Vitamin C and E suppress mitogen-stimulated peripheral blood mononuclear cells in vitro

BACKGROUND/AIMS: The antioxidant properties of vitamin C and E are considered to be important for their anti-inflammatory activity. Recently, antioxidant resveratrol was found to suppress neopterin production and tryptophan degradation in mitogen-treated human peripheral blood mononuclear cells. METHODS: In this study, the effects of vitamin C and E were investigated in unstimulated peripheral blood mononuclear cells and in cells stimulated with the mitogens phytohaemagglutinin and concanavalin A in vitro. RESULTS: The mitogens induced a significant production of neopterin and a degradation of tryptophan. Vitamin C (0.1-10 microM) and E (5-100 microM) suppressed these immunobiological pathways in a dose-dependent way (p < 0.01). CONCLUSION: Neopterin production and tryptophan degradation in monocyte-derived macrophages are both triggered by the pro-inflammatory cytokine interferon-gamma. Thus, their concurrent suppression by vitamin C and E suggests an effect on the formation and release of this cytokine by stimulated T cells. These findings may be related to the general health benefits which are associated with the antioxidant nature of these vitamins.     

T gamma cells in sarcoidosis: E rosetting monocytes suppress lymphocyte transformation

Increased suppressor T gamma lymphocytes have been described in sarcoidosis. We have shown that a proportion of these cells are esterase-positive, phagocytic, adherent to plastic and stain with an anti-monocyte serum. Removal of these cells or the addition of indomethacin increases the lymphocyte transformation to Con A. Transformation was still reduced in spite of preincubation with plastic and the addition of indomethacin suggesting that a further abnormality exists. Thus, within the increased number of T gamma cells there exists a population of activated monocytes which rosette with sheep red blood cells and could therefore be mistaken for T cells.     

The effects of kwashiorkor serum on lymphocyte transformation in vitro.

The serum from twelve children with kwashiorkor was deficient in its ability to support lymphocyte transformation in vitro, whereas lymphocytes from these children responded to phytohaemagglutinin and al-ogeneic lymphocytes in a relatively normal manner when cultured in normal serum. This serum abnormality improved with therapy and could not be clearly correlated with the degree of malnutrition, the presence or absence of infection or other laboratory manifestations of kwashiorkor. These observations indicate that defective cellular immune reactions in kwashiorkor may be symptomatic of a lack of some humoral factor and do not necessarily reflect an intrinsic cellular defect.     

Plasmodium falciparum products enhance human lymphocyte transformation by Epstein-Barr virus

Supernatants obtained from the in vitro culture of Plasmodium falciparum infected erythrocytes induced prolonged lymphocyte survival in culture for more than 8 weeks in six cultures and permanent cell lines were established in four of these. The cells in the latter showed lymphoblastoid features similar to those seen in parallel cultures to which transforming Epstein-Barr (EB) virus instead of P. falciparum derived substances had been added. Cells from the same donors stimulated with other mitogens (pokeweed mitogen, Salmonella paratyphi culture supernatants) ceased to proliferate and died after 3-4 weeks. A 195 Kd polypeptide obtained from P. falciparum parasites also exhibited the potential to transform normal lymphocytes. Characterization of the cell lines indicated a B lymphocyte origin and the presence of EB virus in these lines suggests the possibility that P. falciparum products may activate latent EB virus genomes. These observations appear relevant to both the choice of P. falciparum derived antigens as vaccines, and to the interaction of EB virus and malaria in the aetiology of African Burkitt's lymphoma (BL).     

Triticum mosaic poacevirus enlists P1 rather than HC-Pro to suppress RNA silencing-mediated host defense

Triticum mosaic virus (TriMV) is the type species of the newly established Poacevirus genus in the family Potyviridae. In this study, we demonstrate that in contrast to the helper component-proteinase (HC-Pro) of Potyvirus species, the P1 proteins of TriMV and Sugarcane streak mosaic poacevirus function in suppression of RNA silencing (SRS). TriMV P1 effectively suppressed silencing induced by single- or double-stranded RNAs (ss/ds RNAs), and disrupted the systemic spread of silencing signals at a step after silencing signal production. Interestingly, contrary to enhanced SRS activity of potyviral HC-Pro by co-expression with P1, the presence of TriMV HC-Pro reduced SRS activity of TriMV P1. Furthermore, TriMV P1 suppressed systemic silencing triggered by dsRNA more efficiently than the HC-Pro of Turnip mosaic potyvirus. Furthermore, TriMV P1 enhanced the pathogenicity of a heterologous virus. Our results established poaceviral P1 as a potent RNA silencing suppressor that probably employs a novel mechanism to suppress RNA silencing-based antiviral defense.     

