Sodium salicylate and bile acid-induced colonic secretion in the rat

Previous studies by us suggested that luminally administered sodium salicylate blocks dihydroxy-bile acid-induced colonic secretion in the rat. In the present study, an in vivo rat cecal loop technique is employed to compare the effects of luminally administered and parenterally administered sodium salicylate upon chenodeoxycholic acid-induced colonic secretion. In our experiment, inoculation of four mM chenodeoxycholic acid into the rat cecum produced net secretion of water and sodium which was not reversed by preincubation of this bile acid with eight mM of sodium salicylate. Similarly, an intravenous bolus of either five mg of sodium salicylate per kg of body weight or 50 mg of sodium salicylate per kg of body weight failed to block salt or water secretion. Furthermore, 30 minute incubation of chenodeoxycholic acid with sodium salicylate produced neither reduction of in vitro aqueous bile acid concentration nor inhibition of ex vivo bile acid-facilitated hypotonic red cell hemolysis. These data suggest that sodium salicylate fails to sequester bile acids from aqueous solution and fails to block bile acid-mediated colonic secretion in the rat.     

Marijuana and immunity: tetrahydrocannabinol mediated inhibition of lymphocyte blastogenesis

The effects of delta-9-tetrahydrocannabinol (THC) and its major metabolite, 11-OH delta-9-tetrahydrocannabinol (11-OH THC) on mitogen driven lymphocyte blastogenic transformation (LBT) were studied. THC inhibited LBT responses to the T lymphocyte mitogens phytohemagglutinin and concanavalin A but not the B lymphocyte stimulant pokeweed mitogen. The metabolite 11-OH THC caused a comparable, but less significant, inhibition of LBT responses to the T cell mitogens. These inhibitions were dependent upon the drug doses and the time of incubation with the lymphocytes. There was no significant inhibitory activity of THC to the LBT when it was added 24 or 48 h after mitogen. In addition, exposure of lymphocytes to THC for 3 h, followed by removal of the drug prior to addition of mitogen had no effect on the cells' ability to respond to the mitogen. Thus, there appears to be a specified temporal period during which exposure of lymphocytes to THC results in an inhibition of blastogenesis. Interleukin 2 (30 units/ml) could not preclude the THC induced inhibition of lymphocyte blastogenesis. We conclude that THC and 11-OH THC inhibit T lymphocyte blastogenesis. However, unlike the THC mediated inhibition of natural killer cell activity (as shown previously), the process is not responsive to the cytokine interleukin 2.     

Production of macrophage-, granulocyte-, granulocyte-macrophage- and multi-colony-stimulating factor by peripheral blood cells

The specific cell sources and signals for induction of various colony-stimulating factors (CSF) in peripheral blood mononuclear cells (PBMC), purified T lymphocyte and monocyte (Mo) populations have been investigated. In the absence of exogenous activating stimuli, human PBMC, T cells and Mo failed to produce stable cytoplasmic mRNA for CSF for macrophages (M-CSF or CSF-1), for granulocytes (G-CSF), for granulocytes and macrophages (GM-CSF) and for multilineage CSF [multi-CSF, interleukin (IL) 3] and thus failed to release CSF proteins. However, after stimulation with phorbol myristate acetate and phytohemagglutinin, M-, G-, GM- and multi-CSF mRNA became detectable in PBMC, resulting in the secretion of the respective proteins. Identical culture conditions resulted in synthesis of only G- and M-CSF by purified Mo, whereas purified T lymphocytes produced GM-CSF and multi-CSF only. When Mo or T lymphocytes were exposed to recombinant human interferon-gamma or were stimulated by triggering the epitopes recognized by the monoclonal antibodies anti-Tll2 and Tll3, respectively, again disparate CSF expression patterns were found to be associated with both cell species. Moreover, IL2-receptive T lymphocytes showed the same distinct pattern of CSF secretion when activated by recombinant human IL2.     

