correlation for lymphocytes treated with MC-LR for 6, 12 and 18 h. Therefore, our findings indicate that microcystin-LR-induced DNA damage observed in the comet assay may be related to the early stages of apoptosis due to cytotoxicity but not genotoxicity. In addition, we examined the DNA repair kinetics in lymphocytes

  17. Genotoxicity of di-butyl-phthalate and di-iso-butyl-phthalate in human lymphocytes and mucosal cells.

    PubMed

    Kleinsasser, N H; Wallner, B C; Kastenbauer, E R; Weissacher, H; Harréus, U A

    2001-01-01

    The genotoxicity of phthalates, widely used plasticizers, has been shown previously for di-butyl-phthalate (DBP) and di-iso-butyl-phthalate (DBP) in human mucosal cells of the upper aerodigestive tract in a previous study using the Comet assay. Furthermore, higher genotoxic sensitivities of patients with squamous cell carcinomas of either the larynx or the oropharynx compared to non-tumor patients were described. Other authors have demonstrated DNA damage by a different phthalate in human lymphocytes. It was the aim of the present study to determine whether there is a correlation between the genotoxic sensitivities to DBP and its isomer DiBP in either mucosal cells or lymphocytes. The single-cell microgel electrophoresis assay (Comet assay) was applied to detect DNA strand breaks in human epithelial cells of the upper aerodigestive tract (n=132 specimens). Human mucosa was harvested from the oropharynx in non-tumor patients and patients with squamous cell carcinomas of the oropharynx. Laryngeal mucosa of patients with laryngeal squamous cell carcinomas was harvested as well. Peripheral lymphocytes (n=49 specimens) were separated from peripheral blood. Xenobiotics investigated were DBP, DiBP, and N'methyl-N'-nitro-N-nitrosoguanidine (MNNG) as positive control, respectively. For statistical analysis, the SPSS correlation analysis according to Pearson and the Wilcoxon test were performed. Genotoxicity was found for DBP and DiBP in epithelial cells and lymphocytes (P<0.001). MNNG caused severe DNA damage. In analyzing DBP and DiBP results, genotoxic impacts in mucosal cells showed an intermediate correlation (r=0.570). Correlation in lymphocytes was the same (r=0.570). Phthalates have been investigated as a potential health hazard for a variety of reasons, including possible xenoestrogenic impact, peroxisome proliferation, and membrane destabilization. The present investigation suggests a correlated DNA-damaging impact of DBP and DiBP in human mucosal cells and in

  18. Correlation between radiation dose and p53 protein expression levels in human lymphocytes.

    PubMed

    Cavalcanti, Mariana B; Fernandes, Thiago S; Silva, Edvane B; Amaral, Ademir

    2015-09-01

    The aim of this research was to evaluate the relationship between p53 protein levels and absorbed doses from in vitro irradiated human lymphocytes. For this, samples of blood from 23 donors were irradiated with 0.5; 1; 2; and 4 Gy from a Cobalt-60 source, and the percentages of lymphocytes expressing p53 were scored using Flow Cytometry. The subjects were divided into 3 groups, in accordance with the p53 levels expressed per radiation dose: low (Group I), high (Group II), and excessive levels (Group III). For all groups, the analyses showed that the p53 expression levels increase with the absorbed dose. Particularly for groups I and II, the correlation between this protein expression and the dose follows the linear-quadratic model, such as for radioinduced chromosomal aberrations. In conclusion, our findings indicate possible applications of this approach in evaluating individual radiosensitivity prior to radiotherapeutical procedures as well as in medical surveillance of occupationally exposed workers. Furthermore, due to the rapidity of flow-cytometric analyses, the methodology here employed would play an important role in emergency responses to a large-scale radiation incident where many people may have been exposed. PMID:26312422

  19. Micronuclei Assessment of The Radioprotective Effects of Melatonin and Vitamin C in Human Lymphocytes

    PubMed Central

    Rostami, Aram; Moosavi, Seyed Akbar; Dianat Moghadam, Hassan; Bolookat, Eftekhar Rajab

    2016-01-01

    Objective Critical macromolecules such as DNA maybe damaged by free radicals that are generated from the interaction of ionizing radiation with biological systems. Melatonin and vitamin C have been shown to be direct free radical scavengers. The aim of this study was to investigate the in vivo/in vitro radioprotective effects of melatonin and vitamin C separately and combined against genotoxicity induced by 6 MV x-ray irradiation in human cultured blood lymphocytes. Materials and Methods In this experimental study, fifteen volunteers were divided into three groups of melatonin, vitamin C and melatonin plus vitamin C treatment. Peripheral blood samples were collected from each group before, and 1, 2 and 3 hours after melatonin and vitamin C administration (separately and combined). The blood samples were then irradiated with 200 cGy of 6 MV x-ray. In order to characterize chromosomal aberrations, the lymphocyte samples were cultured with mitogenic stimulus on cytokinesisblocked binucleated cells. Results The samples collected 1hour after melatonin and vitamin C (separately and combined) ingestion exhibited a significant decrease in the incidence of micronuclei compared with their control group (P<0.05). The maximum synergic protection and reduction in frequency of micronuclei (57%) was observed 1 hour after vitamin C and melatonin administration combined. Conclusion We conclude that simultaneous administration of melatonin and vitamin C as radioprotector substances before irradiation may reduce genotoxicity caused by x-ray irradiation. PMID:27054118

  20. Proliferative kinetics of human lymphocytes in culture measured by autoradiography and sister chromatid differential staining

    SciTech Connect

    Morimoto, K.; Sato, M.; Koizumi, A.

    1983-01-01

    A simple combination of autoradiography, to determine when a cell synthesized DNA, and sister chromatid differential staining, to determine how many times a cell has divided, was used to follow up the proliferating fate of human lymphocytes in culture. Cells were incubated continuously with 5-bromodeoxyuridine (BrdU) and pulse-labelled with 0.1 ..mu..Ci/ml (/sup 3/H)thymidine at various times after stimulation with phytohemagglutinin (PHA). The cells were then harvested at 4 h intervals up to 72 h, and the percentage of labelled mitoses was determined separately in first, second, or third division cells. The data showed that the cycling cells, whether they began cycling at earlier or later times after stimulation, had about the same generation times of 12-14 h. This confirms that the heterogeneity of cell generations seen in short-term lymphocyte cultures is in large part due to the difference in the times when cells began cell cycling in response to PHA. 34 references, 2 figures, 1 table.

  1. Effects of ascorbic acid and sodium ascorbate on cyclic nucleotide metabolism in human lymphocytes.

    PubMed

    Atkinson, J P; Weiss, A; Ito, M; Kelly, J; Parker, C W

    1979-01-01

    L-ascorbic acid (LAA) augmented cGMP many-fold in highly purified human peripheral blood lymphocytes. The cGMP response occurred within 10 sec and persisted for at least 60 min. D-ascorbic acid (DAA) and dehydroascorbic acid (DHAA) were also equally active in enhancing cGMP concentrations but metabolic precursors of ascorbic acid and other inorganic acids did not increase cGMP levels. Determination of the amount of DHAA contaminating the LAA precluded the possibility that it was solely responsible for the enhanced cGMP levels. The sodium or calcium salts of ascorbic acid did not increase cGMP concentrations. If these neutralized preparations were acidified, increased cGMP concentrations were then noted. In broken cell preparations, LAA, DAA, and DHAA and to a lesser extent sodium ascorbate (NaA) enhanced guanylate cyclase activity while neither inhibited cAMP or cGMP phosphodiesterase (PDE) activity. The possible role of H2O2, fatty acid liberation, prostaglandin production, oxidizing-reducing agents, and free radical formation in mediating the effects of ascorbic acid on cGMP levels were evaluated, but none of these potential mechanisms were definitively proven to be a required intermediary for the cGMP enhancing activity of ascorbic acid. LAA, DHAA or NaA did not induce lymphocyte transformation or modulate lectin-induced mitogenesis. PMID:36416

  2. Frequency domain electrical conductivity measurements of the passive electrical properties of human lymphocytes.

    PubMed

    Bordi, F; Cametti, C; Rosi, A; Calcabrini, A

    1993-11-21

    An extensive set of electrical conductivity measurements of human lymphocyte suspensions has been carried out in the frequency range from 1 kHz to 100 MHz, where the surface polarization due to the Maxwell-Wagner effect occurs. The data have been analyzed according to well-established heterogeneous system theories and the passive electrical parameters of both the cytoplasmic and nuclear membranes have been obtained. Moreover, a further analysis to take into account the roughness of the membrane surface on the basis of a fractal model yields new estimates for the membrane conductivity and the membrane permittivity, independently of the surface architecture of the cell. These findings are confirmed by measurements carried out at higher frequencies, in the range from 1 MHz to 1 GHz, on lymphocytes dispersed in both hyperosmotic and hypoosmotic media, that influence the surface complexity of the membrane due to the microvillous protrusions. The surface roughness of the cell is described by a macroscopic parameter (the fractal dimension) whose variations are associated to the progressive swelling of the cell, as the osmolality of the solution is changed. PMID:8241253

  3. Effects of halothane on the human beta-adrenergic receptor of lymphocyte membranes

    SciTech Connect

    Marty, J.; Nivoche, Y.; Nimier, M.; Rocchiccioli, C.; Luscombe, F.; Henzel, D.; Loiseau, A.; Desmonts, J.M.

    1987-12-01

    The effects of halothane on beta-adrenergic receptor antagonist interaction were studied using the membranes of human lymphocytes as a model. Membrane preparations of lymphocytes were obtained from blood samples withdrawn from seven healthy young volunteers. Beta-receptor studies were performed using (-)/sup 125/I iodocyanopindolol (/sup 125/ICP) binding. Non-specific binding was determined in the presence of (-)isoproterenol. Beta-receptor density (Bmax) and the dissociation constant (KD) for /sup 125/ICP were determined from saturation curves. Beta-receptor affinity for agonists evaluated by the IC50 (the concentration of isoproterenol required to inhibit 50% of specific /sup 125/ICP binding) and the dissociation constant (KL) for isoproterenol was established from competition curves. The effect of halothane 1%, in an air oxygen mixture (oxygen fraction: 0.3) administered by tonometry during ligand membrane incubation, on beta-adrenergic receptor, was compared to that of control experiments not exposed to halothane. Halothane produced a moderate but significant decrease of Bmax (-10%) and a significant increase in non-specific binding (+30%), while KD, IC50, and KL were unchanged. The authors conclude that halothane, in vitro, decreases beta-adrenergic receptor density. This effect could be mediated by an alteration of the receptor in the membrane due to action of halothane on the lipid phase of the membrane.

  4. Induction of sister chromatid exchanges by coal dust and tobacco snuff extracts in human peripheral lymphocytes

    SciTech Connect

    Tucker, J.D.; Ong, T.

    1985-01-01

    The organic solvent extracts of sub-bituminous coal dust and tobacco snuff, both together and separately, were tested for the induction of sister chromatid exchanges (SCEs) in human peripheral lymphocytes. The results indicate that these extracts induced SCEs, and that when tested together synergistically induced SCEs in two of three donors. Studies with the organic solvent extracts of all five ranks of coal indicate that the extracts of bituminous, lignite, and peat, but not anthracite, induced SCEs. Similar experiments conducted with water extracts, induced SCEs, and that anthracite was equivocal. To determine whether individuals differed in their SCE responses to coal dust extracts, lymphocytes from five donors were tested with organic solvent extracts of bituminous and sub-bituminous coal. An analysis of variance indicates that the SCE response was significantly influenced by the donor and each of the two coal extracts. The findings presented here suggest that coal dust, with or without tobacco snuff, may play a role in the elevated incidence of gastric cancer in coal miners. Because water extracts of some ranks of coal induced SCEs, there exists the possibility of adverse environmental effects due to coal leachates.

  5. Growth hormone protects human lymphocytes from irradiation-induced cell death

    PubMed Central

    Lempereur, Laurence; Brambilla, Daria; Maria Scoto, Giovanna; D'Alcamo, Maria; Goffin, Vincent; Crosta, Lucia; Palmucci, Tullio; Rampello, Liborio; Bernardini, Renato; Cantarella, Giuseppina

    2003-01-01

    Undesired effects of cancer radiotherapy mainly affect the hematopoietic system. Growth hormone (GH) participates in both hematopoiesis and modulation of the immune response. We report both r-hGH cell death prevention and restoration of secretory capacities of irradiated human peripheral blood lymphocytes (PBL) in vitro. r-hGH induced cell survival and increased proliferation of irradiated cells. Western blot analysis indicated that these effects of GH were paralleled by increased expression of the antiapoptotic protein Bcl-2. r-hGH restored mitogen-stimulated release of IL-2 by PBL. Preincubation of irradiated lymphocytes with the growth hormone receptor (GHR) antagonists B2036 and G120 K abrogated r-hGH-dependent IL-2 release. These results demonstrate that r-hGH protects irradiated PBL from death in a specific, receptor-mediated manner. Such effect of r-hGH on PBL involves activation of the antiapoptotic gene bcl-2 and prevention of cell death, associated with preserved functional cell capacity. Finally, potential use of GH as an immunopotentiating agent could be envisioned during radiation therapy of cancer. PMID:12721095

  6. Biodosimetry of ionizing radiation by selective painting of prematurely condensed chromosomes in human lymphocytes

    NASA Technical Reports Server (NTRS)

    Durante, M.; George, K.; Yang, T. C.

    1997-01-01

    Painting of interphase chromosomes can be useful for biodosimetric purposes in particular cases such as radiation therapy, accidental exposure to very high radiation doses and exposure to densely ionizing radiation, for example during space missions. Biodosimetry of charged-particle radiation is analyzed in the present paper. Target cells were human peripheral blood lymphocytes irradiated in vitro with gamma rays, protons and iron ions. After exposure, lymphocytes were incubated for different times to allow repair of radiation-induced damage and then fused to mitotic hamster cells to promote premature condensation in the interphase chromosomes. Chromosome spreads were then hybridized with whole-chromosome DNA probes labeled with fluorescent stains. Dose-response curves for the induction of chromatin fragments shortly after exposure, as well as the kinetics of rejoining and misrejoining, were not markedly dependent on linear energy transfer. However, after exposure to heavy ions, more aberrations were scored in the interphase cells after incubation for repair than in metaphase samples harvested at the first postirradiation mitosis. On the other hand, no significant differences were observed in the two samples after exposure to sparsely ionizing radiation. These results suggest that interphase chromosome painting can be a useful tool for biodosimetry of particle radiation.

  7. Karyotypes of Human Lymphocytes Exposed to High-Energy Iron Ions

    NASA Technical Reports Server (NTRS)

    Durante, M.; George, K.; Wu, H.; Cucinotta, F. A.

    2002-01-01

    Chromosomal aberrations were analyzed using multicolor fluorescence in situ hybridization (mFISH) in human peripheral blood lymphocytes after in vitro exposure to gamma rays or accelerated (56)Fe ions (1 GeV/nucleon, 145 keV/microm) at Brookhaven National Laboratory (Upton, NY). Doses of 0.3 and 3 Gy were used for both radiation types. Chromosomes were prematurely condensed by a phosphatase inhibitor (calyculin A) to avoid the population selection bias observed at metaphase as a result of the severe cell cycle delays induced by heavy ions. A total of 1053 karyotypes (G(2) and M phases) were analyzed in irradiated lymphocytes. Results revealed different distribution patterns for chromosomal aberrations after low- and high-LET radiation exposures: Heavy ions induced a much higher fraction of cells with multiple aberrations, while the majority of the aberrant cells induced by low doses of gamma rays contained a single aberration. The high fraction of complex-type exchanges after heavy ions leads to an overestimation of simple-type asymmetrical interchanges (dicentrics) from analysis of Giemsa-stained samples. However, even after a dose of 3 Gy iron ions, about 30% of the cells presented no complex-type exchanges. The involvement of individual chromosomes in exchanges was similar for densely and sparsely ionizing radiation, and no statistically significant evidence of a nonrandom involvement of specific chromosomes was detected.

  8. Reduction of chrysotile asbestos-induced genotoxicity in human peripheral blood lymphocytes by garlic extract.

    PubMed

    Bhattacharya, Kunal; Yadava, Santosh; Papp, Thilo; Schiffmann, Dietmar; Rahman, Qamar

    2004-11-28

    Asbestos fibers are well known environmental carcinogen, however, the underlying mechanisms of their action have still not clearly been identified. Asbestos is capable of depleting glutathione and generating reactive oxygen species (ROS), which are important mediators of damage in biological system. Asbestos-induced mutagenecity, may be mediated by the generation. It is known that a number of scavengers and antioxidants attenuate asbestos-induced ROS release. Furthermore, it is known that garlic, contains numerous sulfur compounds and glutathione precursors which act as antioxidants and also demonstrate anticarcinogenic properties. The aim of this study was to investigate whether garlic extract has any influence on asbestos-mediated genotoxicity. As an assay system, we applied the micronucleus assay, sister chromatid exchanges, and chromosomal aberrations with human peripheral blood lymphocytes, which has already been used to analyze the genotoxicity of asbestos fibers. Our results indicate that garlic extract, when administered to the lymphocytes cell culture simultaneously with chrysotile reduced the rates of micronucleus formation, sister chromatid exchanges, and chromosomal aberrations significantly. We conclude that garlic extract may be an efficient, physiologically tolerable quencher of asbestos-mediated genotoxicity. PMID:15454308

  9. CD8low CD100− T Cells Identify a Novel CD8 T Cell Subset Associated with Viral Control during Human Hantaan Virus Infection

    PubMed Central

    Liu, Bei; Ma, Ying; Zhang, Yusi; Zhang, Chunmei; Yi, Jing; Zhuang, Ran; Yu, Haitao; Yang, Angang

    2015-01-01

    ABSTRACT Hantaan virus (HTNV) infection can cause a severe lethal hemorrhagic fever with renal syndrome (HFRS) in humans. CD8+ T cells play a critical role in combating HTNV infections. However, the contributions of different CD8+ T cell subsets to the immune response against viral infection are poorly understood. Here, we identified a novel subset of CD8+ T cells characterized by the CD8low CD100− phenotype in HFRS patients. The CD8low CD100− subset accounted for a median of 14.3% of the total CD8+ T cells in early phase of HFRS, and this percentage subsequently declined in the late phase of infection, whereas this subset was absent in healthy controls. Furthermore, the CD8low CD100− cells were associated with high activation and expressed high levels of cytolytic effector molecules and exhibited a distinct expression profile of effector CD8+ T cells (CCR7+/− CD45RA− CD127high CD27int CD28low CD62L−). When stimulated with specific HTNV nucleocapsid protein-derived peptide pools, most responding CD8+ cells (gamma interferon [IFN-γ] positive and/or tumor necrosis factor alpha [TNF-α] positive) were CD8low CD100− cells. The frequency of CD8low CD100− cells among HTNV-specific CD8+ T cells was higher in milder cases than in more severe cases. Importantly, the proportion of the CD8low CD100− subset among CD8+ T cells in early phase of HFRS was negatively correlated with the HTNV viral load, suggesting that CD8low CD100− cells may be associated with viral clearance. The contraction of the CD8low CD100− subset in late phase of infection may be related to the consistently high expression levels of PD-1. These results may provide new insights into our understanding of CD8+ T cell-mediated protective immunity as well as immune homeostasis after HTNV infection in humans. IMPORTANCE CD8+ T cells play important roles in the antiviral immune response. We found that the proportion of CD8low CD100− cells among CD8+ T cells from HFRS patients was

  10. Rejoining and misrejoining of radiation-induced chromatin breaks. I. experiments with human lymphocytes

    NASA Technical Reports Server (NTRS)

    Durante, M.; George, K.; Wu, H.; Yang, T. C.