Microtechnique for quantitative evaluation of in vitro lymphocyte transformation

A microtechnique is described to quantitate in vitro transformation of lymphocytes from man and marmosets following stimulation by PHA and different antigens. The ¹⁴C-thymidine uptake of lymphocytes grown in cultures of 0·06–0·2 ml of whole blood was measured by liquid scintillation. The magnitude, consistency and reproducibility of the results were similar to those achieved using cultures of purified lymphocytes or cell-rich plasma but fewer technical procedures were required.     

Human transfer factor in vitro. II. Augmentation of lymphocyte transformation to phytohaemagglutinin

This paper describes experiments in which human peripheral blood lymphocyte transformation by phytohaemagglutinin (PHA) is augmented by the addition of human dialysable transfer factor. Dextran-separated peripheral blood leucocytes were cultured for 4 days with a range of PHA concentrations so as to produce low, moderate, high or very high incorporation of [³H]thymidine. When transfer factor preparations were added to the cultures, in concentrations similar to those augmenting lymphocyte transformation to tuberculin PPD, augmentation of PHA responses was observed in two-thirds of the experiments (25/38). In these experiments the extent of augmentation was proportional to the level of DNA synthesis induced by PHA in the absence of added transfer factor. It appears that preparations of transfer factor are able to augment lymphocyte transformation responses to PHA in vitro, but that this effect occurs with less regularity than does augmentation of lymphocyte transformation to tuberculin PPD.     

Low extracellular Mg2+ concentrations suppress phagocytosis in vitro by alveolar macrophages from rats.

The aim of this study was to assess the effects of extracellular Mg2+ concentrations on phagocytosis in vitro by alveolar macrophages from rats. Phagocytosis was suppressed in the presence of low, but not high, Mg2+ concentrations. Vanadate, a Mg(2+)-ATPase inhibitor, suppressed phagocytosis. In the presence of a low Mg2+ concentration or vanadate, the cytosolic free Ca2+ concentration ([Ca2+]i) increased, but the cytosolic free Mg2+ concentration did not. These results suggest that low extracellular Mg2+ concentrations and vanadate suppress phagocytosis by rat alveolar macrophages by increasing [Ca2+]i.     

Lymphocyte transformation in vitro. III. Mechanism of stimulation in the mixed lymphocyte culture

In view of the correlation between the mixed lymphocyte culture and the HL-A locus, experiments were performed to determine whether the reactivity in the mixed lymphocyte culture is elicited by the HL-A antigens as such.

Fresh lymphocytes were mixed with either allogeneic fibroblasts or allogeneic lymphocytes treated with various metabolic inhibitors or by heating which abolishes their capacity to tranfsorm in vitro.

No stimulation of the normal untreated lymphocytes was observed in any of these mixed cultures.

However, the HL-A antigens 4b and 7b were not affected quantitatively by this treatment as determined serologically.

The conclusion is drawn that mixed lymphocyte culture reactivity is not merely a reaction to HL-A antigens, and the consequences of this are discussed.

     

Lymphocyte transformation in vitro. II. Mixed lymphocyte culture in relation to leucocyte antigens

By means of family studies the relation between leucocyte antigens and reactivity in mixed lymphocyte culture was investigated.     

Cell-mediated immunity to Chlamydia pneumoniae measured as lymphocyte blast transformation in vitro.

The purpose of the present study was to analyze Chlamydia pneumoniae-induced, antigen-specific, cell-mediated immunity. Peripheral blood mononuclear cells of four persons infected with C. pneumoniae Kajaani 6 and 17 healthy volunteers were stimulated with antigen composed of whole elementary bodies of C. pneumoniae Kajaani 6 (CP-Ag). Definitive antigen-specific lymphoproliferation (LP) responses were developed after recent infection. The LP responses of healthy people to CP-Ag varied considerably. There was no clear correlation between LP responses to CP-Ag and those to an antigen prepared from Chlamydia trachomatis serotype L2 (r > or = 0.50, P < 0.1). A larger study is required to demonstrate whether the LP responses to CP-Ag can be used for the diagnosis of C. pneumoniae infection.     

Colony growth in vitro of mitogen-stimulated mouse B lymphocytes.