Antibodies reactive with the B1 molecule inhibit cell cycle progression but not activation of human B lymphocytes

The B1 cell surface molecule (CD20) is a 35-kDa phosphoprotein expressed by B lineage cells during most stages of differentiation. Some monoclonal antibodies reactive with B1 induce activation while others, anti-B1a, inhibit B lymphocyte function. To further determine the requirement of B1 molecule function in proliferation and differentiation the effects of anti-B1a antibody binding on early cellular activation events were examined. Immunoglobulin secretion of lymphocyte cultures stimulated with pokeweed mitogen was maximally inhibited if the antibody (1-10 micrograms/ml) was added during the first 24 h of culture. However, even high antibody concentrations were unable to inhibit increases in free intracellular Ca2+ concentrations immediately following cross-linkage of cell surface immunoglobulin, or inhibit cell enlargement and the expression of transferrin and interleukin 2 receptors. Antibody binding to B1 inhibited RNA synthesis (37-80%) and progression through cell cycle following activation. In contrast, proliferation induced by phorbol myristate acetate was not inhibited by antibody binding to the B1 molecule. The findings that the earliest activation events and phorbol myristate acetate-induced proliferation were not inhibited by antibody binding to B1 suggest that inhibition is due to the blocking of a step of the activation process required for cell cycle progression and differentiation, rather than blocking initial signal transduction across the membrane or providing a negative or suppressive signal.     

Effect of isoprinosine on interleukin 1 and 2 production and on suppressor cell activity in pokeweed mitogen stimulated cultures of B and T cells

Isoprinosine appeared to potentiate the production of interleukin 1 and interleukin 2 in cultures of lipopolysaccharide stimulated human monocytes and phytohemagglutinin stimulated cultures of blood mononuclear cells respectively at pharmacological drug levels. Administration of the drug in vivo was associated with increased activity of radiation sensitive suppressor T cell activity against immunoglobulin production in pokeweed mitogen stimulated cultures of B and T cells. Addition of isoprinosine to the latter cultures in vitro appeared to enhance immunoglobulin production consistent with inhibition of suppressor cell activity or stimulation of helper activity. It is not clear from these studies whether the contrasting effects of the drug in vitro and in vivo represent different actions of metabolites or alteration of the proportion of lymphocyte subsets in the circulation. Further studies are required to answer these questions and to determine whether the changes persist with long term administration of the drug.     

Enhancement of bovine mammary mononuclear cell proliferation by autologous serum

Blood and mammary secretions were collected from involuted mammary glands of five primiparous Holstein cows to determine the effects of serum and mammary secretion skims on mononuclear cell proliferation. Proliferation was stimulated using concana‐valin A, phytohemagglutinin, pokeweed mitogen, and a mixed leukocyte assay. Autologous mammary secretion skims suppressed blood mononuclear cell proliferation. Suppression was reduced slightly when secretion was added 24 h following initiation of cultures. Addition of autologous serum enhanced mammary gland mononuclear cell proliferation markedly, in some cases equalling or exceeding responses of blood mononuclear cells from the same donors. Ability of serum to restore mammary mononuclear cell proliferation suggests that mammary mononuclear cells may be as competent as blood mononuclear cells if inhibitory effects of mammary secretion can be overcome.     

Expanded adipose-derived stem cells suppress mixed lymphocyte reaction by secretion of prostaglandin E2

Multipotent mesenchymal stem cells (MSCs) in adult tissue are known to be less immunogenic and immunosuppressive. Previous study showed that primary cultures of human adipose-derived stem cells (ADSCs) shared their immunomodulatory properties with other MSCs. However, whether passaged human ADSCs can retain their immunomodulatory effect after in vitro expansion remains unknown. In addition, the mechanism of ADSC-mediated immunomodulatory effect remains to be elucidated. This study aimed to investigate these issues by using passaged human ADSCs as an in vitro study model. Flow cytometry showed that passaged ADSCs expressed human leukocyte antigen (HLA) class I but not class II molecules, which could be induced to express to a high level with interferon-gamma (IFN-gamma) treatment. The study found that passaged ADSCs could not elicit lymphocyte proliferation after co-culturing with them, even after IFN-gamma treatment. In addition, either IFN-gamma-treated or non-treated ADSCs could inhibit phytohemagglutinin (PHA)-stimulated lymphocyte proliferation. Moreover, passaged ADSCs could serve as the third-party cells to inhibited two-way mixed lymphocyte reaction (MLR). Further study using a transwell system also showed that this type of immunosuppressive effect was not cell-cell contact dependent. In defining possible soluble factors, we found that passaged ADSCs significantly increased their secretion of prostaglandin E2 (PGE2), but not transforming growth factor-beta (TGF-beta) and hepatocyte growth factor (HGF), when they were co-cultured with MLR. Furthermore, the result demonstrated that only PGE2 production inhibitor indomethacine, but not TGF-beta- and HGF-neutralizing antibodies, could significantly counteract ADSC-mediated suppression on allogeneic lymphocyte proliferation. These results indicated that in vitro expanded ADSCs retain low immunogenicity and immunosuppressive effect, and PGE2 might be the major soluble factor involved in the in vitro inhibition of allogeneic lymphocyte reaction.     