    1996-01-01

    Fluorescence in situ hybridization with a composite probe for human chromosome 4 and a probe that stained all centromeres was used to study gamma-ray induced breakage, rejoining and misrejoining in prematurely condensed chromosomes in human lymphocytes. Dose-response curves for the induction of all types of aberrations in prematurely condensed human chromosomes 4 were determined immediately after irradiation and after 8 h postirradiation incubation. In addition, aberrations were measured after various incubation times from 0 to 18 h after a dose of 7 Gy. Unrejoined chromosome breaks were the most frequent type of aberration observed immediately after irradiation. Approximately 15% of total aberrations observed were chromosome exchanges. After 8 h postirradiation incubation, the frequency of breaks in prematurely condensed chromosomes declined to about 20% of the initial value, and chromosomal exchanges became the most frequent aberration. Results of metaphase analysis were similar to those for prematurely condensed chromosomes after 8 h incubation with the exception that a significantly lower frequency of fragments was observed. Symmetrical and asymmetrical interchanges were found at similar frequencies at all doses. No complex exchanges were observed in lymphocyte chromosomes immediately after exposure. They accounted for about 1% of total exchanges in metaphase chromosomes at doses <3 Gy and about 14% at 7 Gy. Incomplete exchanges amounted to approximately 15% of total exchanges at all doses. The kinetics of break rejoining was exponential, and the frequency of exchanges increased with kinetics similar to that observed for the rejoining of the breaks. This increase in the total exchanges as a function of the time between irradiation and fusion was due to a rapid increase in reciprocal interchanges, and a slower increase in complex exchanges; the frequency of incomplete exchanges increased initially, then decreased with prolonged incubation to the level observed

  11. Cytogenetic Low-Dose Hyperradiosensitivity Is Observed in Human Peripheral Blood Lymphocytes

    SciTech Connect

    Seth, Isheeta; Joiner, Michael C.; Tucker, James D.

    2015-01-01

    Purpose: The shape of the ionizing radiation response curve at very low doses has been the subject of considerable debate. Linear-no-threshold (LNT) models are widely used to estimate risks associated with low-dose exposures. However, the low-dose hyperradiosensitivity (HRS) phenomenon, in which cells are especially sensitive at low doses but then show increased radioresistance at higher doses, provides evidence of nonlinearity in the low-dose region. HRS is more prominent in the G2 phase of the cell cycle than in the G0/G1 or S phases. Here we provide the first cytogenetic mechanistic evidence of low-dose HRS in human peripheral blood lymphocytes using structural chromosomal aberrations. Methods and Materials: Human peripheral blood lymphocytes from 2 normal healthy female donors were acutely exposed to cobalt 60 γ rays in either G0 or G2 using closely spaced doses ranging from 0 to 1.5 Gy. Structural chromosomal aberrations were enumerated, and the slopes of the regression lines at low doses (0-0.4 Gy) were compared with doses of 0.5 Gy and above. Results: HRS was clearly evident in both donors for cells irradiated in G2. No HRS was observed in cells irradiated in G0. The radiation effect per unit dose was 2.5- to 3.5-fold higher for doses ≤0.4 Gy than for doses >0.5 Gy. Conclusions: These data provide the first cytogenetic evidence for the existence of HRS in human cells irradiated in G2 and suggest that LNT models may not always be optimal for making radiation risk assessments at low doses.

  12. Gene expression profile of human lymphocytes exposed to (211)At alpha particles.

    PubMed

    Turtoi, A; Brown, I; Schläger, M; Schneeweiss, F H A

    2010-08-01

    In this study, the Whole Human Genome 44K DNA microarray assay was used for the first time to obtain gene expression profiles in human peripheral blood lymphocytes 2 h after exposure (in suspension) to 6.78 MeV mean energy alpha particles from extracellular (211)At. Lymphocytes were exposed to fluences of 0.3-9.6 x 10(6) alpha particles/cm(2) [corresponding to mean absorbed alpha-particle doses (D(alpha)) of 0.05-1.60 Gy] over 30 min. Significantly modulated expression was identified in 338 early-response genes. Up-regulated expression was evident in 183 early-response genes, while the remaining 155 were down-regulated. Over half of the up-regulated genes and 40% of the down-regulated genes had a known biological process related primarily to cell growth and maintenance and cell communication. Genes associated with cell death were found only in the up-regulated genes and those with development only in the down-regulated genes. Eight selected early-response genes that displayed a sustained up- or down-regulation (CD36, HSPA2, MS4A6A, NFIL3, IL1F9, IRX5, RASL11B and SULT1B1) were further validated in alpha-particle-irradiated lymphocytes of two human individuals using the TaqMan(R) RT-qPCR technique. The results confirmed the observed microarray gene expression patterns. The expression modulation profiles of IL1F9, IRX5, RASL11B and SULT1B1 genes demonstrated similar trends in the two individuals studied. However, no significant linear correlation between increasing relative gene expression and the alpha-particle dose was evident. The results suggest the possibility that a panel of genes that react to alpha-particle radiation does exist and that they merit further study in a greater number of individuals to determine their possible value regarding alpha-particle biodosimetry. PMID:20681779

  13. STAT5 phosphorylation in T cell subsets from septic patients in response to recombinant human interleukin-7: a pilot study.

    PubMed

    Demaret, Julie; Dupont, Guillaume; Venet, Fabienne; Friggeri, Arnaud; Lepape, Alain; Rimmelé, Thomas; Morel, Jérôme; Monneret, Guillaume

    2015-04-01

    Septic shock is characterized by lymphocyte alterations associated with increased risk of nosocomial infections and mortality. IL-7, a cytokine required for T cell survival, is thought as a novel therapy for septic patients with severe lymphopenia. We assessed CD4(+) lymphocyte responsiveness to rhIL-7 in septic shock patients ex vivo. Thirteen septic shock patients and 10 controls were included. The MFI of pSTAT5, a key signaling molecule for IL-7, was measured by flow cytometry in CD4(+)FOXP3- (Teffs) and CD4(+)FOXP3(+) (Tregs) lymphocytes after whole-blood incubation with increasing doses of rhIL-7. The basal level of pSTAT5 in nonstimulated T cells was higher in patients. However, the maximal activation level in response to the highest doses of rhIL-7 was similar in both groups. Importantly, low doses of rhIL-7 preferentially activated Teff versus Treg in patients and nonsurvivors tended to present with decreased pSTAT5 expression. This pilot study is the first to highlight, in septic patients, the interest of pSTAT5 measurement in whole blood for the monitoring of rhIL-7 therapy. Such a method could represent an innovative, biologic tool for monitoring leukocyte pharmacological responses to biotherapies in daily clinical practice in other clinical contexts. PMID:25691382

  14. The impact of lymphocyte isolation on induced DNA damage in human blood samples measured by the comet assay.

    PubMed

    Bausinger, Julia; Speit, Günter

    2016-09-01

    The comet assay is frequently used in human biomonitoring for the detection of exposure to genotoxic agents. Peripheral blood samples are most frequently used and tested either as whole blood or after isolation of lymphocytes (i.e. peripheral blood mononuclear cells, PBMC). To investigate a potential impact of lymphocyte isolation on induced DNA damage in human blood samples, we exposed blood ex vivo to mutagens with different modes of genotoxic action. The comet assay was performed either directly with whole blood at the end of the exposure period or with lymphocytes isolated directly after exposure. In addition to the recommended standard protocol for lymphocyte isolation, a shortened protocol was established to optimise the isolation procedure. The results indicate that the effects of induced DNA strand breaks and alkali-labile sites induced by ionising radiation and alkylants, respectively, are significantly reduced in isolated lymphocytes. In contrast, oxidative DNA base damage (induced by potassium bromate) and stable bulky adducts (induced by benzo[a]pyrene-7,8-dihydrodiol-9,10-epoxide; BPDE) seem to be less affected. Our findings suggest that in vivo-induced DNA damage might also be reduced in isolated lymphocytes in comparison with the whole blood depending of the types of DNA damage induced. Because only small genotoxic effects can generally be expected in human biomonitoring studies with the comet assay after occupational and environmental exposure to genotoxic agents, any loss might be relevant and should be avoided. The possibility of such effects and their potential impact on variability of comet assay results in human biomonitoring should be considered when performing or evaluating such kind of studies. PMID:27154923

  15. Setae from larvae of the northern processionary moth (Thaumetopoea pinivora, TP) stimulate proliferation of human blood lymphocytes in vitro.

    PubMed

    Holm, Göran; Andersson, Margareta; Ekberg, Monica; Fagrell, Bengt; Sjöberg, Jan; Bottai, Matteo; Björkholm, Magnus

    2014-01-01

    Larvae of the Northern pine processionary moth (Thaumetopoea pinivora, TP) carry microscopic needles (setae), which by penetrating skin and mucous membranes, may cause inflammatory/immune derived symptoms in man. In the present study the stimulatory effects of setae on human blood lymphocytes in vitro was investigated. Blood mononuclear cells were separated from venous blood or buffy coat of ten healthy individuals, six previously exposed to setae and four with no known exposure. Lymphoproliferation was measured as uptake of 3H-thymidine. Setae were prepared from TP larvae. Setae and saline setae extracts stimulated proliferation of T-lymphocytes in the presence of monocytic cells. Stimulation was pronounced in cells from persons who had been exposed to setae, and weak in cells from non-exposed donors. Chitin also induced lymphocyte proliferation in most donors, but to a lesser extent and independently of donor's previous exposure to setae. In conclusion, setae contain molecules that in the presence of monocytes activate human T-lymphocytes to proliferation. The antigenic nature of stimulatory molecules was supported by the significantly stronger lymphocyte response in persons previously exposed to setae than in non-exposed donors. The nature of such molecules remains to be defined. PMID:25531291

  16. Modulation of radiation-induced and mitomycin C-induced chromosome damage by apigenin in human lymphocytes in vitro

    PubMed Central

    Sharma, Narinder K.

    2013-01-01

    Apigenin (APG), a flavone, is known to exhibit antioxidant, antimutagenic and antitumorigenic activity, both in vivo and in vitro. The aim of this study is to investigate the modulatory effects of APG on human lymphocytes after irradiation with gamma rays (3 Gy) or treatment with the antineoplastic agent, mitomycin C (MMC), in vitro. Cytogenetic biomarkers such as chromosome aberrations (CAs), sister chromatid exchanges (SCEs) and cytochalasin-B blocked micronuclei (CBMN), were studied in blood lymphocytes treated with radiation, or antineoplastic agent (MMC), and APG. Whole blood lymphocytes were cultured in vitro using a standard protocol. No significant differences were found in the frequency of CAs or micronuclei (MN) in human peripheral blood lymphocytes irradiated with gamma rays (3 Gy) and then post-treated with APG. There was an increase in the frequency of SCEs per cell in APG-treated samples compared with the controls. Lymphocytes treated with MMC in the presence of APG exhibited a significant decrease (P < 0.01) in the frequency of SCEs compared with MMC treatment alone. The data for the MN test indicated that APG treatment significantly reduced (P < 0.01) the frequency of MMC-induced MN. PMID:23764456

  17. Setae from Larvae of the Northern Processionary Moth (Thaumetopoea pinivora, TP) Stimulate Proliferation of Human Blood Lymphocytes In Vitro

    PubMed Central

    Holm, Göran; Andersson, Margareta; Ekberg, Monica; Fagrell, Bengt; Sjöberg, Jan; Bottai, Matteo; Björkholm, Magnus

    2014-01-01

    Larvae of the Northern pine processionary moth (Thaumetopoea pinivora, TP) carry microscopic needles (setae), which by penetrating skin and mucous membranes, may cause inflammatory/immune derived symptoms in man. In the present study the stimulatory effects of setae on human blood lymphocytes in vitro was investigated. Blood mononuclear cells were separated from venous blood or buffy coat of ten healthy individuals, six previously exposed to setae and four with no known exposure. Lymphoproliferation was measured as uptake of 3H-thymidine. Setae were prepared from TP larvae. Setae and saline setae extracts stimulated proliferation of T-lymphocytes in the presence of monocytic cells. Stimulation was pronounced in cells from persons who had been exposed to setae, and weak in cells from non-exposed donors. Chitin also induced lymphocyte proliferation in most donors, but to a lesser extent and independently of donor's previous exposure to setae. In conclusion, setae contain molecules that in the presence of monocytes activate human T-lymphocytes to proliferation. The antigenic nature of stimulatory molecules was supported by the significantly stronger lymphocyte response in persons previously exposed to setae than in non-exposed donors. The nature of such molecules remains to be defined. PMID:25531291

  18. Siglec-1-positive plasmacytoid dendritic cells (pDCs) in human peripheral blood: A semi-mature and myeloid-like subset imbalanced during protective and autoimmune responses.

    PubMed

    Wilhelm, Theresa R; Taddeo, Adriano; Winter, Oliver; Schulz, Axel Ronald; Mälzer, Julia-Nora; Domingo, Cristina; Biesen, Robert; Alexander, Tobias; Thiel, Andreas; Radbruch, Andreas; Hiepe, Falk; Gerl, Velia

    2016-02-01

    Plasmacytoid dendritic cells (pDCs) play a central role in the pathogenesis of systemic lupus erythematosus (SLE) as IFN-α producers and promoters of T-cell activation or tolerance. Here, we demonstrated by flow-cytometry and confocal microscopy that Siglec-1, a molecule involved in the regulation of adaptive immunoresponses, is expressed in a subset of semi-mature, myeloid-like pDCs in human blood. These pDCs express lower BDCA-2 and CD123 and higher HLA-DR and CD11c than Siglec-1-negative pDCs and do not produce IFN-α via TLR7/TLR9 engagement. In vitro, Siglec-1 expression was induced in Siglec-1-negative pDCs by influenza virus. Proportions of Siglec-1-positive/Siglec-1-negative pDCs were higher in SLE than in healthy controls and correlated with disease activity. Healthy donors immunized with yellow fever vaccine YFV-17D displayed different kinetics of the two pDC subsets during protective immune response. PDCs can be subdivided into two subsets according to Siglec-1 expression. These subsets may play specific roles in (auto)immune responses. PMID:26674280

  19. [Physicochemical properties of the human lymphocyte mitogenic factor induced by phytohemagglutinin].

    PubMed

    Voĭtenok, N N; Varivotskaia, N V; Murzenok, P P; Potemkina, N D

    1976-08-01

    Some physical and chemical properties of the mitogenic factor (MF) produced in vitro by PHA-stimulated human lymphocytes were investigated. Treatment of the culture medium with proteases proved to decrease the MF activity sharply. The MF was found to have an elution pattern in Sephadex and Bio-gel gels similar to proteins with the molecular weight of 20 000--30 000 daltons (peak--25 000). In experiments with the MF fractionation by disc-electrophoresis on 5% polyacylamide gel the MF formed a few discrete peaks in the fractions corresponding to the mobility of alpha1- and alpha2-globulins and transferrin. In isoelectric focusing the MF formed 3 fractions in the pH range of 4.5--8.3. Functional heterogeneity of the MF is suggested. PMID:1026298

  20. Telomerase-based pharmacologic enhancement of antiviral function of human CD8+ T lymphocytes.

    PubMed

    Fauce, Steven Russell; Jamieson, Beth D; Chin, Allison C; Mitsuyasu, Ronald T; Parish, Stan T; Ng, Hwee L; Kitchen, Christina M Ramirez; Yang, Otto O; Harley, Calvin B; Effros, Rita B

    2008-11-15

    Telomerase reverse transcribes telomere DNA onto the ends of linear chromosomes and retards cellular aging. In contrast to most normal somatic cells, which show little or no telomerase activity, immune cells up-regulate telomerase in concert with activation. Nevertheless, during aging and chronic HIV-1 infection, there are high proportions of dysfunctional CD8(+) CTL with short telomeres, suggesting that telomerase is limiting. The present study shows that exposure of CD8(+) T lymphocytes from HIV-infected human donors to a small molecule telomerase activator (TAT2) modestly retards telomere shortening, increases proliferative potential, and, importantly, enhances cytokine/chemokine production and antiviral activity. The enhanced antiviral effects were abrogated in the presence of a potent and specific telomerase inhibitor, suggesting that TAT2 acts primarily through telomerase activation. Our study is the first to use a pharmacological telomerase-based approach to enhance immune function, thus directly addressing the telomere loss immunopathologic facet of chronic viral infection. PMID:18981163

  1. Calcium transport by ionophorous peptides in dog and human lymphocytes detected by quin-2 fluorescence.

    PubMed

    Deber, C M; Hsu, L C

    1986-01-29

    Synthetic peptides of structure cyclo(Glu(OBz)-Sar-Gly-(N-R)Gly)2 (I), electrogenic Ca2+-selective carriers in phospholipid vesicle membranes, are shown to mediate the uptake of Ca2+ ions into the cytoplasm of dog and human lymphocytes. Ca2+ transport by DECYL-2E (I, R = n-decyl) - monitored by measurements of the fluorescence of an intracellular dye, quin-2 - occurred at a rate comparable to that produced by electroneutral Ca2+ ionophores ionomycin and Br-A23187. Fluorescence quenching experiments using Mn2+ suggested a greater selectivity by DECYL-2E for Ca2+/Mn2+ vs. the other two ionophores. The result that Ca2+ ions can traverse biological membranes bound in a neutral cavity consisting exclusively of peptide carbonyl ligands may imply the functional significance of binding sites of similar structures in membrane transport proteins. PMID:3947349

  2. Radiosensitivity of chromosomes in two successive mitotic cycles of human lymphocytes

    SciTech Connect

    Luchnik, N.V.; Poryadkova, N.A.

