Sendai virus induces high levels of tumor necrosis factor mRNA in human peripheral blood leukocytes.

Sendai virus induces human peripheral blood leukocytes to produce high levels of tumor necrosis factor (TNF) mRNA. TNF mRNA can represent as much as 0.6% of the total mRNA. Kinetic studies indicate that the level of TNF mRNA peaks about 2 hours before that of IFN-alpha mRNA produced in the same system. Although the peak levels of TNF and IFN-alpha mRNA were similar, TNF in the culture supernatants was at a 200 fold lower level than IFN-alpha. Cloning and sequence analysis of TNF cDNA isolated from peripheral blood leukocytes RNA showed that normal human cells in response to Sendai virus produce TNF identical to that previously isolated and cloned from tumor-derived cell lines. A bacterial expression system was used to produce the cloned TNF at a maximum level of 2 X 10(6) units per ml of culture.     

Inhibitory action of azelastine on cytokine production from human peripheral blood leukocytes in vitro

Effects of azelastine (AZ) on human peripheral blood leukocytes (PBL) in culture were examined. Addition of 10.0 μg/ml of AZ resulted in a marked inhibition of PBL blastic activity, but lower concentrations (1.0 and 0.5 μg/ml) had no demonstrable suppressive effects on the activation. Interleukin (IL)-2, IL-3 and IL-4 production from PBL in response to Concanavalin A stimulation was also strongly suppressed when the cells were cultured in the presence of AZ. This suppression was observed even when lower concentrations (1.0 and 0.5 μg/ml) of AZ were added to cell cultures     

Dengue virus replication enhancement in peripheral blood leukocytes from immune human beings.

Dengue 2 virus replication in peripheral blood leukocyte cultures from 10 of 13 Asian or Polynesian subjects with actively acquired dengue immunity and two of three infants with passively acquired dengue antibody. Only 2 of 11 cultures from nonimmune infants or children supported viral replication. This study establishes a parallel in the biological behavior of human and simian PBL with respect to the immunological dependence of dengue virus replication in vitro. Elucidation of the mechanism (or mechamisms) regulating growth of dengue virus in leukocytes from immune hosts may contribute to an understanding of the role that virus-leukocyte interactions play in the pathogenesis of human dengue illness.     

Pleiotrophin induces expression of inflammatory cytokines in peripheral blood mononuclear cells

Pleiotrophin (PTN) is a polypeptide that belongs to a family of heparin-binding growth factor, which displays mitogenic activity for a wide variety of cells. Since PTN induces the proliferation of immune cells the mechanism of action was investigated. In the present study, we show for the first time that PTN induces the expression of inflammatory cytokines including TNF-alpha, IL-1beta and IL-6 in quiescent human peripheral blood mononuclear cells (PBMC). These results emphasize the importance of PTN in the regulation of inflammatory processes. Elucidation of the mechanisms by which a host factor such PTN regulates cytokines production will significantly advance our understanding of endothelium-immunity interactions.     

Genotoxic effect of ozone in human peripheral blood leukocytes

The genotoxic effect of ozone was studied in human leukocytes in vitro, using the single cell gel electrophoresis (SCGE) assay. Cell treatment for 1 h at 37 degrees C with 0.9-5.3 mM O(3) resulted in a dose-dependent increase of DNA damage, comparable to that induced by 4-40 mM of H(2)O(2), used as a positive control. This effect of ozone was reversed by post-treatment incubation of the cells for 45-90 min at 37 degrees C, and prevented by pre-incubation of the cells with catalase (20 microg/ml). These results demonstrate that O(3) induces DNA-damage in primary human leukocytes. The damage is rapidly repaired, and probably mediated by the formation of H(2)O(2).     

Plasma membrane fluidity gradients of human peripheral blood leukocytes

The shape of the fluidity gradient of the outer hemileaflet of the plasma membrane of normal, living, human white blood cells was determined using a series of n-(9-anthroyloxy) fatty acid probes where n = 2, 3, 6, 7, 9, 11, 12, and 16, to establish a baseline for future studies on the consequences of various pathological states. Fluorescence uptake and steady-state anisotropy values were obtained with a flow cytometer capable of continuous recording over time of vertical and horizontal emission intensities, with the output of these intensities as calculated anisotropy values. The fluorescence uptake of all of the membrane probes was rapid up to about 15 min. The magnitudes of the uptake of fluorescence was, for the n-(9-anthroyloxy) series, in the order 2 greater than 3 greater than 6 greater than 7 greater than 9 greater than 11 = 12 = 16 for neutrophils, lymphocytes, and monocytes. Anisotropy values were constant from 5 to 30 min after addition of the various probes. The orders of the anisotropy magnitudes, indicative of the shapes of the fluidity gradient, were, for neutrophils, 6 greater than 7 greater than 9 greater than 2 = 3 = 11 = 12 greater than 16, for lymphocytes, 7 greater than 6 greater than 9 greater than 11 greater than 2 = 3 greater than 11 = 12 greater than 16, and for monocytes, 9 greater than 7 greater than 6 greater than 11 greater than 2 = 3 greater than 12 greater than 16. The kinetics of anisotropy from 1 to 5 min after addition of the probes differed for each of the three cell types. Probes with an n-value less than or equal to the maxima (n = 6, neutrophils; n = 7, lymphocytes; n = 9, monocytes) rapidly (1.2 min) reached equilibrium, whereas those probes with n-values greater than the maxima took progressively longer times to equilibrate as n increased. This behavior is consistent with the existence of an energy barrier just below the approximate region sensed by the probes, which would correspond to just below 6AS for neutrophils, 7AS for lymphocytes, and 9AS for monocytes.     