Modulation in vitro of human natural cytotoxicity, lymphocyte proliferative response to mitogens and cytokine production by essential fatty acids.

Essential fatty acids (EFA) have been shown in animal studies to have a differential effect on various aspects of immune reactivity. However, there have been few studies in humans. Therefore, we elected to investigate the effects of a variety of EFA [gamma-linolenic acid (GLA), eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA)] in vitro on human blood lymphocyte reactivity, cytokine secretion and natural cytotoxicity. The proliferative response to polyclonal mitogens (phytohaemagglutinin, pokeweed mitogen, concanavalin A), as measured by [3H]thymidine incorporation into newly synthesized lymphocytes, was inhibited (P < 0.05) by all EFAs tested, in a dose-dependent manner (3-15 micrograms/ml). The greatest inhibition of proliferation was caused by EPA and DHA. Similarly, EPA, DHA and GLA significantly reduced cytotoxic activity [expressed as lytic units, using 51 chromium-release assays natural killer (NK) (K562 cells) and lymphokine-activated (LAK) (Daudi cells) cells] (P < 0.05) in a concentration-dependent manner (5-50 micrograms/ml), without affecting cell viability. EPA and DHA exhibited greater suppression than GLA. Furthermore, the inhibition of cell proliferation and suppression of natural cytotoxicity was associated with marked decrease in cytokine [interleukin-1 (IL-1), IL-2, tumour necrosis factor-alpha (TNF-alpha) and interferon-gamma (IFN-gamma)] production in vitro. Our findings demonstrate that EFAs (GLA, EPA, DHA) have the potential to inhibit significantly various aspects of human lymphocyte cell-mediated and humoral immune reactivities.     

Kinetics of inhibition of mitogen-induced proliferation of human lymphocytes by alpha 2-macroglobulin in serum-free medium

The effect of alpha 2-macroglobulin (alpha 2M) on the ability of human lymphocytes to proliferate in response to stimuli by 3 mitogens, concanavalin A, phytohaemagglutinin and pokeweed mitogen was investigated in in vitro lymphocyte cultures in serum-free medium. The following experiments were performed: 1. lymphocytes were treated with alpha 2M prior to stimulation with the mitogens; 2. alpha 2M and mitogens were added simultaneously to the lymphocyte cultures; and 3. alpha 2M was added to the lymphocyte cultures after they were stimulated with the mitogens. In all cases, alpha 2M was found to inhibit mitogen induced proliferation of the lymphocytes as evaluated by (6-3H)-thymidine uptake of the cells. The mechanisms involved in the inhibition of lymphocyte proliferation by alpha 2M are discussed.     

Impaired Mitogen-Induced Interferon-γ Production in Rheumatoid Arthritis and Related Diseases

Peripheral blood lymphocytes from patients with rheumatoid arthritis (RA), systemic lupus erythematosus (SLE), progressive systemic sclerosis (PSS), Reiter's disease, osteoarthritis, and from healthy volunteers were investigated for interferon-gamma (IFN-gamma) production after mitogen activation. Phytohaemagglutinin stimulation revealed an impaired IFN-gamma production in RA, SLE, and PSS but normal levels in Reiter's disease and osteoarthritis. In RA this deficiency was also seen after pokeweed mitogen, OKT3, and concanavalin A activation. No major differences were found in interleukin 2 (IL-2) production and cell proliferation. The IL-2 receptor expression was reduced on stimulated RA lymphocytes. The deficient IFN-gamma production was compensated in RA by co-stimulation of PHA or OKT3 with phorbol myristic acetate (PMA). In addition, the combination of the calcium ionophore A 23187 and PMA induced a strong IFN-gamma secretion in all patient groups and in the controls.     