    1988-11-01

    A culture of human lymphocytes was irradiated with /gamma/-quanta in a dose of 0.5 Gy with different ratios of cells in first (M1) and second (M2) mitotic cycle and the frequency of aberrations induced at stage G2 was analyzed. With increase in interval of time between the start of culturing and irradiation, total yield of aberrations increased in a regular way. However, if the M1:M2 ratio is considered, then it turns out that in M2 chromosomes are /approximately/1.5 times more sensitive than in M1: within the limits of each cycle, radiosensitivity is constant and does not depend on its duration. It was established in accordance with data of other authors that 5-bromodeoxyuridine (5-BdU) increases radiosensitivity materially.

  3. Portable device for magnetic stimulation: Assessment survival and proliferation in human lymphocytes

    NASA Astrophysics Data System (ADS)

    Pérez, H.; Cordova-Fraga, T.; López-Briones, S.; Martínez-Espinosa, J. C.; Rosas, E. F.; Espinoza, A.; Villagómez-Castro, J. C.; Sosa, M.; Topsu, S.; Bernal-Alvarado, J. J.

    2013-09-01

    A device's instrumentation for magnetic stimulation on human lymphocytes is presented. This is a new procedure to stimulate growing cells with ferrofluid in vortices of magnetic field. The stimulation of magnetic vortices was provided at five different frequencies, from 100 to 2500 Hz and intensities from 1.13 to 4.13 mT. To improve the stimulation effects, a paramagnetic ferrofluid was added on the cell culture medium. The results suggest that the frequency changes and the magnetic field variation produce an important increase in the number of proliferating cells as well as in the cellular viability. This new magnetic stimulation modality could trigger an intracellular mechanism to induce cell proliferation and cellular survival only on mitogen stimulated cells.

  4. Chromosome aberrations induced in human lymphocytes by D-T neutrons

    SciTech Connect

    Lloyd, D.C.; Edwards, A.A.; Prosser, J.S.; Bolton, D.; Sherwin, A.G.

    1984-06-01

    Unstable chromosome aberrations induced by in vitro irradiation with D-T neutrons have been analyzed in human blood lymphocytes. With respect to 250 kVp X rays a maximum limiting RBE at low doses of 4.1 was obtained for dicentric aberrations. Using aberrations as markers in mixed cultures of irradiated and unirradiated cells permits an assessment of interphase death plus mitotic delay. The low-dose RBE for this effect is 2.5. Assuming all unstable aberrations observed at metaphase would lead to cell death by nondisjunction allows an assessment of mitotic death. The low-dose RBE for this effect is 4.5. The data are compared with similar work obtained earlier with /sup 242/Cm ..cap alpha.. particles. The application of the present work to cytogenetic assessment of dose after accidental exposure to D-T neutrons is discussed.

  5. Alteration of membrane transductive mechanisms induced by ethanol in human lymphocyte cultures.

    PubMed

    Fanò, G; Belia, S; Mariggiò, M A; Antonica, A; Agea, E; Spinozzi, F

    1993-03-01

    Ethanol, in millimolar concentrations, significantly modifies different transductive systems in human lymphocyte cultures. In particular, the presence of alcohol in the medium more than doubles the [Ca2+]i (from 70-90 to 200-250 nM), increasing Ca2+ fluxes from outside, and inhibits the active transport carried out by the calcium pump. The Ca2+ release from intracellular stores is not involved because 10 mM EGTA in the medium completely abolished the rise of [Ca2+]i. Since IP3 levels and cAMP concentrations are also involved in ethanol events (although with opposite effects), it seems that the alcohol may have a specific target on cell membranes (G-proteins) which influence many transductive pathways. PMID:8388700

  6. Clastogenic effects of food additive citric acid in human peripheral lymphocytes

    PubMed Central

    Ünal, Fatma; Yüzbaşıoğlu, Deniz; Aksoy, Hüseyin

    2008-01-01

    Clastogenic properties of the food additive citric acid, commonly used as an antioxidant, were analysed in human peripheral blood lymphocytes. Citric acid induced a significant increase of chromosomal aberrations (CAs) at all the concentrations and treatment periods tested. Citric acid significantly decreased mitotic index (MI) at 100 and 200 μg ml−1 concentrations at 24 h, and in all concentrations at 48 h. However, it did not decrease the replication index (RI) significantly. Citric acid also significantly increased sister chromatid exchanges (SCEs) at 100 and 200 μg ml−1 concentrations at 24 h, and in all concentrations at 48 h. This chemical significantly increased the micronuclei frequency (MN) compared to the negative control. It also decreased the cytokinesis-block proliferation index (CBPI), but this result was not statistically significant. PMID:19002851

  7. Comparison of histamine release from human blood monocytes, lymphocytes, adenoidal and skin mast cells.

    PubMed

    Schmutzler, W; Bolsmann, K; Zwadlo-Klarwasser, G

    1995-01-01

    Monocytes and lymphocytes from human blood contain 0.043 +/- 0.007 and 0.053 +/- 0.014 pg histamine/cell, respectively, which can be released by a number of stimulants (A 23187, C5a, substance P, specific allergen). The release process takes 60-120 min to reach its end point, in contrast to tissue mast cells which complete the release within 1-3 min. Both, ketotifen (10(-7) - 10(-5) M) and disodium cromoglycate (10(-5) - 10(-3) M) inhibited histamine release dose dependently up to 40-45%, which might be particularly relevant during the later stages of acute allergic or pseudoallergic reactions. PMID:7542070

  8. Monoclonal antibody (H107) inhibiting IgE binding to Fc epsilon R(+) human lymphocytes.

    PubMed

    Noro, N; Yoshioka, A; Adachi, M; Yasuda, K; Masuda, T; Yodoi, J

    1986-08-15

    A hybridoma-producing monoclonal antibody blocking the binding of human IgE to lymphocytes Fc receptor (Fc epsilon R) was established by the fusion of murine myeloma cells. P3X63.653.Ag8, with BALB/c spleen cells immunized with Fc epsilon R(+) human B lymphoblastoid cell line cells, RPMI1788. A clone of the hybridoma (H107) produced a monoclonal IgG2b antibody that inhibited the rosette formation of Fc epsilon R(+) human B lymphoblastoid cell line cells (RPMI1788, RPMI8866, CESS, Dakiki, and IM9) with fixed ox red blood cells (ORBC) conjugated with human IgE (IgE-ORBC). In contrast, the rosette formation with IgG-conjugated ORBC (IgG-ORBC) on Fc gamma R(+), Fc epsilon R(-) Daudi cells were not affected by the H107 antibodies. A close association of Fc epsilon R and the antigenic determinant recognized by H107 antibody was suggested by the following results. First, the bindings of 125I-labeled IgE (125I-IgE) or 125I-labeled H107 IgG2b antibody (125I-H107) to RPMI8866 cells were inhibited by cold human IgE and H107 IgG2b but not by other classes of human Ig (IgA and IgG), MPC11 IgG2b, or unrelated monoclonal antibodies. Second, H107 antibody reacted with Fc epsilon R(+) B cell lines but not with Fc epsilon R(-) B cell lines as determined by an indirect immunofluorescence. Third, Fc epsilon R(+) cells were depleted by the incubation in the dish coated with H107 antibody or IgE but not in the dish coated with unrelated antibodies. Finally, there was a correlation between the increase of Fc epsilon R(+) cells and that of H107(+) cells in the peripheral blood lymphocytes of the patients with atopic dermatitis. The surface antigens on Fc epsilon R(+) RPMI8866 cells recognized by H107 antibodies had the molecular size of 45,000 as determined by immunoprecipitation and sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis. PMID:2942602

  9. Age-related increases in human lymphocyte DNA damage: is there a role of aerobic fitness?

    PubMed

    Soares, Jorge Pinto; Mota, Maria Paula; Duarte, José Alberto; Collins, Andrew; Gaivão, Isabel

    2013-12-01

    Oxidative stress has been advanced as one of the major causes of damage to DNA and other macromolecules. Although physical exercise may also increase oxidative stress, an important role has been recognized for regular exercise in improving the overall functionality of the body, as indicated by an increase in maximal aerobic uptake ((V)O2max), and in resistance to cell damage. The aims of this study were 1) to evaluate the association between DNA damage in human lymphocytes and age and 2) to evaluate the association between DNA damage in human lymphocytes and ((V)O2max. The sample was composed of 36 healthy and nonsmoking males, aged from 20 to 84 years. ((V)O2max was evaluated through the Bruce protocol with direct measurement of oxygen consumption. The comet assay was used to evaluate the DNA damage, strand breaks and formamidopyrimidine DNA glycosylase (FPG)-sensitive sites. We found a positive correlation of age with DNA strand breaks but not with FPG-sensitive sites. ((V)O2max was significantly inversely related with DNA strand breaks, but this relation disappeared when adjusted for age. A significantly positive relation between ((V)O2max and FPG-sensitive sites was verified. In conclusion, our results showed that younger subjects have lower DNA strand breaks and higher (V)O2max compared with older subjects and FPG-sensitive sites are positively related with ((V)O2max, probably as transient damage due to the acute effects of daily physical activity. PMID:24446564

  10. CD57 in human natural killer cells and T-lymphocytes.

    PubMed

    Kared, Hassen; Martelli, Serena; Ng, Tze Pin; Pender, Sylvia L F; Larbi, Anis

    2016-04-01

    The CD57 antigen (alternatively HNK-1, LEU-7, or L2) is routinely used to identify terminally differentiated 'senescent' cells with reduced proliferative capacity and altered functional properties. In this article, we review current understanding of the attributes of CD57-expressing T-cells and NK cells in both health and disease and discuss how this marker can inform researchers about their likely functions in human blood and tissues in vivo. While CD57 expression on human lymphocytes indicates an inability to proliferate, these cells also display high cytotoxic potential, and CD57(pos) NK cells exhibit both memory-like features and potent effector functions. Accordingly, frequencies of CD57-expressing cells in blood and tissues have been correlated with clinical prognosis in chronic infections or various cancers and with human aging. Functional modulation of senescent CD57(pos) T-cells and mature CD57(pos) NK cells may therefore represent innovative strategies for protection against human immunological aging and/or various chronic diseases. PMID:26850637

  11. Expression profile of Eph receptors and ephrin ligands in healthy human B lymphocytes and chronic lymphocytic leukemia B-cells.

    PubMed

    Alonso-C, Luis M; Trinidad, Eva M A; de Garcillan, Beatriz; Ballesteros, Monica; Castellanos, Milagros; Cotillo, Ignacio; Muñoz, Juan J; Zapata, Agustin G

    2009-03-01

    Increasing information relates some Eph receptors and their ligands, ephrins (EFN), with the immune system. Herein, we found that normal B-cells from peripheral blood (PB) and lymph nodes (LN) showed a differential expression of certain Eph/EFN members, some of them being modulated upon in vitro stimulation including EFNA1, EFNA4, EphB6 and EphA10. In contrast, PB CLL B-cells showed a more heterogeneous Eph/EFN profile than their normal PB B-cell counterparts, expressing Eph/EFN members frequently found within the LN and activated B-cells, specially EFNA4, EphB6 and EphA10. Two of them, EphB6 and EFNA4 were further related with the clinical course of CLL patients. EphB6 expression correlated with a high content of ZAP-70 mRNA and a poor prognosis. High serum levels of a soluble EFNA4 isoform positively correlated with increasing peripheral blood lymphocyte counts and lymphadenopathy. These findings suggest that Eph/EFN might be relevant in normal B-cell biology and could represent new potential prognostic markers and therapeutic targets for CLL. PMID:18819711

  12. Human T lymphocyte migration towards the supernatants of human rhinovirus infected airway epithelial cells: influence of exercise and carbohydrate intake.

    PubMed

    Bishop, Nicolette C; Walker, Gary J; Gleeson, Michael; Wallace, Fiona A; Hewitt, Colin R A

    2009-01-01

    Physical stress induces a marked redistribution of T lymphocytes that may be influenced by carbohydrate (CHO) availability, yet the effect of these on T lymphocyte migration towards infected tissue is unknown. Therefore, the aim of this study was to determine the effect of strenuous exercise and CHO ingestion on subsequent ex vivo lymphocyte migration towards the supernatants of a Human Rhinovirus (HRV)-infected bronchial epithelial cell line. In a randomised, cross-over, double-blind design, 7 trained males ran for 2 h at 60% VO2peak on two occasions with regular ingestion of either a 6.4% w/v glucose and maltodextrin solution (CHO trial) or placebo solution (PLA trial). Plasma glucose concentration was higher on CHO than PLA after exercise (P<0.05). Migration of CD4+ and CD8+ cells and their CD45RA+ and CD45RO+ subpopulations towards supernatants from HRV-infected cells decreased following exercise (main effect for exercise, P<0.01 for CD4+, CD4+CD45RA+ and CD4+CD45RO+; P<0.05 for CD8+, CD8+CD45RA+ and CD8+CD45RO+). Migration of CD4+ cells and CD4+CD45RA+ cells was approximately 35% and approximately 30% higher, respectively, on CHO than PLA at 1 h post-exercise (interaction, P<0.05 for both) and was higher on CHO than PLA for all other subpopulations (P<0.05, main effect for trial). There was little effect of exercise or CHO on migration of these cells towards uninfected (control) cell supernatants or on the proportion of these cells within the peripheral blood mononuclear cell population. The findings of this study suggest that physical stress reduces T cell migration towards HRV-infected cell supernatants and that ingestion of CHO can lessen this effect. PMID:19957874

  13. LARGE DELETIONS ARE TOLERATED AT THE HPRT LOCUS OF IN-VIVO DERIVED HUMAN T-LYMPHOCYTES

    EPA Science Inventory

    A cloning assay was used to recover hprt T-lymphocytes from adult human males. nalysis of crude cellular extracts by polymerase chain reactions (PCRs) demonstrated that 8% (18/218) of the hprt mutations were due to total deletion of the hprt gene. ourteen of the 18 mutants were e...

  14. CYTOGENETIC COMPARISON OF THE RESPONSES OF MOUSE AND HUMAN PERIPHERAL BLOOD LYMPHOCYTES TO 60CO GAMMA RADIATION (JOURNAL VERSION)

    EPA Science Inventory

    Experiments were conducted to compare the chromosome damaging effects of (60)Co gamma radiation on mouse and human peripheral blood lymphocytes (PBLs). Either whole blood or isolated and pelleted mononuclear leucocytes (MNLs) were irradiated with a (60)Co unit to yield exposures ...

  15. MULTIPLEX PCR ANALYSIS OF IN VIVO-ARISING DELETION MUTATIONS IN THE HPRT GENE OF HUMAN T-LYMPHOCYTES

    EPA Science Inventory

    A multiplex polymerase chain reaction (PCR) procedure was adapted for the rapid and efficient evaluation of the hypoxanthine guanine phosphoribosyltransferase (hprt) gene in human T-lymphocytes for deletions. he hprt clonal assay was used to isolate in-vivo-arising hprt-deficient...

  16. DELETION MUTATIONS IN THE HPRT GENE OF T-LYMPHOCYTES AS A BIOMARKER FOR GENOMIC REARRANGEMENTS IMPORTANT IN HUMAN CANCERS

    EPA Science Inventory

    The DNA sequence of 11 in vivo-arising intragenic deletion breaksite junctions occurring in the hypoxanthine guanine phosphoribosyltransferase gene of human T-lymphocytes was determined and deletions ranged in size from 16 bp to 4057 bp. o extensive homology was found at the dele...

  17. Effect of environmental exposures to lead and cadmium on human lymphocytic detoxifying enzymes

    SciTech Connect

    D'Souza, S.J.; Narurkar, L.M.; Narurkar, M.V. )

    1994-09-01

    Lead (Pb) is among the most toxic heavy elements in the atmosphere. Aerosol lead enters the human blood stream by way of the respiratory tract and indirectly, by surface disposition in the alimentary tract followed by adsorption. Lead pollution is also known to occur through its presence in petrol, pain, glazed vessels and solder. Atmospheric lead pollution may be predominantly high around factories manufacturing Pb alloys. Lead toxicity is associated with inhibition of [alpha]-aminolevulinic acid dehydrase (ALAD) activity, rise in the blood porphyrin, inhibition of ATPase in erthrocytes, decreased blood haemoglobin and anemia. Elevated lead concentrations in pregnant women have been shown to cause hypertension and birth defects. Lead is also known to interact with other elements such as Fe, Zn, Ca and Cu in biological systems. Cadmium (Cd) is not essential for human body. It enters the human environment as a contaminant. Human intake of Cd is chiefly through the food chain (about 400-500 [mu]g/wk). Analysis of neuropsy material shows that smokers accumulate much more Cd than nonsmokers. Chronic Cd poisoning produces proteinuere and affects the proximal tubules of kidney, causing the formation of kidney stones. The reported hypertensive effect of Cd in man has been associated with high Cd/Zn ratio in kidney. Studies on air pollution have shown that Cd concentration in air could be positively correlated with heart disease, hypertension and arteriosclerosis. The present investigation was aimed at assessing the usefulness of human lymphocytic detoxicating enzyme activities and their ratios in an assessment of human health-risks during environmental exposures to Pb and Cd. The human subjects investigated comprised those exposed to highly contaminated lead and cadmium areas in the state of Maharashtra, India. 17 refs., 2 figs.

  18. Human tonsil B-cell lymphoma 6 (BCL6)-expressing CD4+ T-cell subset specialized for B-cell help outside germinal centers.