Limited effect of selected organic pollutants on cytokine production by peripheral blood leukocytes

To test the hypothesis that some persistent organic pollutants contribute to the increased prevalence of allergic disease, the effects of selected compounds on cytokine production by PBMC from control and allergic donors were evaluated. Cells were cultured for six days in the presence of a xenobiotic (PCB 153, hexachlorobenzene, pentachlorobenzene, pentachlorophenol, lindane, atrazine or DMSO vehicle) with phytohemagglutinin (PHA) or Dermatophagoides pteronyssinus extract, then for one day in the presence of PHA + phorbol 12-myristate 13-acetate. PCB 153 reduced the levels of IL-10, IFN-gamma and TNF-alpha. Hexachlorobenzene reduced the levels of IL-5, IL-10 and IFN-gamma. Pentachlorobenzene reduced IL-6 levels. Pentachlorophenol reduced IL-5 levels. Lindane and atrazine reduced both IL-5 and IFN-gamma. In addition, lindane reduced TNF-alpha levels. As these compounds had similar effects on cells from allergic and non-allergic donors, and as these effects were, in all cases, very limited indeed, the potential deleterious impact of the xenobiotics tested on the allergic response seems unlikely.     

Influenza virus infection induces functional alterations in peripheral blood lymphocytes

This report describes alterations in functional responses to lectin-induced stimulation of peripheral blood lymphocytes and in the natural killer cell (NKC) activity, of college students, obtained during an outbreak of influenza A/Philippines/2/82(H3N2) virus infection. These results are compared with similar observations in college students with an acute, febrile, noninfluenzal respiratory illness that occurred during the same outbreak. The lymphopenia typical of influenza during acute illness was shown to be due to a reduction in both T and B cells without alteration in the CD4:CD8 ratio. In addition, phytohemagglutinin and concanavalin A responses were reduced and NKC activity was increased, while pokeweed mitogen reactivity was unaltered at the time of admission to the study. Patients with noninfluenzal illness showed early polymorphonuclear leukocytosis and a similar lymphopenia. Lymphocyte functions were virtually unchanged during acute illness in noninfluenza patients. The relatively specific alterations in lymphocyte responses to lectin-induced stimulation in influenza patients may indicate that the peripheral T cells are incapable of activation via the CD3 or CD2 activation pathways. In addition, increased NKC activity in the periphery may be reflective of increased NKC activity in the lung. Influenza-infected individuals with higher NKC activity at the time of admission to the study also took longer to recover. Finally, the early lymphopenia and the later neutropenia in the influenza-infected patient may represent migration of these cells from the circulation to the infected respiratory tract as a consequence of infection.     

Adiponectin induces the anti-inflammatory cytokines IL-10 and IL-1RA in human leukocytes

There is increasing evidence that the adipose tissue and immunologic processes are closely linked. The most abundant protein within the adipocyte is adiponectin. Our current work reports that adiponectin has potent immuno-suppressive properties, as it induces the production of the anti-inflammatory mediators IL-10 and IL-1RA in primary human monocytes, monocyte-derived macrophages, and dendritic cells. In addition, adiponectin significantly impaired the production of the pro-inflammatory cytokine IFN-gamma in human macrophages. Moreover, adiponectin-treated macrophages exhibit a reduced phagocytotic and allo-stimulatory capacity. However, we could not detect any functional deficits or phenotypic changes in adiponectin-treated monocytes and monocyte-derived DC. In summary, the presented data support the idea that adiponectin might be of critical relevance for cytokine regulation in obesity and fatty liver diseases affecting primarily macrophage functions. This might represent a fundamental link between over-nutrition and an impaired inflammatory immune response.     

Functional activities of rosette separated human peripheral blood leukocytes.