The ability of mouse B-mitogen-induced lymphocytes to grow and develop into colonies in a soft agar system was studied. Prerequisite conditions for the colony formation of mouse lymphocytes from inguinal lymph nodes of strains ICR C3H/eB and C3H were their suspension in a liquid medium and stimulation with polyclonal B-cell activators such as bacterial lipopolysaccharide (LPS), purified protein derivative (PPD) or dextran sulphate (DxS) prior to being seeded on a soft agar culture medium. After 3-5 days of culture, colonies of 50-350 cells or more per clone developed. A linear relationship was found between the number of cells seeded and the number of colonies growing. Of the cells seeded, only a limited population of the mitogen-stimulated lymphocytes has the potential to divide and to develop into colonies. The largest number of colonies was obtained by culturing lymph node cells of ICR mice and using LPS as mitogens. Two sublines of C3H were found to respond differently to LPS: C3H/HeJ mice were low responders while C3H/eB mice were high responders. Experiments with inbred, congenitally athymic nude (nu/nu) mice known to be deficient in T cells showed that LPS-stimulated lymphocytes were capable of forming colonies. The morphology of the colony cells, as well as the fact that they stain positively for cell-membrane immunoglobulins, suggest that the colonies developed from B lymphocytes.     

Deficiency in kwashiorkor serum of factors required for optimal lymphocyte transformation in vitro.

Blastogenic responses of normal human peripheral blood lymphocytes cultured in media supplemented with serum from children with kwashiorkor were, on average, 47.7% of those observed when the same cells were cultured in the presence of normal AB serum. Incorporation of radioactive uridine was also diminished in the presence of normal AB serum. Incorporation of radioactive uridine was also diminished in the presence of kwashiorkor serum indicating that lectin-induced RNA synthesis was also affected. The kwashiorkor serum effect was not due to a cytotoxic action nor could it be attributed to the presence of saccharides or other inhibitors of the inducing lectins. Mixing experiments showed that kwashiorkor serum was not inhibitory, but that it lacked factors present in normal serum that are required for optimal lymphocyte blastogenesis. The deficiency of these factors could largely be rectified by supplementing kwashiorkor serum with an ultrafiltrate of normal serum containing components with molecular weights of less than 500 Daltons. We conclude that nutritional deprivation of severity sufficient to cause kwashiorkor leads to a deficiency of low molecular weight lymphocyte growth factors. This lack may contribute to the immunodeficiency associated with the disease.     

Silica-induced pulmonary fibrosis involves the reaction of particles with interstitial rather than alveolar macrophages

Macrophage-derived products have been implicated in fibroblast stimulation following particle deposition in the lung. To assess the role of macrophages in the alveolus versus those in the interstitium in the induction of pulmonary fibrosis, we compared the pulmonary response to silica when phagocytosis occurred predominantly in each of these compartments. One group of mice received intratracheal silica which was phagocytosed largely by alveolar macrophages (AM). A second group was exposed to whole body irradiation prior to receiving the same dose of silica. This prevented the usual efflux of PMN and monocytes into the air sacs, allowing passage of silica particles across the alveolar epithelium to reach the interstitial macrophages (IM). In the irradiation plus silica group, many large interstitial granulomas were formed at 2-4 weeks, and collagen levels were significantly greater than in all other groups at 16 weeks. More silica was found in a lung tissue residue and in lymph nodes of these animals. Pulmonary fibrosis was limited to interstitial areas where there was a high level of retained silica, whereas peripheral regions of the lung, where free AM containing silica were found, did not show fibrosis of the alveolar walls. The results suggest that factors secreted by IM in response to silica are more effective in stimulating fibrogenesis than secretions made by the AM into the alveolar space.     

Human transfer factor in vitro. I. Augmentation of lymphocyte transformation to tuberculin PPD

There is recent interest in whether dialysable transfer factor can specifically increase immune responses when added to lymphoid cells in vitro. This report demonstrates that transfer factor preparations (human leucocyte dialysates) `augment' rather than `transfer' lymphocyte transformation responses (DNA synthesis) to tuberculin PPD in vitro and that the magnitude of this augmentation is proportional to the level of DNA synthesis induced by PPD in the absence of added transfer factor. Experiments showed that transfer factor preparations from Mantoux-positive or Mantoux-negative `donors' were equally effective in augmenting `recipient' lymphocyte transformation responses to PPD. Thus the extent of augmentation was related, not to the tuberculin sensitivity of the transfer factor donors, but to that of the recipients. In the absence of tuberculin PPD, transfer factor preparations sometimes stimulated lymphocyte DNA synthesis, but the extent of this was small and inconstant. The results therefore provide evidence for an antigen-dependent, but not an antigen-specific effect of transfer factor in increasing lymphocyte DNA synthesis in vitro. It is suggested that leucocyte dialysates contain an augmenting factor which may facilitate the response of antigen-sensitive cells to PPD in vitro, or may facilitate the recruitment into DNA synthesis of cell populations responding to mitogenic lymphokine produced during lymphocyte transformation.