    PubMed

    Bentebibel, Salah-Eddine; Schmitt, Nathalie; Banchereau, Jacques; Ueno, Hideki

    2011-08-16

    T follicular helper (Tfh) cells represent a Th subset engaged in the help of B-cell responses in germinal centers (GCs). Tfh cells abundantly express the transcription repressor B-cell lymphoma 6 (Bcl6), a factor that is necessary and sufficient for their development in vivo. Whether Tfh or Tfh-committed cells are involved in the help of B cells outside GCs remains unclear, particularly in humans. In this study, we identified a previously undefined BCL6-expressing CD4(+) T-cell subset in human tonsils. This subset expressed IL-7 receptor and chemokine receptor 5 (CXCR5) and inducible costimulator (ICOS) at low levels (CXCR5(lo)ICOS(lo)), and it was found exclusively outside GCs. CXCR5(lo)ICOS(lo) CD4(+) T cells secreted larger amounts of IL-21 and IL-10 than CXCR5(hi)ICOS(hi) GC-Tfh cells upon activation, and they induced proliferation and differentiation of naïve B cells into Ig-producing cells more efficiently than GC-Tfh cells. However, this subset lacked the capacity to help GC-B cells because of the induction of apoptosis of GC-B cells through the FAS/FAS-ligand interaction. CXCR5(lo)ICOS(lo) CD4(+) T cells expressed equivalent amounts of BCL6 transcript with CXCR5(hi)ICOS(hi) GC-Tfh cells, but they expressed less Bcl6 protein. Collectively, our study indicates that CXCR5(lo)ICOS(lo) CD4(+) T cells in human tonsils represent Tfh lineage-committed cells that provide help to naïve and memory B cells outside GCs. PMID:21808017

  19. Relation between clinical mature and immature lymphocyte cells in human peripheral blood and their spatial label free scattering patterns

    NASA Astrophysics Data System (ADS)

    Zhang, Lu; Zhao, Xin; Zhang, Zhenxi; Zhao, Hong; Chen, Wei; Yuan, Li

    2016-07-01

    A single living cell's light scattering pattern (LSP) in the horizontal plane, which has been denoted as the cell's "2D fingerprint," may provide a powerful label-free detection tool in clinical applications. We have recently studied the LSP in spatial scattering planes, denoted as the cell's "3D fingerprint," for mature and immature lymphocyte cells in human peripheral blood. The effects of membrane size, morphology, and the existence of the nucleus on the spatial LSP are discussed. In order to distinguish clinical label-free mature and immature lymphocytes, the special features of the spatial LSP are studied by statistical method in both the spatial and frequency domains. Spatial LSP provides rich information on the cell's morphology and contents, which can distinguish mature from immature lymphocyte cells and hence ultimately it may be a useful label-free technique for clinical leukemia diagnosis.

  20. [Structural and Functional Modifications of Human Lymphocytes in Dynamics of UV-Induced Development of Their Apoptosis].

    PubMed

    Nakvasina, M A; Lidokhova, O V; Domanskya, T L; Artyukhov, V G; Ryazantsev, S V

    2015-01-01

    The flow-cytometric analysis of condition of human peripheral blood lymphocytes in dynamics of development of the apoptosis induced by exposure to UV-light (240-390 nanometers) at a dose of 1510 J/m2 was carried out. Superficial architectonics and changes of the level of a membrane potential and functional activity of succinatedehydrogenase of mitochondrions are studied. The contribution of extracellular calcium to the processes of lymphocyte cellular death is revealed. The ideas about dynamics of the intracellular events leading to the death of photomodified lymphocytes 1-6 h after their UV-radiation exposure are specified and added on the basis of the analysis of our new and already published results. PMID:26964347

  1. Relation between clinical mature and immature lymphocyte cells in human peripheral blood and their spatial label free scattering patterns.

    PubMed

    Zhang, Lu; Zhao, Xin; Zhang, Zhenxi; Zhao, Hong; Chen, Wei; Yuan, Li

    2016-07-01

    A single living cell's light scattering pattern (LSP) in the horizontal plane, which has been denoted as the cell's "2D fingerprint," may provide a powerful label-free detection tool in clinical applications. We have recently studied the LSP in spatial scattering planes, denoted as the cell's "3D fingerprint," for mature and immature lymphocyte cells in human peripheral blood. The effects of membrane size, morphology, and the existence of the nucleus on the spatial LSP are discussed. In order to distinguish clinical label-free mature and immature lymphocytes, the special features of the spatial LSP are studied by statistical method in both the spatial and frequency domains. Spatial LSP provides rich information on the cell's morphology and contents, which can distinguish mature from immature lymphocyte cells and hence ultimately it may be a useful label-free technique for clinical leukemia diagnosis. PMID:27475572

  2. Immunoregulatory effects on T lymphocytes by human mesenchymal stromal cells isolated from bone marrow, amniotic fluid, and placenta.

    PubMed

    Mareschi, Katia; Castiglia, Sara; Sanavio, Fiorella; Rustichelli, Deborah; Muraro, Michela; Defedele, Davide; Bergallo, Massimiliano; Fagioli, Franca

    2016-02-01

    Mesenchymal stromal cells (MSCs) are a promising tool in cell therapies because of their multipotent, bystander, and immunomodulatory properties. Although bone marrow represents the main source of MSCs, there remains a need to identify a stem cell source that is safe and easily accessible and yields large numbers of cells without provoking debates over ethics. In this study, MSCs isolated from amniotic fluid and placenta were compared with bone marrow MSCs. Their immunomodulatory properties were studied in total activated T cells (peripheral blood mononuclear cells) stimulated with phytohemagglutinin (PHA-PBMCs). In particular, an in vitro co-culture system was established to study: (i) the effect on T-lymphocyte proliferation; (ii) the presence of T regulatory lymphocytes (Treg); (iii) the immunophenotype of various T subsets (Th1 and Th2 naïve, memory, effector lymphocytes); (iv) cytokine release and master gene expression to verify Th1, Th2, and Th17 polarization; and (v) IDO production. Under all co-culture conditions with PHA-PBMCs and MSCs (independently of tissue origin), data revealed: (i) T proliferation inhibition; (ii) increase in naïve T and decrease in memory T cells; (iii) increase in T regulatory lymphocytes; (iv) strong Th2 polarization associated with increased interleukin-10 and interleukin-4 levels, Th1 inhibition (significant decreases in interleukin-2, tumor necrosis factor-α, interferon-γ, and interleukin-12) and Th17 induction (production of high concentrations of interleukins-6 and -17); (v) indoleamine-2,3-dioxygenase mRNA induction in MSCs co-cultured with PHA-PBMCs. AF-MSCs had a more potent immunomodulatory effect on T cells than BM-MSCs, only slightly higher than that of placenta MSCs. This study indicates that MSCs isolated from fetal tissues may be considered a good alternative to BM-MSCs for clinical applications. PMID:26577566

  3. Inhibition of human peripheral blood lymphocyte function by protoporphyrin and longwave ultraviolet light

    SciTech Connect

    Barrett, K.E.; Yen, A.; Montisano, D.; Gigli, I.; Bigby, T.D.

    1994-10-01

    Modulation of immunologic effector cells by exogenous photoactive substances has been advanced as an underlying mechanism for the efficacy of various photochemotherapeutic regimens. It is also possible that endogenous photosensitizers, such as protoporphyrin, could similarly modify the function of immune cell types. The authors examined the effects of protoporphyrin plus longwave UV light on the ability of human PBL to proliferate in response to mitogens. Noncytotoxic dosages of protoporphyrin plus UV light suppressed PHA-stimulated proliferation of both PBMC and enriched T cells. CD8{sup +} cells were more sensitive to this inhibitory effect than CD4{sup +} cells. The inhibitory effect was also observed when proliferation was induced by the combination of a phorbol ester and ionomycin. Inhibition of PBMC proliferation was associated with inhibition of IL-2 secretion but proliferation was not restored with exogenous IL-2. Instead, the effect of protoporphyrin plus UV light may be on IL-2R. Cells treated with protoporphyrin and UV light did not display the increase in CD25 and {beta}-chain of the IL-2R induced by PHA in control cells. In contrast to the effects of protoporphyrin and UV light on IL-2 and IL-2R {alpha}-chain protein expression, the accumulation of mRNA for these proteins induced by PHA was unaffected. None of the effects of protoporphyrin plus UV light on lymphocytes were observed in control experiments where cells were treated with either protoporphyrin or UV light alone. They conclude that biologically relevant dosages of protoporphyrin and UV light modify the function of circulating lymphocytes. 26 refs., 8 figs., 1 tab.

  4. Glucocorticoids increase the synthesis of immunoglobulin E by interleukin 4-stimulated human lymphocytes.

    PubMed Central

    Wu, C Y; Sarfati, M; Heusser, C; Fournier, S; Rubio-Trujillo, M; Peleman, R; Delespesse, G

    1991-01-01

    This study indicates that hydrocortisone (HC) markedly increases the synthesis of immunoglobulin E (IgE) by interleukin 4 (IL-4)-stimulated human lymphocytes. The effect is glucocorticoid specific and is obtained with low concentrations of HC (0.1-10 microM). In both the early and the late phase of the IL-4-induced response HC exerts its effects which are respectively IL-4 dependent and IL-4 independent. The IgE potentiation cannot be explained by the inhibition of interferon-gamma (IFN-gamma) production since it is observed in the absence of endogenous secretion of IFN-gamma. HC inhibits the production of IgE-binding factors (soluble CD23) and the expression of the low-affinity receptor for IgE, also known as the (Fc epsilon RII) CD23 antigen; however, the residual expression of Fc epsilon RII by IL-4- and HC-treated peripheral blood mononuclear cells (PBMCs) is important since the IgE response of these cells is markedly inhibited by anti-CD23 monoclonal antibody. HC acts mainly by amplifying the cellular interactions between monocytes and lymphocytes; indeed, HC has no effect on monocyte-depleted PBMCs, and moreover, monocytes cannot be replaced by soluble factors. Most importantly, T cells are not required for the induction of IgE synthesis by costimulation with IL-4 and HC. However, the IgE response of rigorously T cell-depleted PBMCs may be further increased by the addition of T cells. Further analysis of the permissive effect of HC on the synthesis of IgE by T cell-depleted PBMCs suggests that HC acts in synergy with IL-4 to trigger the activation and the differentiation of B cells into IgE-producing cells. PMID:1825666

  5. Melittin induced cytogenetic damage, oxidative stress and changes in gene expression in human peripheral blood lymphocytes.

    PubMed

    Gajski, Goran; Domijan, Ana-Marija; Žegura, Bojana; Štern, Alja; Gerić, Marko; Novak Jovanović, Ivana; Vrhovac, Ivana; Madunić, Josip; Breljak, Davorka; Filipič, Metka; Garaj-Vrhovac, Vera

    2016-02-01

    Melittin (MEL) is the main constituent and principal toxin of bee venom. It is a small basic peptide, consisting of a known amino acid sequence, with powerful haemolytic activity. Since MEL is a nonspecific cytolytic peptide that attacks lipid membranes thus leading to toxicity, the presumption is that it could have significant therapeutic benefits. The aim was to evaluate the cyto/genotoxic effects of MEL in human peripheral blood lymphocytes (HPBLs) and the molecular mechanisms involved using a multi-biomarker approach. We found that MEL was cytotoxic for HPBLs in a dose- and time-dependent manner. It also induced morphological changes in the cell membrane, granulation and lysis of exposed cells. After treating HPBLs with non-cytotoxic concentrations of MEL, we observed increased DNA damage including oxidative DNA damage as well as increased formation of micronuclei and nuclear buds, and decreased lymphocyte proliferation determined by comet and micronucleus assays. The observed genotoxicity coincided with increased formation of reactive oxygen species, reduction of glutathione level, increased lipid peroxidation and phospholipase C activity, showing the induction of oxidative stress. MEL also modulated the expression of selected genes involved in DNA damage response (TP53, CDKN1A, GADD45α, MDM), oxidative stress (CAT, SOD1, GPX1, GSR and GCLC) and apoptosis (BAX, BCL-2, CAS-3 and CAS-7). Results indicate that MEL is genotoxic to HPBLs and provide evidence that oxidative stress is involved in its DNA damaging effects. MEL toxicity towards normal cells has to be considered if used for potential therapeutic purposes. PMID:26704293

  6. Chromosome aberration yields and apoptosis in human lymphocytes irradiated with Fe-ions of differing LET

    NASA Astrophysics Data System (ADS)

    Lee, R.; Nasonova, E.; Ritter, S.

    In the present paper the relationship between cell cycle delays induced by Fe-ions of differing LET and the aberration yield observable in human lymphocytes at mitosis was examined. Cells of the same donor were irradiated with 990 MeV/n Fe-ions (LET = 155 keV/μm), 200 MeV/n Fe-ions (LET = 440 keV/μm) and X-rays and aberrations were measured in first cycle mitoses harvested at different times after 48 84 h in culture and in prematurely condensed G2-cells (PCCs) collected at 48 h using calyculin A. Analysis of the time-course of chromosomal damage in first cycle metaphases revealed that the aberration frequency was similar after X-ray irradiation, but increased two and seven fold after exposure to 990 and 200 MeV/n Fe-ions, respectively. Consequently, RBEs derived from late sampling times were significantly higher than those obtained at early times. The PCC-data suggest that the delayed entry of heavily damaged cells into mitosis results especially from a prolonged arrest in G2. Preliminary data obtained for 4.1 MeV/n Cr-ions (LET = 3160 keV/μm) revealed, that these delays are even more pronounced for low energy Fe-like particles. Additionally, for the different radiation qualities, BrdU-labeling indices and apoptotic indices were determined at several time-points. Only the exposure to low energy Fe-like particles affected the entry of lymphocytes into S-phase and generated a significant apoptotic response indicating that under this particular exposure condition a large proportion of heavily damaged cells is rapidly eliminated from the cell population. The significance of this observation for the estimation of the health risk associated with space radiation remains to be elucidated.

  7. Human lymphocytes express the transcriptional regulator, Wilms tumor 1: The role of WT1 in mediating nitric oxide-dependent repression of lymphocyte proliferation

    SciTech Connect

    Marcet-Palacios, Marcelo; Davoine, Francis; Adamko, Darryl J.; Moqbel, Redwan; Befus, A. Dean

    2007-11-16

    The inhibitory roles of nitric oxide (NO) in T cell proliferation have been observed and studied extensively over the last two decades. Despite efforts, the fundamental pathway by which NO exerts its inhibitory actions remains to be elucidated although recent evidence suggests that the transcription factor Wilms tumor 1 (WT1) may be important. WT1 has been linked to numerous developmental pathways in particular nephrogenesis. Due to its roles in development and cell proliferation, polymorphisms within the WT1 gene can result in malignancies such as leukemia and Wilms tumor. WT1 functions as a transcriptional regulator and its activity is controlled through phosphorylation by protein kinase A (PKA). PKA-dependent WT1 phosphorylation results in translocation of WT1 from the nucleus to the cytosol, a process that interferes with WT1 transcriptional activities. In the current study we demonstrate that WT1 is expressed in human lymphocytes. Using the proliferative compound PHA we induced T cell proliferation and growth correlated with an increase in the expression of WT1 measured by RT-PCR, flow cytometry and immunoblot. Co-stimulation with the NO donor SNOG at concentrations of 0, 100, 300 and 600 {mu}M reduced in a concentration dependent way the PHA-induced upregulation of WT1 that correlated with a reduction in T cell proliferation. We conclude that WT1 might be an important component of the NO-dependent regulation of T lymphocyte proliferation and potential function.

  8. Results of space experiment program "Interferon". I. Production of interferon in vitro by human lymphocytes aboard space laboratory Solyut-6 ("Interferon I") and influence of space flight on lymphocyte functions of cosmonauts ("Interferon III").

    PubMed

    Tálas, M; Bátkai, L; Stöger, I; Nagy, L; Hiros, L; Konstantinova, I; Rykova, M; Mozgovaya, I; Guseva, O; Kozharinov, V

    1983-01-01

    The results of the biological space experiment "Interferon" performed by two international cosmonaut crews aboard the space laboratory Solyut-6 are reported. Human lymphocytes separated from the blood of healthy donors and placed into "Interferon I" equipment could be kept for 7 days in suspension culture under spaceflight conditions. Interferon production could be induced in human lymphocytes by preparations of different origin, such as virus, synthetic polyribonucleotides, bacterial protein and plant pigment. An increased lymphocyte interferon production was observed in the space laboratory as compared to the ground control. A decrease of induced interferon production and natural killer cell activity was observed in the cosmonauts' lymphocytes on the 1st day on Earth after 7 days spaceflight. PMID:6659855

  9. Effects of ISS equivalent ionizing radiation dose on Human T-lymphocytes

    NASA Astrophysics Data System (ADS)

    Meloni, Maria Antonia; Pani, Giuseppe; Benotmane, Rafi; Mastroleo, Felice; Aboul-El-Ardat, Khalil; Janssen, Ann; Leysen, Liselotte; Vanhavere, Filip; Leys, Natalie; Galleri, Grazia; Pippia, Proto; Baatout, Sarah

    One of the objectives of the current international space programs is to investigate the effects of cosmic environment on Humans. It is known that during a long exposure to the space conditions, including ionizing radiations and microgravity, the immune system of the astronauts is impaired. In past years several experiments were performed to identify responsible factors of in vitro mitogenic activation process in human T-lymphocytes under simulated microgravity effect and during dedicated space missions. It come out that the lack of immune response in microgravity occurs at the cellular and molecular level. In order to evaluate effects on pure primary T-lymphocytes from peripheral blood exposed to International Space Station (ISS)-like ionizing radiation, we applied a mixture of Cesium-137, as representative of low energy particles, and Californium-252, as representative of hight energy particles, with rate similar to those monitored inside the ISS during previous space mission (Goossens et all. 2006). This facility is available at SCK•CEN (Belgium) (Mastroleo et al., 2009). Although the dose received by the cells was relatively low, flow cytometry analysis 24 hours after irradiation showed a decrease in cell viability coupled with the increase of the caspase-3 activity. However, Bcl-2 activity did not seem to be affected by the radiation. Furthermore, activation of cells induced an increase of the cell size and alteration of cellular morphology. Cell cycle as well as 8-oxo-G were also modified upon radiation and activation. Gene expression analysis shows a modulation of genes rather as a consequence of exposure than with the activation status. 330 genes have been identified to be significantly modulated in function of the time and have been grouped in four different cluster representing significant expression profiles. Preliminary functional analysis shows mainly genes involved in the immune response and inflammatory diseases as well as oxidative stress and

  10. Cytogenetic effects of space-relevant hze-particles in human blood lymphocytes

    NASA Astrophysics Data System (ADS)

    Lee, R.; Nasonova, E.; Ritter, S.

    The analysis of aberrations in human lymphocytes collected 48 h after exposure is used since the 1960s to estimate the radiation risk. However, evidence is increasing that this protocol is not reliable in the case of high LET exposure, because particle induced cell cycle delays influence the aberration yield. To contribute to this issue lymphocytes obtained from a healthy donor were irradiated with Fe-ions (200 MeV/n, 440 keV/μ m), iron-like particles (˜ 4 MeV/n Ni- and Cr-ions, ˜ 4000 keV/μ m) and X-rays. Directly after irradiation PHA and BrdU was added to the cell culture medium. Aberrations were measured in first mitoses collected at 48, 60 and 72 h post-irradiation following colcemid treatment and in prematurely condensed G2-cells (PCCs) at 48 h using calyculin A. Samples were stained with the FPG-technique to allow cell cycle discrimination. Additionally, the mitotic index, the BrdU-labelling index and the number of apoptotic cells were determined at several time-points. Analysis of the BrdU-labelling indices and the mitotic indices revealed a dose- and LET-dependent delay in the cell cycle progression. Cells that reached the first mitosis 48 h after high LET exposure carried only a few aberrations. However, cells that entered the first mitosis 60 to 72 h after high LET exposure carried at least five times more aberrations than those collected at 48 h. The analysis of chromosomal damage in G2-PCCs showed that the delayed entry of severely damaged cells into mitosis results from a prolonged arrest in G2. Conversely, after X-ray exposure a stable aberration-yield was observed in lymphocytes collected at different time-points post-irradiation and the number of aberrations measured in G2-PCCs was only slightly higher than in metaphase cells. Furthermore, only in samples exposed to stopping heavy charged particles a high frequency of apoptotic cells was detected indicating that under this exposure conditions a large proportion of heavily damaged cells is

  11. Erythropoietin receptor is expressed on human peripheral blood T and B lymphocytes and monocytes and is modulated by recombinant human erythropoietin treatment.