Subpopulations of human peripheral blood leukocytes were isolated by rosette formation and tested for functional activity. E -rosette-forming cells (E-RFC) and EAC-RFC were separated from non-resetting cells by sedimentation on Ficoll-Hypaque gradients. The efficiency of the method and the purity of the resulting subpopulations were high. E-RFC responded to PHA Con A, allogeneic leukocytes, and PPD, with higher levels of proliferative reactivity than the unseparated lymphocytes while E-RFC depleted, EAC-RFC, and null cells showed only low levels of reactivity. Reactivity to PWM and tetanus toxoid was also restricted to the E-RFC subpopulation, but was lower than that of unseparated cells. A staphylococcal antigen preparation triggered lymphoproliferative reactivity in the E-RFC, E-RFC depleted, EAC-RFC, and the null cell subpopulations. 51Cr release lymphocyte cytotoxicity against a human lymphoblast target cell line was found in the E-RFC and null cell fractions but was not observed with the EAC-RFC subpopulation.     

Evaluation of human peripheral blood leukocytes for mast cell tryptase

Murine monoclonal and goat polyclonal antibodies against tryptase, the dominant neutral protease and protein component in secretory granules of human mast cells, were used to assess the presence of tryptase in peripheral leukocytes. Carnoy's fluid-fixed cytocentrifuge preparations of enriched populations of lymphocytes, monocytes, eosinophils, and neutrophils showed no reactivity with anti-tryptase antibodies by a sensitive indirect immunoperoxidase procedure. Dispersed human lung mast cells showed strong granular cytoplasmic staining with both antibodies, whereas only approximately 50% of the peripheral blood basophils detectable with Wright's stain were detected with anti-tryptase antibodies, and these showed a staining pattern that was faint, granular, and cytoplasmic at high concentrations of antibody. At lower antibody concentrations mast cell staining was still intense, whereas basophils were not stained. Extracts of neutrophils and lymphocytes of up to 90% purity had undetectable amounts of tryptase by an ELISA sandwich immunoassay, as well as undetectable enzymatic activity with tosyl-L-gly-pro-lys-p-nitroanilide (a sensitive substrate for tryptase) in the presence of soybean trypsin inhibitor. Extracts of basophil-enriched (6 to 50% purity) preparations contained 0.046 +/- 0.013 pg of tryptase per basophil by the immunoassay along with 2 X 10(-9) +/- 0.8 X 10(-9) U of tryptase-like enzyme activity per basophil, compared with corresponding values of 12 pg, 480 X 10(-9) U of tryptase per human lung mast cell. Thus very small amounts of tryptase are present in human basophils (approximately 0.4% of that found in mast cells), but not in other peripheral leukocytes.     

The varicella-zoster virus induces apoptosis in vitro in subpopulations of primary human peripheral blood mononuclear cells

Varicella-zoster virus (VZV), a member of Herpesviridae, subfamily alpha-Herpesvirinae, is pathogenic exclusively in the human. Chickenpox is the result of primary infection of VZV. During the viremic stage, VZV infects peripheral blood mononuclear cells (PBMC) and spreads to the periphery. In skin cells it causes typical lesions. Apoptosis has been demonstrated in different cell types by other alpha-herpesviruses. VZV-infected T lymphocytes, B lymphocytes, and monocytes, respectively, were examined in this in vitro study by flow cytometry, immunofluorescence and electron microscopy. All infected cell types showed signs of apoptosis: a lower DNA content, DNA fragmentation, loss of membrane integrity, and an altered nuclear morphology. The results observed led to the suggestion that VZV can induce apoptosis during infection in vivo in the PBMC subpopulations.     

A macrophage derived blastogenic factor -MBF- induces cytokines and cytotoxic effectors in human peripheral blood mononuclear cells.

A soluble macrophage-derived blastogenic factor, previously reported as MBF, is secreted from macrophages activated with galactose oxidase. It was previously shown that MBF is able to induce IFN-gamma production and proliferation of T lymphocytes. In this study we found that MBF is able to induce in human peripheral blood mononuclear cells (PBMC) production of interleukin 1 (IL-1) beta, interleukin 2 (IL-2) and tumor necrosis factor (TNF) alpha and generation of MHC-unrestricted cytotoxic activity. The induction of killer cells is likely to rely on IFN-gamma production in that in PBMC treated with a monoclonal antibody (Mab) against IFN-gamma, the MBF induced cytotoxic activity was drastically reduced. A comparison of MBF induced cytotoxic effectors with those induced by IL-2 showed that both cytotoxic effectors pertain to NK lineage, in that they were CD3- and CD16+. On the contrary, the precursors of MBF and IL-2 induced killer cells were different; MBF cytotoxic precursor cells were highly sensitive to L-Leucine methyl ester (Leu-OME), a drug able to eliminate monocytes and NK cells, whereas IL-2 cytotoxic precursors were unaffected by this drug.     

A comparison of the susceptibility to fusion of cell monolayers of human origin by inactivated Sendai virus

Cell monolayers of human foetal liver and lung, SV40 transformed foetal liver, HeLa cells and tonsils, showed a wide range of susceptibility to fusion by inactivated Sendai virus. Monolayers from human leucocytes, thyroid, foetal skin and thymus were also susceptible to fusion. All cells tested were found to be more susceptible to fusion in monolayers than in suspension.