CD8+CD11b+ peripheral blood T lymphocytes contain lymphokine-activated killer cell precursors

CD3+CD8+ cells, purified from peripheral blood T cells by depletion of CD4+ lymphocytes, were tested for their cytotoxic activity against K-562 or HL-60 cells. Freshly prepared cells had no cytotoxic function, but upon exposure to recombinant interleukin 2 (rIL2) in vitro they acquired a lymphokine-activated killer (LAK) activity. Fractionation of CD3+CD8+ cells into CD11b+ and CD11b- cells demonstrated that all the cells with the potential of developing LAK cell functions were within the CD8+CD11b+ subset. These cells lacked natural killer cell markers such as CD16 or NKH1, but a proportion of them stained for Leu-7. Furthermore, they expressed the alpha/beta chains, but not the gamma/delta chains, of the T cell receptor, as could be determined by staining with the appropriate monoclonal antibodies. CD8+CD11b+ cells had a large granular lymphocyte morphology, as shown by both cytochemical and electron microscopy analyses. They proliferated in response to IL2 in vitro and developed cytotoxic functions against a number of natural killer resistant targets. Their response to phytohemagglutinin or pokeweed mitogen was very weak or absent. By contrast, CD8+CD11b- cells failed to generate LAK cells in response to rIL2, did not show a large granular lymphocyte morphology, but responded vigorously to phytohemagglutinin or pokeweed mitogen. Purified CD8+CD11b+ cells were cloned by limiting dilution in the presence of rIL2. The observed cloning efficiency of 19 +/- 0.3% indicates that a fraction of the cells only could respond to IL2. Furthermore, only 50% of the clones obtained had a LAK cell function. These data show that CD8+CD11b+ cells represent a heterogeneous cell population. Nevertheless this cell subset probably represents the major source of LAK cell progenitors within the circulating T cells.     

Inhibition of in vitro human lymphocyte response by the pneumococcal toxin pneumolysin

The effects of pneumolysin, a sulfhydryl-activated cytolytic toxin produced by Streptococcus pneumoniae, on the in vitro human lymphocyte response was examined. The toxin, at concentrations of one to five hemolytic units per ml, caused marked inhibition of the response of lymphocytes to concanavalin A, phytohemagglutinin, pokeweed mitogen, and protein A. The response was assessed by measuring both [3H]thymidine incorporation and the ability of lymphocytes to produce immunoglobulins and lymphokine activity. The effects of pneumolysin were irreversible, could be prevented by pretreatment of the toxin with cholesterol, and were not related to a direct cytotoxic effect on the lymphocytes. Pneumolysin appeared to act at the initiation phase of the immune response and had no effect on lymphocytes committed to DNA synthesis or to the synthesis and secretion of immunoglobulins. Furthermore, pneumolysin-mediated inhibition of the lymphocyte response was not due to the inhibition of binding of mitogens to leukocytes and is likely to be related to effects on membrane-mediated signals essential for lymphocyte triggering. This may be one means by which pneumolysin plays a role in the pathogenesis of pneumococcal infections.     

Recombinant interleukin 2 inhibits pokeweed mitogen-induced proliferation of human adult peripheral blood and cord blood mononuclear cells

We studied the effects of recombinant interleukin 2 (rIL-2) on pokeweed mitogen (PWM)-induced proliferation of unfractionated human mononuclear cells from the peripheral blood (PBMCs) and cord blood (CBMCs). rIL-2 was found to inhibit PWM-induced proliferation of both PBMCs and CBMCs in a dose-dependent manner. The response was statistically significant at concentration 10 U/ml. The inhibition of adult PBMC proliferation was relatively mild, but in newborns up to 75% inhibition was found. The inhibitory effect of rIL-2 was best detectable at day 4, at the same time as PWM-induced proliferation peaked. However, also at days 8 and 10, the response induced by PWM plus rIL-2 was much lower than that induced by rIL-2 alone. These finding suggest that the inhibition was due to rIL-2-mediated stimulation of the function of PWM-activated suppressor cells. CD8+ T cells were, however, not alone responsible for the rIL-2-induced inhibition of PWM-induced proliferation, since the responses were essentially similar independent of whether CD8+ T cells were present or absent. In addition, the release of interferon-gamma (IFN-gamma) in response to rIL-2 was not a major cause of the inhibitory effects, because rIL-2 inhibited PWM-induced proliferation of CBMCs, which are deficient in producing IFN-gamma. Our results provide novel data to the ongoing discussion of IL-2 as a down-regulator of immune functions.     