    PubMed

    Lisowska, Katarzyna A; Debska-Slizień, Alicja; Bryl, Ewa; Rutkowski, Bolesław; Witkowski, Jacek M

    2010-08-01

    Erythropoietin receptor (EPO-R) appears on the cell surface in the early stages of erythropoiesis. It has also been found on endothelial cells and polymorphonuclear leukocytes, suggesting erythropoietin (EPO) role beyond erythropoiesis itself. Earlier reports have shown that treatment with recombinant human erythropoietin (rhEPO) in chronic renal failure (CRF) patients improves interleukin-2 production and restores the T lymphocyte function. We decided to investigate possible expression of EPO-R on circulating peripheral blood lymphocytes and monocytes of CRF patients in order to assess the possibility of rhEPO direct action on these cells. Flow cytometry was used for detection and quantification of EPO-R, and reverse transcription polymerase chain reaction for detection of the EPO receptor mRNA. Our results show for the first time the existence of EPO-R on cell surface of human T and B lymphocytes and monocytes as well as at the transcriptional activity of the EPO-R gene in these cells, both in healthy and CRF individuals. We have also found significant differences between the numbers of EPO-R molecules on T and B lymphocytes of CRF patients not treated and treated with rhEPO and healthy control. Discovery of EPO-R expression on human lymphocytes suggests that EPO is probably able to directly modulate some signaling pathways important for these cells. PMID:20528849

  12. Human autoimmunity after lymphocyte depletion is caused by homeostatic T-cell proliferation

    PubMed Central

    Jones, Joanne L.; Thompson, Sara A. J.; Loh, Priscilla; Davies, Jessica L.; Tuohy, Orla C.; Curry, Allison J.; Azzopardi, Laura; Hill-Cawthorne, Grant; Fahey, Michael T.; Compston, Alastair; Coles, Alasdair J.

    2013-01-01

    The association between lymphopenia and autoimmunity is recognized, but the underlying mechanisms are poorly understood and have not been studied systematically in humans. People with multiple sclerosis treated with the lymphocyte-depleting monoclonal antibody alemtuzumab offer a unique opportunity to study this phenomenon; one in three people develops clinical autoimmunity, and one in three people develops asymptomatic autoantibodies after treatment. Here, we show that T-cell recovery after alemtuzumab is driven by homeostatic proliferation, leading to the generation of chronically activated (CD28−CD57+), highly proliferative (Ki67+), oligoclonal, memory-like CD4 and CD8 T cells (CCR7−CD45RA− or CCR7−CD45RA+) capable of producing proinflammatory cytokines. Individuals who develop autoimmunity after treatment are no more lymphopenic than their nonautoimmune counterparts, but they show reduced thymopoiesis and generate a more restricted T-cell repertoire. Taken together, these findings demonstrate that homeostatic proliferation drives lymphopenia-associated autoimmunity in humans. PMID:24282306

  13. Studies on the cytotoxicity of diamond nanoparticles against human cancer cells and lymphocytes.

    PubMed

    Adach, Kinga; Fijalkowski, Mateusz; Gajek, Gabriela; Skolimowski, Janusz; Kontek, Renata; Blaszczyk, Alina

    2016-07-25

    Detonation nanodiamonds (DND) are a widely studied group of carbon nanomaterials. They have the ability to adsorb a variety of biomolecules and drugs onto their surfaces, and additionally their surfaces may be subjected to chemical functionalization by covalent bonds. We present a procedure for the purification and surface oxidation of diamond nanoparticles, which were then tested by spectroscopic analysis such as ATR-FTIR, Raman spectroscopy, and thermogravimetric analysis. We also examined the zeta potential of the tested material. Analysis of the cytotoxic effect of nanodiamonds against normal lymphocytes derived from human peripheral blood, the non-small cell lung cancer cell line (A549) and the human colorectal adenocarcinoma cell line (HT29) was performed using MTT colorimetric assay. Evaluation of cell viability was performed after 1-h and 24-h treatment with the tested nanoparticles applied at concentrations ranging from 1 μg/ml to 100 μg/ml. We found that the survival of the examined cells was strongly associated with the presence of serum proteins in the growth medium. The incubation of cells with nanodiamonds in the presence of serum did not exert a significant effect on cell survival, while the cell treatment in a serum-free medium resulted in a decrease in cell survival compared to the negative control. The role of purification and functionalization of nanodiamonds on their cytotoxicity was also demonstrated. PMID:27270448

  14. Chaga mushroom extract inhibits oxidative DNA damage in human lymphocytes as assessed by comet assay.

    PubMed

    Park, Yoo Kyoung; Lee, Hyang Burm; Jeon, Eun-Jae; Jung, Hack Sung; Kang, Myung-Hee

    2004-01-01

    The Chaga mushroom (Inonotus obliquus) is claimed to have beneficial properties for human health, such as anti-bacterial, anti-allergic, anti-inflammatory and antioxidant activities. The antioxidant effects of the mushroom may be partly explained by protection of cell components against free radicals. We evaluated the effect of aqueous Chaga mushroom extracts for their potential for protecting against oxidative damage to DNA in human lymphocytes. Cells were pretreated with various concentrations (10, 50, 100 and 500 microg/mL) of the extract for 1 h at 37 degrees C. Cells were then treated with 100 microM of H2O2 for 5 min as an oxidative stress. Evaluation of oxidative damage was performed using single-cell gel electrophoresis for DNA fragmentation (Comet assay). Using image analysis, the degree of DNA damage was evaluated as the DNA tail moment. Cells pretreated with Chaga extract showed over 40% reduction in DNA fragmentation compared with the positive control (100 micromol H2O2 treatment). Thus, Chaga mushroom treatment affords cellular protection against endogenous DNA damage produced by H2O2. PMID:15630179

  15. Protective Effect of Prolactin against Methylmercury-Induced Mutagenicity and Cytotoxicity on Human Lymphocytes

    PubMed Central

    Silva-Pereira, Liz Carmem; da Rocha, Carlos Alberto Machado; Cunha, Luiz Raimundo Campos da Silva e; da Costa, Edmar Tavares; Guimarães, Ana Paula Araújo; Pontes, Thais Brilhante; Diniz, Domingos Luiz Wanderley Picanço; Leal, Mariana Ferreira; Moreira-Nunes, Caroline Aquino; Burbano, Rommel Rodríguez

    2014-01-01

    Mercury exhibits cytotoxic and mutagenic properties as a result of its effect on tubulin. This toxicity mechanism is related to the production of free radicals that can cause DNA damage. Methylmercury (MeHg) is one of the most toxic of the mercury compounds. It accumulates in the aquatic food chain, eventually reaching the human diet. Several studies have demonstrated that prolactin (PRL) may be differently affected by inorganic and organic mercury based on interference with various neurotransmitters involved in the regulation of PRL secretion. This study evaluated the cytoprotective effect of PRL on human lymphocytes exposed to MeHg in vitro, including observation of the kinetics of HL-60 cells (an acute myeloid leukemia lineage) treated with MeHg and PRL at different concentrations, with both treatments with the individual compounds and combined treatments. All treatments with MeHg produced a significant increase in the frequency of chromatid gaps, however, no significant difference was observed in the chromosomal breaks with any treatment. A dose-dependent increase in the mitotic index was observed for treatments with PRL, which also acts as a co-mitogenic factor, regulating proliferation by modulating the expression of genes that are essential for cell cycle progression and cytoskeleton organization. These properties contribute to the protective action of PRL against the cytotoxic and mutagenic effects of MeHg. PMID:25247425

  16. Protective effect of prolactin against methylmercury-induced mutagenicity and cytotoxicity on human lymphocytes.

    PubMed

    Silva-Pereira, Liz Carmem; da Rocha, Carlos Alberto Machado; Cunha, Luiz Raimundo Campos da Silva E; da Costa, Edmar Tavares; Guimarães, Ana Paula Araújo; Pontes, Thais Brilhante; Diniz, Domingos Luiz Wanderley Picanço; Leal, Mariana Ferreira; Moreira-Nunes, Caroline Aquino; Burbano, Rommel Rodríguez

    2014-09-01

    Mercury exhibits cytotoxic and mutagenic properties as a result of its effect on tubulin. This toxicity mechanism is related to the production of free radicals that can cause DNA damage. Methylmercury (MeHg) is one of the most toxic of the mercury compounds. It accumulates in the aquatic food chain, eventually reaching the human diet. Several studies have demonstrated that prolactin (PRL) may be differently affected by inorganic and organic mercury based on interference with various neurotransmitters involved in the regulation of PRL secretion. This study evaluated the cytoprotective effect of PRL on human lymphocytes exposed to MeHg in vitro, including observation of the kinetics of HL-60 cells (an acute myeloid leukemia lineage) treated with MeHg and PRL at different concentrations, with both treatments with the individual compounds and combined treatments. All treatments with MeHg produced a significant increase in the frequency of chromatid gaps, however, no significant difference was observed in the chromosomal breaks with any treatment. A dose-dependent increase in the mitotic index was observed for treatments with PRL, which also acts as a co-mitogenic factor, regulating proliferation by modulating the expression of genes that are essential for cell cycle progression and cytoskeleton organization. These properties contribute to the protective action of PRL against the cytotoxic and mutagenic effects of MeHg. PMID:25247425

  17. Mouse and human CD8(+) CD28(low) regulatory T lymphocytes differentiate in the thymus.

    PubMed

    Vuddamalay, Yirajen; Attia, Mehdi; Vicente, Rita; Pomié, Céline; Enault, Geneviève; Leobon, Bertrand; Joffre, Olivier; Romagnoli, Paola; van Meerwijk, Joost P M

    2016-06-01

    Regulatory T (Treg) lymphocytes play a central role in the control of immune responses and so maintain immune tolerance and homeostasis. In mice, expression of the CD8 co-receptor and low levels of the co-stimulatory molecule CD28 characterizes a Treg cell population that exerts potent suppressive function in vitro and efficiently controls experimental immunopathology in vivo. It has remained unclear if CD8(+) CD28(low) Treg cells develop in the thymus or represent a population of chronically activated conventional T cells differentiating into Treg cells in the periphery, as suggested by their CD28(low) phenotype. We demonstrate that functional CD8(+) CD28(low) Treg cells are present in the thymus and that these cells develop locally and are not recirculating from the periphery. Differentiation of CD8(+) CD28(low) Treg cells requires MHC class I expression on radioresistant but not on haematopoietic thymic stromal cells. In contrast to other Treg cells, CD8(+) CD28(low) Treg cells develop simultaneously with CD8(+) CD28(high) conventional T cells. We also identified a novel homologous naive CD8(+) CD28(low) T-cell population with immunosuppressive properties in human blood and thymus. Combined, our data demonstrate that CD8(+) CD28(low) cells can develop in the thymus of mice and suggest that the same is true in humans. PMID:26924728

  18. Identification of aneuploidy-inducing agents using cytokinesis-blocked human lymphocytes and an antikinetochore antibody

    SciTech Connect

    Eastmond, D.A.; Tucker, J.D.

    1989-01-01

    The identification of agents causing aneuploidy in humans, a condition associated with carcinogenesis and birth defects, is currently limited due to the highly skilled and time-consuming nature of cytogenetic analyses. We report the development of a new simple and rapid assay to identify aneuploidy-inducing agents (aneuploidogens). The assay involves the chemical- or radiation-induced formation of micronuclei in cytokinesis-blocked human lymphocytes and the use of an antikinetochore antibody to determine whether the micronuclei contain centromeres--a condition indicating a high potential for aneuploidy. All agents tested produced dose-related increases in the frequency of micronucleated cells. The micronucleated cells induced by the known aneuploidogens--colchicine, vincristine sulfate, and diethylstilbestrol--contained kinetochore-positive micronuclei 92, 87, and 76% of the time, respectively. In contrast, the micronucleated cells induced by the potent clastogens--ionizing radiation and sodium arsenite--contained kinetochore-positive micronuclei only 3 and 19% of the time, respectively. These results indicate that this relatively simple assay can discriminate between aneuploidogens and clastogens and may allow a more rapid identification of environmental and therapeutic agents with aneuploidy-inducing potential.

  19. Early T cell signalling is reversibly altered in PD-1+ T lymphocytes infiltrating human tumors.

    PubMed

    Wang, Shu-Fang; Fouquet, Stéphane; Chapon, Maxime; Salmon, Hélène; Regnier, Fabienne; Labroquère, Karine; Badoual, Cécile; Damotte, Diane; Validire, Pierre; Maubec, Eve; Delongchamps, Nicolas B; Cazes, Aurélie; Gibault, Laure; Garcette, Marylène; Dieu-Nosjean, Marie-Caroline; Zerbib, Marc; Avril, Marie-Françoise; Prévost-Blondel, Armelle; Randriamampita, Clotilde; Trautmann, Alain; Bercovici, Nadège

    2011-01-01

    To improve cancer immunotherapy, a better understanding of the weak efficiency of tumor-infiltrating T lymphocytes (TIL) is necessary. We have analyzed the functional state of human TIL immediately after resection of three types of tumors (NSCLC, melanoma and RCC). Several signalling pathways (calcium, phosphorylation of ERK and Akt) and cytokine secretion are affected to different extents in TIL, and show a partial spontaneous recovery within a few hours in culture. The global result is an anergy that is quite distinct from clonal anergy induced in vitro, and closer to adaptive tolerance in mice. PD-1 (programmed death -1) is systematically expressed by TIL and may contribute to their anergy by its mere expression, and not only when it interacts with its ligands PD-L1 or PD-L2, which are not expressed by every tumor. Indeed, the TCR-induced calcium and ERK responses were reduced in peripheral blood T cells transfected with PD-1. Inhibition by sodium stibogluconate of the SHP-1 and SHP-2 phosphatases that associate with several inhibitory receptors including PD-1, relieves part of the anergy apparent in TIL or in PD-1-transfected T cells. This work highlights some of the molecular modifications contributing to functional defects of human TIL. PMID:21408177

  20. The effect of brevenal on brevetoxin-induced DNA damage in human lymphocytes

    PubMed Central

    Sayer, Andrew; Hu, Qing; Bourdelais, Andrea J.; Baden, Daniel G.; Gibson, James E.

    2008-01-01

    Brevenal is a nontoxic short-chain trans-syn polyether that competes with brevetoxin (PbTx) for the active site on voltage-sensitive sodium channels. The PbTxs are highly potent polyether toxins produced during blooms of several species of marine dinoflagellates, most notably Karenia brevis. Blooms of K. brevis have been associated with massive fish kills, marine mammal poisoning, and are potentially responsible for adverse human health effects such as respiratory irritation and airway constriction in beach-goers. Additionally, the consumption of shellfish contaminated with PbTxs results in neurotoxic shellfish poisoning (NSP). The purpose of the present study was to determine whether PbTx could induce DNA damage in a human cell type, the lymphocyte, and if so, whether the damage could be antagonized or ameliorated by brevenal, a brevetoxin antagonist. The DNA damage may occur through both endogenous and exogenous physiological and pathophysiological processes. Unrepaired or erroneously repaired DNA damage may result in gene mutation, chromosome aberration, and modulation of gene regulation, which have been associated with immunotoxicity and carcinogenesis. A single-cell gel electrophoresis assay, or comet assay, was used to determine and compare DNA damage following various treatments. The data were expressed as tail moments, which is the percentage of DNA in the tail multiplied by the length between the center of the head and center of the tail (in arbitrary units). The negative control tail moment was 29.2 (SE=±0.9), whereas the positive control (hydrogen peroxide) was 72.1 (1.5) and solvent (ethanol) was 24.2 (2.1). The PbTx-2 (from Sigma, St. Louis, MO, USA), 10−8 M was 41.3 (3.6), PbTx-9 (Sigma), 10−8 M was 57.0 (5.3), PbTx-2 (from University of North Carolina at Wilmington, UNCW), 10−8 M was 49.4 (9.9), and PbTx-3 (UNCW), 10−8 M was 64.0 (6.4). 1.0 μg/ml brevenal applied 1 h before the PbTxs protected the lymphocytes from DNA damage; PbTx-2 (Sigma

  1. The effect of brevenal on brevetoxin-induced DNA damage in human lymphocytes.

    PubMed

    Sayer, Andrew; Hu, Qing; Bourdelais, Andrea J; Baden, Daniel G; Gibson, James E

    2005-11-01

    Brevenal is a nontoxic short-chain trans-syn polyether that competes with brevetoxin (PbTx) for the active site on voltage-sensitive sodium channels. The PbTxs are highly potent polyether toxins produced during blooms of several species of marine dinoflagellates, most notably Karenia brevis. Blooms of K. brevis have been associated with massive fish kills, marine mammal poisoning, and are potentially responsible for adverse human health effects such as respiratory irritation and airway constriction in beach-goers. Additionally, the consumption of shellfish contaminated with PbTxs results in neurotoxic shellfish poisoning (NSP). The purpose of the present study was to determine whether PbTx could induce DNA damage in a human cell type, the lymphocyte, and if so, whether the damage could be antagonized or ameliorated by brevenal, a brevetoxin antagonist. The DNA damage may occur through both endogenous and exogenous physiological and pathophysiological processes. Unrepaired or erroneously repaired DNA damage may result in gene mutation, chromosome aberration, and modulation of gene regulation, which have been associated with immunotoxicity and carcinogenesis. A single-cell gel electrophoresis assay, or comet assay, was used to determine and compare DNA damage following various treatments. The data were expressed as tail moments, which is the percentage of DNA in the tail multiplied by the length between the center of the head and center of the tail (in arbitrary units). The negative control tail moment was 29.2 (SE=+/-0.9), whereas the positive control (hydrogen peroxide) was 72.1 (1.5) and solvent (ethanol) was 24.2 (2.1). The PbTx-2 (from Sigma, St. Louis, MO, USA), 10(-8) M was 41.3 (3.6), PbTx-9 (Sigma), 10(-8) M was 57.0 (5.3), PbTx-2 (from University of North Carolina at Wilmington, UNCW), 10(-8) M was 49.4 (9.9), and PbTx-3 (UNCW), 10(-8) M was 64.0 (6.4). 1.0 microg/ml brevenal applied 1 h before the PbTxs protected the lymphocytes from DNA damage; PbTx-2

  2. Development of γδ T cell subset responses in gnotobiotic pigs infected with human rotaviruses and colonized with probiotic lactobacilli

    PubMed Central

    Wen, Ke; Li, Guohua; Zhang, Wei; Azevedo, Marli SP; Saif, Linda J; Liu, Fangning; Bui, Tammy; Yousef, Ahmed; Yuan, Lijuan

    2011-01-01

    γδ T cell responses are induced by various viral and bacterial infections. Different γδ T cells contribute to activation and regulation of the inflammatory response and to epithelial repair. How γδ T cells respond to rotavirus infection and how the colonization of probiotics influences the γδ T cell response were unknown. In this study, we evaluated by multicolor flow cytometry the frequencies and distribution of total γδ T cells and three major subsets (CD2−CD8−, CD2+CD8− and CD2+CD8+) in ileum, spleen and blood of gnotobiotic (Gn) pigs at early (3–5 days) and late phases (28 days) after rotavirus infection. The Gn pigs were inoculated with the virulent human rotavirus Wa strain and colonized with a mixture of two strains of probiotics Lactobacillus acidophilus and Lactobacillus reuteri. In naive pigs, the highest frequency of total γδ T cells was found in blood, followed by spleen and ileum at the early age (8–10 days old) whereas in older pigs (32 days of age) the highest frequency of total γδ T cells was found in ileum and spleen followed by blood. Rotavirus infection significantly increased frequencies of intestinal total γδ T cells and the putatively regulatory CD2+CD8+ γδ T cell subset and decreased frequencies of the putatively proinflammatory CD8− subsets in ileum, spleen and blood at post-infection days (PID) 3 or 5. The three γδ T cell subsets distributed and responded differently after rotavirus infection and/or lactobacilli colonization. The CD2+CD8+ subset contributed the most to the expansion of total γδ T cells after rotavirus infection in ileum because more than 77% of the total γδ T cells there were CD2+CD8+ cells. There was an additive effect between lactobacilli and rotavirus in inducing total γδ T cell expansion in ileum at PID 5. The overall effect of lactobacilli colonization versus rotavirus infection on frequencies of the CD2+CD8+ γδ T cell subset in ileum was similar; however, rotavirus-infected pigs

  3. Exogenous Interleukin-2 Administration Corrects the Cell Cycle Perturbation of Lymphocytes from Human Immunodeficiency Virus-Infected Individuals

    PubMed Central

    Paiardini, Mirko; Galati, Domenico; Cervasi, Barbara; Cannavo, Giuseppe; Galluzzi, Luca; Montroni, Maria; Guetard, Denise; Magnani, Mauro; Piedimonte, Giuseppe; Silvestri, Guido

    2001-01-01

    Human immunodeficiency virus (HIV)-induced immunodeficiency is characterized by progressive loss of CD4+ T cells associated with functional abnormalities of the surviving lymphocytes. Increased susceptibility to apoptosis and loss of proper cell cycle control can be observed in lymphocytes from HIV-infected individuals and may contribute to the lymphocyte dysfunction of AIDS patients. To better understand the relation between T-cell activation, apoptosis, and cell cycle perturbation, we studied the effect of exogenous interleukin-2 (IL-2) administration on the intracellular turnover of phase-dependent proteins. Circulating T cells from HIV-infected patients display a marked discrepancy between a metabolic profile typical of G0 and a pattern of expression of phase-dependent proteins that indicates a more-advanced position within the cell cycle. This discrepancy is enhanced by in vitro activation with ConA and ultimately results in a marked increase of apoptotic events. Conversely, treatment of lymphocytes with IL-2 alone restores the phase-specific pattern of expression of cell cycle-dependent proteins and is associated with low levels of apoptosis. Interestingly, exogenous IL-2 administration normalizes the overall intracellular protein turnover, as measured by protein synthesis, half-life of newly synthesised proteins, and total protein ubiquitination, thus providing a possible explanation for the effect of IL-2 on the intracellular kinetics of cell cycle-dependent proteins. The beneficial effect of IL-2 administration is consistent with the possibility of defective IL-2 function in vivo, which is confirmed by the observation that lymphocytes from HIV-infected patients show abnormal endogenous IL-2 paracrine/autocrine function upon in vitro mitogen stimulation. Overall these results confirm that perturbation of cell cycle control contributes to HIV-related lymphocyte dysfunction and, by showing that IL-2 administration can revert this perturbation, suggest a new

  4. Infection frequency of dendritic cells and CD4+ T lymphocytes in spleens of human immunodeficiency virus-positive patients.

    PubMed Central

    McIlroy, D; Autran, B; Cheynier, R; Wain-Hobson, S; Clauvel, J P; Oksenhendler, E; Debré, P; Hosmalin, A

    1995-01-01

    Dendritic cells (DC) are specialized antigen-presenting leukocytes that are responsible for the activation of naive as well as memory T lymphocytes. If infected by human immunodeficiency virus (HIV), DC may transfer virus to CD4+ lymphocytes. However, the question of whether DC are infected in vivo is controversial. As HIV infection is more active in secondary lymphoid organs than in blood, infection of splenic DC isolated from HIV-seropositive patients was investigated. Splenic DC were first enriched and characterized by flow cytometry from HIV- donors. After direct isolation, they were negative for monocyte and T- and B-lymphocyte markers, negative for CD1a, but positive for major histocompatibility complex class II and CD4. After in vitro maturation, major histocompatibility complex class II expression increased, while CD4 expression was lost. Extensive purification from the spleens of seven HIV+ patients was performed by fluorescence-activated cell sorting. The frequency of cells harboring HIV DNA in purified populations was quantified by limiting-dilution PCR. Directly isolated DC (average, 1/3,000; range, 1/720 to 1/18,000) were in each patient 10 to 100 times less infected than CD4+ T lymphocytes (average, 1/52; range, 1/17 to 1/190). On average, 1/1,450 (1/320 to 1/6,100) unseparated mononuclear splenocytes (containing 5% CD4+ lymphocytes) harbored HIV DNA. In conclusion, in these HIV+ patient spleens, DC seem to be infected, but HIV-DNA positive CD4+ T lymphocytes accounted for the vast majority of infected mononuclear splenocytes. PMID:7609039

  5. The effects of age and latent cytomegalovirus infection on the redeployment of CD8+ T cell subsets in response to acute exercise in humans.

    PubMed

    Spielmann, Guillaume; Bollard, Catherine M; Bigley, Austin B; Hanley, Patrick J; Blaney, James W; LaVoy, Emily C P; Pircher, Hanspeter; Simpson, Richard J

    2014-07-01

    Dynamic exercise evokes a rapid redeployment of cytotoxic T cell subsets with high expression of β2 adrenergic receptors, presumably to enhance immunosurveillance during acute stress. As this response is affected by age and infection history, this study examined latent CMV infection as a potential confounder to age-related differences in blood CD8+ T-cell responses to exercise. Healthy young (n=16) and older (n=16) humans counterbalanced by CMV IgG serostatus (positive or negative) exercised for 30-min at ∼80% peak cycling power. Those with CMV redeployed ∼2-times more CD8+ T-cells and ∼6-times more KLRG1+/CD28- and CD45RA+/CCR7- CD8+ subsets than non-infected exercisers. Seronegative older exercisers had an impaired redeployment of total CD8+ T-cells, CD45RA+/CCR7+ and KLRG1-/CD28+ CD8+ subsets compared to young. Redeployed CD8+ T-cell numbers were similar between infected young and old. CMVpp65 specific CD8+ cells in HLA/A2(∗) subjects increased ∼2.7-fold after exercise, a response that was driven by the KLRG1+/CD28-/CD8+ subset. Stimulating PBMCs before and after exercise with CMVpp65 and CMV IE-1 antigens and overlapping peptide pools revealed a 2.1 and 4.4-fold increases in CMVpp65 and CMV IE-1 IFN-γ secreting cells respectively. The breadth of the T cell response was maintained after exercise with the magnitude of the response being amplified across the entire epitope repertoire. To conclude, latent CMV infection overrides age-related impairments in CD8+ T-cell redeployment with exercise. We also show for the first time that many T-cells redeployed with exercise are specific to CMVpp65 and CMV IE-1 antigens, have broad epitope specificity, and are mostly of a high-differentiated effector memory phenotype. PMID:23684819

  6. Allergen-Specific Immunotherapy Alters the Frequency, as well as the FcR and CLR Expression Profiles of Human Dendritic Cell Subsets

    PubMed Central

    Lundberg, Kristina; Rydnert, Frida; Broos, Sissela; Andersson, Morgan; Greiff, Lennart; Lindstedt, Malin

    2016-01-01

    Allergen-specific immunotherapy (AIT) induces tolerance and shifts the Th2 response towards a regulatory T-cell profile. The underlying mechanisms are not fully understood, but dendritic cells (DC) play a vital role as key regulators of T-cell responses. DCs interact with allergens via Fc receptors (FcRs) and via certain C-type lectin receptors (CLRs), including CD209/DC-SIGN, CD206/MR and Dectin-2/CLEC6A. In this study, the effect of AIT on the frequencies as well as the FcR and CLR expression profiles of human DC subsets was assessed. PBMC was isolated from peripheral blood from seven allergic donors before and after 8 weeks and 1 year of subcutaneous AIT, as well as from six non-allergic individuals. Cells were stained with antibodies against DC subset-specific markers and a panel of FcRs and CLRs and analyzed by flow cytometry. After 1 year of AIT, the frequency of CD123+ DCs was increased and a larger proportion expressed FcεRI. Furthermore, the expression of CD206 and Dectin-2 was reduced on CD141+ DCs after 1 year of treatment and CD206 as well as Dectin-1 was additionally down regulated in CD1c+ DCs. Interestingly, levels of DNGR1/CLEC9A on CD141+ DCs were increased by AIT, reaching levels similar to cells isolated from non-allergic controls. The modifications in phenotype and occurrence of specific DC subsets observed during AIT suggest an altered capacity of DC subsets to interact with allergens, which can be part of the mechanisms by which AIT induces allergen tolerance. PMID:26863539

  7. Human gene transfer: Characterization of human tumor-infiltrating lymphocytes as vehicles for retroviral-mediated gene transfer in man

    SciTech Connect

    Kasid, A.; Morecki, S.; Aebersold, P.; Cornetta, K.; Culver, K.; Freeman, S.; Director, E.; Lotze, M.T.; Blaese, R.M.; Anderson, W.F.; Rosenberg, S.A. )

    1990-01-01

    Tumor-infiltrating lymphocytes (TILs) are cells generated from tumor suspensions cultured in interleukin 2 that can mediate cancer regression when adoptively transferred into mice or humans. Since TILs proliferate rapidly in vitro, recirculate, and preferentially localize at the tumor site in vivo, they provide an attractive model for delivery of exogenous genetic material into man. To determine whether efficient gene transfer into TILs is feasible. The authors transduced human TILs with the bacterial gene for neomycin-resistance (Neo{sup R}) using the retroviral vector N2. The transduced TIL populations were stable and polyclonal with respect to the intact Neo{sup R} gene integration and expressed high levels of neomycin phosphotransferase activity. The Neo{sup R} gene insertion did not alter the in vitro growth pattern and interleukin 2 dependence of the transduced TILs. Analyses of T-cell receptor gene rearrangement for {beta}- and {gamma}-chain genes revealed the oligoclonal nature of the TIL populations with no major change in the DNA rearrangement patterns or the levels of mRNA expression of the {beta} and {gamma} chains following transduction and selection of TILs in the neomycin analog G418. Human TILs expressed mRNA for tumor necrosis factors ({alpha} and {beta}) and interleukin 2 receptor P55. This pattern of cytokine-mRNA expression was not significantly altered following the transduction of TILs. The studies demonstrate the feasibility of TILs as suitable cellular vehicles for the introduction of therapeutic genes into patients receiving autologous TILs.

  8. Differential Activation of Human Monocytes and Lymphocytes by Distinct Strains of Trypanosoma cruzi

    PubMed Central

    Magalhães, Luísa M. D.; Viana, Agostinho; Chiari, Egler; Galvão, Lúcia M. C.; Gollob, Kenneth J.; Dutra, Walderez O.

    2015-01-01

    Background Trypanosoma cruzi strains are currently classified into six discrete typing units (DTUs) named TcI to VI. It is known that these DTUs have different geographical distribution, as well as biological features. TcI and TcII are major DTUs found in patients from northern and southern Latin America, respectively. Our hypothesis is that upon infection of human peripheral blood cells, Y strain (Tc II) and Col cl1.7 (Tc I), cause distinct immunological changes, which might influence the clinical course of Chagas disease. Methodology/Principal Findings We evaluated the infectivity of CFSE-stained trypomastigotes of Col cl1.7 and Y strain in human monocytes for 15 and 72 hours, and determined the immunological profile of lymphocytes and monocytes exposed to the different isolates using multiparameter flow cytometry. Our results showed a similar percentage and intensity of monocyte infection by Y and Col cl1.7. We also observed an increased expression of CD80 and CD86 by monocytes infected with Col cl1.7, but not Y strain. IL-10 was significantly higher in monocytes infected with Col cl1.7, as compared to Y strain. Moreover, infection with Col cl1.7, but not Y strain, led to an increased expression of IL-17 by CD8+ T cells. On the other hand, we observed a positive correlation between the expression of TNF-alpha and granzyme A only after infection with Y strain. Conclusion/Significance Our study shows that while Col cl1.7 induces higher monocyte activation and, at the same time, production of IL-10, infection with Y strain leads to a lower monocyte activation but higher inflammatory profile. These results show that TcI and TcII have a distinct immunological impact on human cells during early infection, which might influence disease progression. PMID:26147698

  9. Leukocyte Immunoglobulin-Like Receptor 1-Expressing Human Natural Killer Cell Subsets Differentially Recognize Isolates of Human Cytomegalovirus through the Viral Major Histocompatibility Complex Class I Homolog UL18

    PubMed Central

    Chen, Kevin C.; Banat, Jareer J.

    2016-01-01

    ABSTRACT Immune responses of natural killer (NK) cell are controlled by the balance between activating and inhibitory receptors, but the expression of these receptors varies between cells within an individual. Although NK cells are a component of the innate immune system, particular NK cell subsets expressing Ly49H are positively selected and increase in frequency in response to cytomegalovirus infection in mice. Recent evidence suggests that in humans certain NK subsets also have an increased frequency in the blood of human cytomegalovirus (HCMV)-infected individuals. However, whether these subsets differ in their capacity of direct control of HCMV-infected cells remains unclear. In this study, we developed a novel in vitro assay to assess whether human NK cell subsets have differential abilities to inhibit HCMV growth and dissemination. NK cells expressing or lacking NKG2C did not display any differences in controlling viral dissemination. However, when in vitro-expanded NK cells were used, cells expressing or lacking the inhibitory receptor leukocyte immunoglobulin-like receptor 1 (LIR1) were differentially able to control dissemination. Surprisingly, the ability of LIR1+ NK cells to control virus spread differed between HCMV viral strains, and this phenomenon was dependent on amino acid sequences within the viral ligand UL18. Together, the results here outline an in vitro technique to compare the long-term immune responses of different human NK cell subsets and suggest, for the first time, that phenotypically defined human NK cell subsets may differentially recognize HCMV infections. IMPORTANCE HCMV infection is ubiquitous in most populations; it is not cleared by the host after primary infection but persists for life. The innate and adaptive immune systems control the spread of virus, for which natural killer (NK) cells play a pivotal role. NK cells can respond to HCMV infection by rapid, short-term, nonspecific innate responses, but evidence from murine

  10. Characterization of a cellular inhibitory activity affecting the human autologous mixed lymphocyte reaction

    SciTech Connect

    Haynes, M.K.

    1987-01-01

    The human autologous mixed lymphocyte reaction, peripheral blood T cells are stimulated to proliferate when co-cultured with irradiated, autologous, non-T cells was studied. ({sup 3}H)-thymidine uptake was maximal on day 7 and observable on day 9. Non-adherent and adherent fractions of non-T population and lectin-activated T cell blasts contributed. Unseparated non-T stimulator cells were used. Autologous T cells, when added as irradiated third party cells, reduced AMLR proliferative responses. Depletion of Fc IgG receptor-bearing cells (T{gamma}) from T cell population by (a) IgG-sensitized ox erythrocyte rosetting (EA-rosettes) or (b) adsorption of the T{gamma} cells onto a Sepharose 6MB {approximately} BSA/anti-BSA immune complex column resulted in enhanced AMLR proliferation not attributed solely to an increase in CD8+ AMLR responder cell phenotype. Isolated T{gamma} populations exhibited a reversed helper/suppressor phenotype containing HLA DR+ and CD8+ cells. Dual labeling studies demonstrated 7-24% of CD8+ cells expressed DR antigens. After separation, AMLR proliferative capacity was localized in T-depleted population.

  11. Synthesis of novel tetrahydrobenzazepine derivatives and their cytoprotective effect on human lymphocytes.

    PubMed

    Simic, Milena R; Stankovic, Miroslava; Mandic, Boris M; Tesevic, Vele V; Savic, Vladimir M

    2015-02-01

    Cytoprotective compounds such as amifostine play an important role in chemo- and radiotherapy due to their ability to reduce the side effects of these treatments. Our work was initiated with the intention to design, synthesise and test a new class of heterocyclic compounds that would have an antioxidative profile with the potential to be further developed as cytoprotective agents. The design was based on the privileged tetrahydrobenzazepine scaffold found in many natural products with a wide range of biological properties. This structure was further functionalised with moieties known to possess antioxidative features such as tertiary amine and styrene double bond. A series of eight tetrahydrobenzazepine derivatives of isoquinoline, 3,4-dihydro-β-carboline and pyridine were synthesised employing the Heck reaction as a key transformation. Some of the prepared compounds were tested for their in vitro effects on chromosome aberrations in peripheral human lymphocytes using the cytochalasin-B blocked micronucleus (MN) assay. Three tetrahydrobenzoazepine derivatives showed significant cytoprotective properties, comparable or even better to those of the radioprotective agent amifostine. PMID:25664628

  12. Functional changes in astrocytes by human T-lymphotropic virus type-1 T-lymphocytes.

    PubMed

    Akaoka, H; Szymocha, R; Beurton-Marduel, P; Bernard, A; Belin, M F; Giraudon, P

    2001-10-30

    The human T-lymphotropic virus type-1 (HTLV-1) is the causative agent of a chronic progressive myelopathy (TSP/HAM) in which lesions of the central nervous system (CNS) are associated with infiltration of HTLV-1-infected T-cells. In a model that mimics the interaction between glial and T-cells, we show that transient contact with T-lymphocytes chronically infected with HTLV-1 induce profound metabolic alterations in astrocytes. Within the first week post-contact, an overall activation of astrocyte metabolism was observed as assessed by enhanced uptake of glutamate and glucose, and lactate release. In contrast, longer examination showed a reduced astrocytic accumulation of glutamate. The time course of the change in glutamate uptake was in fact biphasic. Previous observations indicated that HTLV-1 protein Tax-1 was involved in this delayed decrease, via the induction of TNF-alpha. The expression of the glial glutamate transporters, GLAST and GLT-1 decreased in parallel. These decreases in glutamate uptake and transporters' expression were associated with an imbalance in the expression of the catabolic enzymes of glutamate, GS and GDH, presumably due to Tax-1. Given the fact that impairment of glutamate management in astrocytes is able to compromise the functional integrity of neurons and oligodendrocytes, our results altogether give new insights into the physiopathology of TSP/HAM. PMID:11520580

  13. Dose rate effect of pulsed electron beam on micronucleus frequency in human peripheral blood lymphocytes.

    PubMed

    Acharya, Santhosh; Sanjeev, Ganesh; Bhat, Nagesh N; Narayana, Yerol

    2010-03-01

    The micronucleus assay in human peripheral blood lymphocytes is a sensitive indicator of radiation damage and could serve as a biological dosimeter in evaluating suspected overexposure to ionising radiation. Micronucleus (MN) frequency as a measure of chromosomal damage has also extensively been employed to quantify the effects of radiation dose rate on biological systems. Here we studied the effects of 8 MeV pulsed electron beam emitted by Microtron electron accelerator on MN induction at dose rates between 35 Gy min-1 and 352.5 Gy min-1. These dose rates were achieved by varying the pulse repetition rate (PRR). Fricke dosimeter was employed to measure the absorbed dose at different PRR and to ensure uniform dose distribution of the electron beam. To study the dose rate effect, blood samples were irradiated to an absorbed dose of (4.7+/-0.2) Gy at different rates and cytogenetic damage was quantified using the micronucleus assay. The obtained MN frequency showed no dose rate dependence within the studied dose rate range. Our earlier dose effect study using 8 MeV electrons revealed that the response of MN was linear-quadratic. Therefore, in the event of an accident, dose estimation can be made using linear-quadratic dose response parameters, without adding dose rate as a correction factor. PMID:20338871

  14. An infrared spectral signature of human lymphocyte subpopulations from peripheral blood.

    PubMed

    Wald, N; Legat, A; Meyer, C; Speiser, D E; Goormaghtigh, E

    2015-04-01

    Metastatic melanomas are frequently refractory to most adjuvant therapies such as chemotherapies and radiotherapies. Recently, immunotherapies have shown good results in the treatment of some metastatic melanomas. Immune cell infiltration in the tumor has been associated with successful immunotherapy. More generally, tumor infiltrating lymphocytes (TILs) in the primary tumor and in metastases of melanoma patients have been demonstrated to correlate positively with favorable clinical outcomes. Altogether, these findings suggest the importance of being able to identify, quantify and characterize immune infiltration at the tumor site for a better diagnostic and treatment choice. In this paper, we used Fourier Transform Infrared (FTIR) imaging to identify and quantify different subpopulations of T cells: the cytotoxic T cells (CD8+), the helper T cells (CD4+) and the regulatory T cells (T reg). As a proof of concept, we investigated pure populations isolated from human peripheral blood from 6 healthy donors. These subpopulations were isolated from blood samples by magnetic labeling and purities were assessed by Fluorescence Activated Cell Sorting (FACS). The results presented here show that Fourier Transform Infrared (FTIR) imaging followed by supervised Partial Least Square Discriminant Analysis (PLS-DA) allows an accurate identification of CD4+ T cells and CD8+ T cells (>86%). We then developed a PLS regression allowing the quantification of T reg in a different mix of immune cells (e.g. Peripheral Blood Mononuclear Cells (PBMCs)). Altogether, these results demonstrate the sensitivity of infrared imaging to detect the low biological variability observed in T cell subpopulations. PMID:25553786

  15. Characterization of protein kinase C and its isoforms in human T lymphocytes.

    PubMed

    Beyers, A D; Hanekom, C; Rheeder, A; Strachan, A F; Wooten, M W; Nel, A E

    1988-11-15

    Protein kinase C (PKC) regulates numerous T cell functions and is present in abundance in normal human T cells and certain T cell lines. Although crude Triton X-100 soluble material obtained from T cell pellets contains minimal PKC activity, DEAE chromatography revealed that 12 to 37% of cellular PKC was membrane associated, probably due to removal of an inhibitor through column chromatography. As in other tissues, PKC from lymphoid tissue was phospholipid and Ca2+ dependent and diolein reduced the Ca2+ requirements for enzyme activity. Hydroxylapatite chromatography revealed that T cells possess two major peaks of PKC activity. Although, the enzyme in these peaks had similar m.w. and identical iso-electric mobility, the proteins differed with respect to their autophosphorylation sites and immunoreactivity toward an isoform specific antibody. Furthermore, differences in their activities in the presence of phospholipid, diolein, and limiting amounts of Ca2+ imply that these isoforms may be differentially activated. We discuss optimal conditions for activation of PKC and its isoforms for study of T lymphocyte cellular function. PMID:3263426

  16. Apigenin ameliorates gamma radiation-induced cytogenetic alterations in cultured human blood lymphocytes.

    PubMed

    Begum, Naziya; Prasad, N Rajendra; Kanimozhi, G; Hasan, Annie Q

    2012-08-30

    The aim of the present study was to assess the protective effect of apigenin, a dietary flavone, against cytogenetic alterations in human peripheral blood lymphocytes (HPBL) induced by Cobalt-60 radiation (3Gy). Results of MTT [3-(4, 5-dimethyl-2-thiaozolyl)-2,5-diphenyl-2H tetrazolium bromide] assay revealed that 37.2μM of apigenin was found to be non-toxic in HPBL. At this dose (37.2μM) of apigenin, the LD(50) radiation dose of HPBL increased from 2.9Gy to 3.4Gy, which resulted in a DMF of 1.17. Apigenin (37.2μM) treatment 1h before irradiation significantly (p<0.05) reduced DNA damage in irradiated HPBL as measured by comet assay (% tail DNA, tail length, tail moment, and olive tail moment). Moreover, apigenin treatment significantly decreased the frequencies of dicentric (DC), acentric fragments (AF), and acentric rings (AR) in irradiated HPBL. Apigenin pretreatment also reduced the radiation-induced CBMN (cytokinesis blocked micronuclei) anomalies such as micronuclei (MNi), nucleoplasmic bridges (NPB) and nuclear buds (NBUD) in HPBL. These results also showed that there was a significant correlation between NPB and DC frequencies and MNi and AF+AR. Treatment with apigenin alone had no significant effect on DNA damage and chromosomal aberrations in HPBL. Thus, the current studies indicate that apigenin protects HPBL from radiation-induced cytogenetic alterations. PMID:22516036

  17. Immunoresistant human glioma cell clones selected with alloreactive cytotoxic T lymphocytes

    PubMed Central

    Gomez, German G.; Hickey, Michelle J.; Tritz, Richard; Kruse, Carol A.

    2008-01-01

    Summary We previously reported the cellular, functional and cytogenetic characterization of immunoresistant (IR) 13-06-IR29 and 13-06-IR30 human glioma cell clones isolated after immunoselection with alloreactive cytotoxic T lymphocytes (aCTL). Relative to the 13-06-MG parental cells, both clones resisted aCTL lysis at multiple effector to target ratios; the resistant phenotype was maintained for 13-41 cell doublings after cloning and when selective pressure was removed; cross-resistance to other inducers of apoptosis/cell death was also observed (Gomez et al, 2006; Gomez and Kruse, 2007). In this study we further characterize the IR clones for factors that may contribute to the resistance. Data obtained by in-vitro quantitative morphologic and 7-amino actinomycin D flow cytometric assays revealed reduced apoptotic cell death when IR clones were coincubated with aCTL, relative to the parental cells. Since changes in apoptosis were observed, we examined the expression patterns of apoptosis-related genes in several extracts of parental cells and IR clones using pathway-specific cDNA microarray analysis. In general, the apoptotic factors were downregulated in the IR clones. From three separate extracts analyzed separately on microarrays, three factors, ATM, caspases 3 and 8, were statistically downregulated in both IR clones. Immunoblotting of the proteins confirmed the findings. Therefore, a possible mechanism for immunoresistance in gliomas may be achieved by the downregulation of one or more genes in the apoptotic pathway. PMID:19066635

  18. Genomic Instability in Human Lymphocytes from Male Users of Crack Cocaine

    PubMed Central

    de Freitas, Thiago Aley Brites; Palazzo, Roberta Passos; de Andrade, Fabiana Michelsen; Reichert, César Luis; Pechansky, Flávio; Kessler, Félix; de Farias, Caroline Brunetto; de Andrade, Gisele Gomes; Leistner-Segal, Sandra; Maluf, Sharbel Weidner

    2014-01-01

    Recent research suggests that crack cocaine use alters systemic biochemical markers, like oxidative damage and inflammation markers, but very few studies have assessed the potential effects of crack cocaine at the cellular level. We assessed genome instability by means of the comet assay and the cytokinesis-block micronucleus technique in crack cocaine users at the time of admission to a rehabilitation clinic and at two times after the beginning of withdrawal. Thirty one active users of crack cocaine and forty control subjects were evaluated. Comparison between controls and crack cocaine users at the first analysis showed significant differences in the rates of DNA damage (p = 0.037). The frequency of micronuclei (MN) (p < 0.001) and nuclear buds (NBUDs) (p < 0.001) was increased, but not the frequency of nucleoplasmic bridges (NPBs) (p = 0.089). DNA damage decreased only after the end of treatment (p < 0.001). Micronuclei frequency did not decrease after treatment, and nuclear buds increased substantially. The results of this study reveal the genotoxic and mutagenic effects of crack cocaine use in human lymphocytes and pave the way for further research on cellular responses and the possible consequences of DNA damage, such as induction of irreversible neurological disease and cancer. PMID:25264678

  19. Very short-lived and stable mRNAs from resting human lymphocytes.

    PubMed Central

    Berger, S L; Cooper, H L

    1975-01-01

    The kinetics of degradation of newly synthesized cytoplasmic poly(A)-bearing RNA have been examined in resting human lymphocytes. Two classes were identified, a very labile component with a half-life of less than 17 min and a stable component which remains apparently undiminished during 24 hr of observation. Both classes have molecular weights between 2.5 and 3.5 x 10(5) but the stable material has a narrower size distribution and a slightly lower average molecular weight than the short-lived component. The fate of stable RNA synthesized in the resting cell was also examined after growth stimulation with phytohemagglutinin after 2 and 6 hr of treatment. No transfer of stable material into the labile pool could be discerned; the amount of stable material remained constant. The existence of two species of mRNAs with different lifetimes in animal cells provides a potential means for regulation of protein synthesis by controlling the supply of specific messages. Furthermore, such a short-lived mRNA species may explain the observed disparity between the amount of poly(A)-bearing heterogeneous RNA produced in the nucleus and the amount of mature message found in the cytoplasm. PMID:1060069

  20. Large granular lymphocytes are universally increased in human, macaque, and feline lentiviral infection.

    PubMed

    Sprague, Wendy S; Apetrei, Cristian; Avery, Anne C; Peskind, Robert L; Vandewoude, Sue

    2015-10-15

    Large granular lymphocytes (LGLs) have only been anecdotally reported in HIV infection. We previously reported an LGL lymphocytosis in FIV-infected cats associated with a rise in FIV proviral loads and a marked neutropenia that persisted during chronic infection. Extensive immunophenotyping of peripheral blood mononuclear cells in cats chronically infected with FIV were identified LGLs as CD8lo(+)FAS(+); this cell population expanded commensurate with viral load. CD8lo(+)FAS(+) cells expressed similar levels of interferon-γ compared to CD8lo(+)FAS(+) cells from FIV-naive control animals, yet CD3ɛ expression, which was increased on total CD8(+) T cells in FIV-infected cats, was decreased on CD8lo(+)FAS(+) cells. Down-modulation of CD3 expression was reversed after culturing PBMC for 3 days in culture with ConA/IL-2. We identified CD8lo(+)FAS(+) LGLs to be polyclonal T cells lacking CD56 expression. Blood smears from HIV-infected individuals and SIVmac239-infected rhesus macaques revealed increased LGLs compared to HIV/SIV negative counterparts. In humans, there was no correlation with viral load or treatment and in macaques the LGLs arose in acute SIV infection with increases in viremia. This is the first report describing and partially characterizing LGL lymphocytosis in association with lentiviral infections in three different species. PMID:26292765

  1. Effects of anti-human T lymphocyte immune globulins in patients: new or old.

    PubMed

    Wang, Diane C; Wang, Xiangdong; Chen, Chengshui

    2016-09-01

    Multiple studies demonstrated that anti-human T lymphocyte immune globulins (ATG) can decrease the incidence of acute and chronic graft rejection in cell or organ transplants. However, further in-depth study indicates that different subgroups may benefit from either different regimes or alteration of them. Studies among renal transplant patients indicate that low immunological risk patients may not gain the same amount of benefit and thus tilt the risk versus benefit consideration. This may hold true for low immunological risk patients receiving other organ transplants and would be worth further investigation. The recovery time of T cells and natural killer (NK) cells also bears consideration and the impact that it has on the severity and incidence of opportunistic infections closely correlated with the dosage of ATG. The use of lower doses of ATG in combination with other induction medications may offer a solution. The finding that ATG may lose efficacy in cases of multiple transplants or re-transplants in the case of heart transplants may hold true for other transplantations. This may lead to reconsideration of which induction therapies would be most beneficial in the clinical setting. These studies on ATG done on different patient groups will naturally not be applicable to all, but the evidence accrued from them as a whole may offer us new and different perspectives on how to approach and potentially solve the clinical question of how to best reduce the mortality associated with chronic host-versus-graft disease. PMID:27084794

  2. Effects on DNA repair in human lymphocytes exposed to the food dye tartrazine yellow.

    PubMed

    Soares, Bruno Moreira; Araújo, Taíssa Maíra Thomaz; Ramos, Jorge Amando Batista; Pinto, Laine Celestino; Khayat, Bruna Meireles; De Oliveira Bahia, Marcelo; Montenegro, Raquel Carvalho; Burbano, Rommel Mario Rodríguez; Khayat, André Salim

    2015-03-01

    Tartrazine is a food additive that belongs to a class of artificial dyes and contains an azo group. Studies about its genotoxic, cytotoxic and mutagenic effects are controversial and, in some cases, unsatisfactory. This work evaluated the potential in vitro cytotoxicity, genotoxicity and effects on DNA repair of human lymphocytes exposed to the dye. We assessed the cytotoxicity of tartrazine by 3-(4,5-Dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide test and the response of DNA repair through comet assay (alkaline version). We used different concentrations of the dye, ranging from 0.25-64.0 mM. The results demonstrated that tartrazine has no cytotoxic effects. However, this dye had a significant genotoxic effect at all concentrations tested. Although most of the damage was amenable to repair, some damage remained higher than positive control after 24 h of repair. These data demonstrate that tartrazine may be harmful to health and its prolonged use could trigger carcinogenesis. PMID:25750299

  3. Study on the cytogenetic changes induced by benzene and hydroquinone in human lymphocytes.

    PubMed

    Peng, D; Jiaxing, W; Chunhui, H; Weiyi, P; Xiaomin, W

    2012-04-01

    Benzene (BN) is a prototypical hematotoxicant, genotoxic carcinogen, and ubiquitous environmental pollutant. Although the molecular mechanisms of BN-induced cytotoxicity and genotoxic damage are poorly understood in humans, previous studies suggested that bioactivated BN metabolites are capable of oxidative stress, cell cycle arrest, apoptosis, and DNA damage. The objective of the current study was to investigate the BN-induced cytogenetic changes and underlying mechanisms based on these hypotheses. Peripheral blood lymphocytes (PBLs) might be the targets for BN-induced cytotoxicity and genotoxicity, and therefore DNA damage responses of PBLs after exposure to different concentrations of BN (0.25, 3.5, 50 μmol/L) or BN metabolite, hydroquinone (HQ; 50, 150, 450 μmol/L) were studied in vitro. Microculture tetrazolium assay, flow cytometry, 2',7'-dichlorodihydrofluorescein-diacetate assay, comet assay, micronuclei assay, and attenuated total reflectance microspectroscope were chosen for this study. Based on the results, we reached the conclusion that different concentrations of BN or HQ significantly inhibited cell growth, induced the arrest of S phase and G2/M phase, and increased late apoptosis in a concentration-dependent manner. Furthermore, evidence was also provided to support the conclusion that BN and HQ induced DNA strand breaks and chromosomal mutations in PBL, which indicated the genotoxicity of BN and HQ. Current evidence has indicated that multiple mechanisms including dysfunction of cell cycle, programmed cell death, oxidative stress, and DNA lesions are likely to contribute to BN-induced cytogenetic changes. PMID:22297702

  4. Effect of insulin on low-density-lipoprotein metabolism in human lymphocytes in vitro.

    PubMed Central

    Suresh, S; Warty, V; Virji, M; Sanghvi, A

    1986-01-01

    The metabolism of low-density lipoproteins (LDL) in vitro in the presence of insulin was studied in freshly isolated human peripheral-blood lymphocytes. Insulin appeared to decrease the binding affinity of 125I-LDL to its cell-surface receptor, without any change in apparent Vmax or in the number of LDL receptors. As a consequence, the absolute amounts of 125I-LDL internalized and degraded were lower in the presence of insulin than in its abscence, although the fraction of internalized 125I-LDL degraded in either instance was quite similar. 3-Hydroxy-3-methylglutaryl-CoA reductase activity, and hence cholesterol synthesis, were stimulated by insulin. This effect of insulin was independent of the inhibitory effect of LDL on cholesterol synthesis. At the same time, acid cholesterol esterase and acyl-CoA: cholesterol O-acetyltransferase activities were lower in cells incubated with insulin than in controls. The net effect of these metabolic alterations seems to be that cells accumulate greater quantities of free and esterified cholesterol when treated with insulin. PMID:3513764

  5. Biochemical manipulation of intracellular glutathione levels influences cytotoxicity to isolated human lymphocytes by sulfur mustard

    SciTech Connect

    Gross, C.L.; Innace, J.K.; Hovatter, R.C.; Meier, H.L.; Smith, W.J.

    1993-12-31

    Glutathione (GSH) is the major nonprotein thiol that can protect cells from damage due to electrophilic alkylating agents by forming conjugates with the agent. Sulfur mustard (HD) is an electrophilic alkylating agent that has potent mutagenic, carcinogenic, cytotoxic, and vesicant properties. Compounds that elevate or reduce intracellular levels of GSH may produce changes in cytotoxicity induced by sulfur mustard. Pretreatment of human peripheral blood lymphocytes (PBL) for 72 hr with 1 mM buthionine sulfoximine (BSO), which reduces intracellular GSH content to approximately 26% of control, appears to sensitize these in vitro cells to the cytotoxic effects of 10 AM HD but not to higher HD concentrations. Pretreatment of PBL for 48 hr with 10 mM N-acetyl cysteine (NA C), which elevates intracellular glutathione levels to 122% of control, appears to partially protect these in vitro cells from the cytotoxic effects of 10 LAIHD but not to higher HD concentrations. Augmentation of intracellular levels of glutathione may provide partial protection against cytotoxicity of sulfur mustard.

  6. [Cytogenetic effect of thaliblastine in a culture of human peripheral blood lymphocytes].

    PubMed

    Khadzhidekova, V; Krusteva, E; Bulanova, M; Ivanov, V; Benova, D

    1988-12-01

    The mutagenicity of thaliblastine (Bulgarian potential antitumor drug) was investigated in vitro in lymphocytes from healthy donors, and in vivo in lymphocytes of oncological patients after thaliblastine administration. No increase in the rate of chromosome aberrations was noted with increasing thaliblastine concentrations in vitro and in the course of therapy in vivo. Some polyploid metaphases were found in the lymphocytes of the patients treated with thaliblastine, as a result of the statmokinetic effect of the drug. Thaliblastine exerts extraordinarily slight mutagenic effect, as compared with other cytostatics. PMID:2977980

  7. Unstimulated human CD4 lymphocytes express a cytoplasmic immature form of the common cytokine receptor gamma-chain.

    PubMed

    Bani, L; Pasquier, V; Kryworuchko, M; Salamero, J; Thèze, J

    2001-07-01

    As a component of various cytokine receptors, common cytokine receptor gamma-chain (gamma(c)) is essential in the development of the immune system and plays an important role in different stages of inflammatory and immune responses. Here we establish that resting CD4 T cells and the Jurkat CD4 T cell line do not express the mature form of gamma(c) (64 kDa) recognized by mAb Tugh4. However, these cells constitutively transcribe the corresponding gamma(c) gene. This apparent paradox was solved by the demonstration that polyclonal anti-gamma(c) Abs detected endoglycosidase-H-sensitive immature forms of gamma(c) (54-58 kDa) expressed by quiescent CD4 T lymphocytes and Jurkat cells. Immature gamma(c) is characterized as an intracellular component localized in the endoplasmic reticulum. Pulse-chase analysis shows that the immature gamma(c) is rapidly degraded after synthesis. After activation of CD4 T lymphocytes, and as seen in the CD4 T cell line Kit 225, the endoglycosidase-H-resistant mature form of gamma(c) is detectable at the cell surface and in the endosomal compartment. For the first time, our results demonstrate that a cytokine receptor chain may be constitutively produced as an immature form. Furthermore, this supports the notion that expression of the functional form of gamma(c) may require intracellular interactions with lineage- or subset-specific molecular partners. PMID:11418669

  8. The assessment of genotoxicity of carbamazepine using cytokinesis-block (CB) micronucleus assay in cultured human blood lymphocytes.

    PubMed

    Celik, Ayla

    2006-01-01

    The genotoxic effect of CBZ has been investigated in few studies. There is little evidence linking carbamazepine (CBZ) with any genotoxic effects, particularly in vitro micronucleus test using cytogenesis-block technique. In this study, the genotoxicity of the antiepileptic drug, carbamazepine, was tested using cytokinesis-block (CB) micronucleus assay. In vitro analysis was performed in human blood lymphocytes from four healthy persons at five different concentrations of carbamazepine (6, 8, 10, 12, 14 microg/mL). Genotoxic potential and cytotoxic effects of carbamazepine were evaluated by using micronucleus assay and cytokinesis-block proliferation index (CBPI), called the parameter of cytotoxicity in human peripheral blood lymphocyte cultures, respectively. The results of this study indicate that CBZ caused the genotoxic effect under in vitro conditions, except at the dose of 6 microg/mL, and cytotoxic effects of carbamazepine were revealed by a decrease in the cytokinesis-block proliferation index at all the concentrations. PMID:16707330

  9. Human T-lymphotropic virus type I-associated myelopathy and tax gene expression in CD4+ T lymphocytes.

    PubMed

    Moritoyo, T; Reinhart, T A; Moritoyo, H; Sato, E; Izumo, S; Osame, M; Haase, A T

    1996-07-01

    Infection by human T-lymphotropic virus type I (HTLV-I) is associated with adult T-cell leukemia and a slowly progressive disease of the central nervous system (CNS), HTLV-I-associated myelopathy/tropical spastic paraparesis, characterized pathologically by inflammation and white matter degeneration in the spinal cord. One of the explanations for the tissue destruction is that HTLV-I infects cells in the CNS, or HTLV-I-infected CD4+ T lymphocytes enter the CNS, and this drives local expansion of virus-specific CD8+ cytotoxic T lymphocytes, which along with cytokines cause the pathological changes. Because both in the circulation and in the cerebrospinal fluid, CD8+ cytotoxic T lymphocytes are primarily reactive to the product of the HTLV-I tax gene, we sought evidence of expression of this gene within cells in the inflammatory lesions. After using double-label in situ hybridization techniques, we now report definitive localization of HTLV-I tax gene expression in CD4+ T lymphocytes in areas of inflammation and white matter destruction. These findings lend support to a hypothetical scheme of neuropathogenesis in which HTLV-I tax gene expression provokes and sustains an immunopathological process that progressively destroys myelin and axons in the spinal cord. PMID:8687197

  10. Protective Effect of Carvacrol on Oxidative Stress and Cellular DNA Damage Induced by UVB Irradiation in Human Peripheral Lymphocytes.

    PubMed

    Aristatile, Balakrishnan; Al-Numair, Khalid S; Al-Assaf, Abdullah H; Veeramani, Chinnadurai; Pugalendi, Kodukkur Viswanathan

    2015-11-01

    Exposure to ultraviolet B (UVB; 280-320 nm) radiation induces the formation of reactive oxygen species (ROS) in the biological system. In this study, we examined the protective effect of carvacrol on UVB-induced lipid peroxidation and oxidative DNA damage with reference to alterations in cellular an-tioxidant status in human lymphocytes. A series of in vitro assays (hydroxyl radical, superoxide, nitric oxide, DPPH (2,2-Diphenyl-1-picryl hydrazyl), and ABTS (2,2-azino-bis-3-ethylbenzothiazoline-6-sulfonic acid) radical scavenging assays) demonstrate antioxidant property of carvacrol in our study. UVB exposure significantly increased thiobarbituric acid reactive substances (TBARS), lipid hydroperoxides (LHPs), % tail DNA and tail moment; decreased % cell viability and antioxidant status in UVB-irradiated lymphocytes. Treatment with carvacrol 30 min prior to UVB-exposure resulted in a significant decline of TBARS, LHP, % tail DNA, and tail moment and increased % cell viability as carvacrol concentration increased. UVB irradiated lymphocytes with carvacrol alone (at 10 μg/mL) gave no significant change in cell viability, TBARS, LHP, % tail DNA, and tail moment when compared with normal lymphocytes. On the basis of our results, we conclude that carvacrol, a dietary antioxidant, mediates its protective effect through modulation of UVB-induced ROS. PMID:26768646

  11. Relation of impaired lymphocyte proliferative function to other major human immunodeficiency virus type 1-induced immunological changes.

    PubMed Central

    Bass, H Z; Fahey, J L; Nishanian, P; Detels, R; Cumberland, W; Kemeny, M; Plaeger, S

    1997-01-01

    Human immunodeficiency virus (HIV) type 1 (HIV-1) induces impairment of immune function reflected in reduced lymphocyte proliferative responses. Many other immune changes are induced by HIV-1, but their relationship to lymphocyte functional defects is not known. The present study was designed to correlate functional defects with other HIV disease parameters. Cryopreserved samples from 118 HIV-1-positive subjects and 40 seronegative individuals were examined. The main findings were that impaired proliferative responses to mitogens correlated with (i) decreased cell surface expression of the interleukin-2 receptor (CD25), (ii) increased expression of HLA-DR antigens on CD4 cells, (iii) reduced CD4 and increased CD8 cell numbers, and (iv) increased levels of serum immune complex dissociated p24 antigen. However, impaired function was not associated with increased serum neopterin, beta2-microglobulin, or soluble interleukin-2 receptor or with CD38 antigen expression on lymphocytes. In summary, proliferative functional impairment correlated with some, but not all, immunological changes associated with HIV-1 infection. Most of the phenotypic markers that correlated with altered function are cell surface molecules with significant roles in lymphocyte proliferation and were associated primarily with CD4 cells, compatible with the view that dysregulation of CD4 cells is responsible for impaired function. PMID:9008283

  12. Effects of synthetic and naturally occurring flavonoids on mitogen-induced proliferation of human peripheral-blood lymphocytes

    SciTech Connect

    Hirano, Toshihiko; Oka, Kitaro; Kawashima, Etsuko; Akiba, Mitsuo )

    1989-01-01

    Examination was made of the effects of 17 synthetic and naturally occurring flavonoids on human lymphocyte proliferation in the presence of concanavalin A as a mitogen. Twelve of the flavonoids examined were mono-hydroxy of methoxy derivatives. The mitogen-induced response of lymphocytes was evaluated from the extent of the incorporation of ({sup 3}H)thymidine into cells in vitro. All the compounds showed inhibitory effects; 4.5-77.7% of ({sup 3}H) thymidine incorporation was blocked by an 1.0 {mu}g/ml concentration. The viability of lymphocytes before and after treatment, as assessed by a dye exclusion test, indicated no change, and thus the flavonoids may inhibit DNA synthesis. The flavonoids possessing 5-hydroxyl, 5-methoxyl and 6-methoxyl groups, and those with cyclohexyl instead of phenyl substituent (i.e. 2-cyclohexyl-benzopyran-4-one), showed the greatest inhibition. The inhibitory effect of any one of them was less than one half that of prednisolone, but essentially the same or somewhat exceeding that of bredinine of azathioprine. It would thus appear that the well-known anti-inflammatory effects of flavonoids may possibly arise in part from the inhibition of the proliferative response of lymphocytes.

  13. TRIP6 and LPP, but not Zyxin, are present at a subset of telomeres in human cells

    PubMed Central

    Sheppard, Samantha A; Savinova, Tatiana

    2011-01-01

    The protection of chromosome ends requires the inhibition of DNA damage responses at telomeres. This inhibition is exerted in great part by the shelterin complex, known to prevent inappropriate ATM and ATR activation. The molecular mechanisms by which shelterin protects telomeres are incompletely understood. Recently, we have implicated for the first time a class of molecules, LIM domain proteins, in telomere protection. This protection occurred through interaction with shelterin, possibly through POT1, and required the pair of LIM proteins TRIP6 and LPP, themselves part of the Zyxin family. The domain similarity between TRIP6, LPP and Zyxin led us to ask whether the latter also interacted with telomeres. Here, we show that there is specificity in the association of LIM proteins with telomeres: Zyxin, despite a high degree of similarity with TRIP6 and LPP, was not detected at telomeres, nor found in a complex with shelterin. TRIP6 and LPP, however, were detected by immunofluorescence at a small subset of telomeres, perhaps those that are critically short. We speculate that specific LIM proteins are part of complex events occurring in the context of the telomere dysfunction response and are possibly at play during the induction of senescence. PMID:21519191

  14. In vitro genotoxicity and cytotoxicity of a particular combination of pemetrexed and cefixime in human peripheral blood lymphocytes.

    PubMed

    Istifli, Erman Salih; Topaktaş, Mehmet

    2015-01-01

    This study aims to find the genotoxic and cytotoxic effects of a particular combination of pemetrexed (PMX) and cefixime (CFX) in human peripheral blood lymphocytes. Chromosome aberration (CA), sister chromatid exchange (SCE), and micronucleus (MN) tests were used to assess genotoxicity. Whereas, the cytotoxicity was evaluated by using mitotic index (MI), proliferation index (PI), and nuclear division index (NDI). Our tests were proceeded with concentrations of 12.5 + 450, 25 + 800, 37.5 + 1150, and 50 + 1500 μg/mL of a mixture of PMX and CFX separately for 24 hr and 48 hr. The combination of PMX + CFX did not induce the CA or SCE in human peripheral blood lymphocytes when compared with both the control and the solvent control. MN in human peripheral blood lymphocytes was not significantly increased after treatment with a particular combination of PMX + CFX. However, PMX + CFX significantly decreased the MI, PI and NDI at all concentrations for 24- and 48-hr treatment periods when compared with both controls. Generally, PMX + CFX inhibited cell proliferation more than positive control (MMC) and showed a higher cytotoxic effect than MMC at both treatment periods. These results were compared with individual effects of PMX and CFX. As a result, it was observed that a particular combination of PMX + CFX was not genotoxic. However, the combination synergistically increase cytotoxicity in human peripheral blood lymphocytes. PMID:25653913

  15. Behavior and Properties of Mature Lytic Granules at the Immunological Synapse of Human Cytotoxic T Lymphocytes

    PubMed Central

    Ming, Min; Schirra, Claudia; Becherer, Ute; Stevens, David R.; Rettig, Jens

    2015-01-01

    Killing of virally infected cells or tumor cells by cytotoxic T lymphocytes requires targeting of lytic granules to the junction between the CTL and its target. We used whole-cell patch clamp to measure the cell capacitance at fixed intracellular [Ca2+] to study fusion of lytic granules in human CTLs. Expression of a fluorescently labeled human granzyme B construct allowed identification of lytic granule fusion using total internal reflection fluorescence microscopy. In this way capacitance steps due to lytic granule fusion were identified. Our goal was to determine the size of fusing lytic granules and to describe their behavior at the plasma membrane. On average, 5.02 ± 3.09 (mean ± s.d.) lytic granules were released per CTL. The amplitude of lytic granule fusion events was ~ 3.3 fF consistent with a diameter of about 325 nm. Fusion latency was biphasic with time constants of 15.9 and 106 seconds. The dwell time of fusing lytic granules was exponentially distributed with a mean dwell time of 28.5 seconds. Fusion ended in spite of the continued presence of granules at the immune synapse. The mobility of fusing granules at the membrane was indistinguishable from that of lytic granules which failed to fuse. While dwelling at the plasma membrane lytic granules exhibit mobility consistent with docking interspersed with short periods of greater mobility. The failure of lytic granules to fuse when visible in TIRF at the membrane may indicate that a membrane-confined reaction is rate limiting. PMID:26296096

  16. Cannibalism of live lymphocytes by human metastatic but not primary melanoma cells.

    PubMed

    Lugini, Luana; Matarrese, Paola; Tinari, Antonella; Lozupone, Francesco; Federici, Cristina; Iessi, Elisabetta; Gentile, Massimo; Luciani, Francesca; Parmiani, Giorgio; Rivoltini, Licia; Malorni, Walter; Fais, Stefano

    2006-04-01

    The phenomenon of cell cannibalism, which generally refers to the engulfment of cells within other cells, was described in malignant tumors, but its biological significance is still largely unknown. In the present study, we investigated the occurrence, the in vivo relevance, and the underlying mechanisms of cannibalism in human melanoma. As first evidence, we observed that tumor cannibalism was clearly detectable in vivo in metastatic lesions of melanoma and often involved T cells, which could be found in a degraded state within tumor cells. Then, in vitro experiments confirmed that cannibalism of T cells was a property of metastatic melanoma cells but not of primary melanoma cells. In particular, morphologic analyses, including time-lapse cinematography and electron microscopy, revealed a sequence of events, in which metastatic melanoma cells were able to engulf and digest live autologous melanoma-specific CD8(+) T cells. Importantly, this cannibalistic activity significantly increased metastatic melanoma cell survival, particularly under starvation condition, supporting the evidence that tumor cells may use the eating of live lymphocytes as a way to "feed" in condition of low nutrient supply. The mechanism underlying cannibalism involved a complex framework, including lysosomal protease cathepsin B activity, caveolae formation, and ezrin cytoskeleton integrity and function. In conclusion, our study shows that human metastatic melanoma cells may eat live T cells, which are instead programmed to kill them, suggesting a novel mechanism of tumor immune escape. Moreover, our data suggest that cannibalism may represent a sort of "feeding" activity aimed at sustaining survival and progression of malignant tumor cells in an unfavorable microenvironment. PMID:16585188

  17. Human ecalectin, a variant of human galectin-9, is a novel eosinophil chemoattractant produced by T lymphocytes.

    PubMed

    Matsumoto, R; Matsumoto, H; Seki, M; Hata, M; Asano, Y; Kanegasaki, S; Stevens, R L; Hirashima, M

    1998-07-01

    A 1.6-kilobase pair cDNA was isolated from a human T-cell-derived expression library that encodes a novel eosinophil chemoattractant (designated ecalectin) expressed during allergic and parasitic responses. Based on its deduced amino acid sequence, ecalectin is a 36-kDa protein consisting of 323 amino acids. Although ecalectin lacks a hydrophobic signal peptide, it is secreted from mammalian cells. Ecalectin is not related to any known cytokine or chemokine but rather is a variant of human galectin-9, a member of the large family of animal lectins that have affinity for beta-galactosides. Recombinant ecalectin, expressed in COS cells and insect cells, exhibited potent eosinophil chemoattractant activity and attracted eosinophils in vitro and in vivo in a dose-dependent manner but not neutrophils, lymphocytes, or monocytes. The finding that the ecalectin transcript is present in abundance in various lymphatic tissues and that its expression increases substantially in antigen-activated peripheral blood mononuclear cells suggests that ecalectin is an important T-cell-derived regulator of eosinophil recruitment in tissues during inflammatory reactions. We believe that this is the first report of the expression of an immunoregulatory galectin expressed by a T-cell line that is selective for eosinophils. PMID:9642261

  18. Evaluation of T and B lymphocyte membrane markers in human non-Hodgkin malignant lymphomata.

    PubMed Central

    Brouet, J. C.; Labaume, S.; Seligmann, M.

    1975-01-01

    Lymphoma cells from 25 patients were studied for the prese