Effects of immune activation on quinolinic acid and neuroactive kynurenines in the mouse.

Accumulation of quinolinic acid and neuroactive kynurenines derived from tryptophan are of potential significance in human neuropathologic diseases because of their neurotoxic and convulsant properties. Clinical studies have established that sustained elevations of quinolinic acid, L-kynurenine and kynurenic acid within the cerebrospinal fluid occur in patients with a broad spectrum of inflammatory diseases and correlate with markers of immune activation and interferon-gamma activity. The present study describes an animal model that replicates these clinical observations and investigates the role of interferon-gamma as a mediator between immune activation and increased kynurenine pathway metabolism. Marked elevations in quinolinic acid, L-kynurenine and 3-hydroxykynurenine as well as an increased ratio of quinolinic acid: kynurenic acid in brain occurred 24 h after systemic pokeweed mitogen administration to C57BL6 mice. In plasma, L-tryptophan and kynurenic acid levels were reduced by pokeweed mitogen, while the concentrations of L-kynurenine, 3-hydroxykynurenine and quinolinic acid were increased. Interferon-gamma, pokeweed mitogen and lipopolysaccharide induced indoleamine-2,3-dioxygenase, the first enzyme of the kynurenine pathway, and increased both L-kynurenine and quinolinic acid concentrations of brain and systemic tissues, particularly in the lung, gastrointestinal tract and spleen. In contrast, hepatic tryptophan-2,3-dioxygenase activity was either reduced or unaffected. Increases in kynurenine pathway metabolism were sustained in mice given daily injections of interferon-gamma for seven days and subsequent responses to interferon-gamma were further enhanced. In contrast, daily administration of lipopolysaccharide was associated with subsequent attenuated responsiveness (tolerance) to lipopolysaccharide, pokeweed mitogen and interferon-gamma. Systemic administration of a monoclonal antibody to mouse interferon-gamma either attenuated or abolished the responses of kynurenine pathway metabolism to pokeweed mitogen and interferon-gamma. We conclude that acute and chronic increases in quinolinic acid and neuroactive kynurenines follow immune stimulation in mice, and result from indoleamine-2,3-dioxygenase induction. The results demonstrate that interferon-gamma is an important mediator between immune stimulation and indoleamine-2,3-dioxygenase induction. These increases in kynurenine pathway metabolism closely parallel the responses documented in patients with a broad spectrum of inflammatory diseases. Mice treated with immune stimuli are a useful model to investigate the relationships between immune activation and kynurenine pathway metabolism.     

Effect of interleukin-2 and methylprednisolone on in vitro transformation of uremic lymphocytes

The functional relationship in vitro between mitogen-induced lymphocyte transformation, lymphocyte response to interleukin-2 (IL-2) and steroid, and production of IL-2 was examined in patients with chronic renal failure on hemodialysis (HD) or on continuous ambulatory peritoneal dialysis (CAPD). The lymphocyte responses to optimal stimulation with phytohemagglutinin, concanavalin A, and pokeweed mitogen were depressed in lymphocyte cultures from HD patients, while CAPD lymphocyte cultures responded normally. However, at suboptimal phytohemagglutinin stimulation both CAPD lymphocyte and HD lymphocyte responses were subnormal. Uremic lymphocyte cultures were more sensitive to the immunosuppressive effect of methylprednisolone. Addition of IL-2 normalized the phytohemagglutinin responses of suboptimally stimulated CAPD lymphocyte cultures and clearly improved the mitogen responses of the HD lymphocyte cultures. Furthermore, the increased uremic lymphocyte sensitivity to methylprednisolone was normalized by addition of IL-2 to the cultures. The measured IL-2 production had clearly decreased in the HD cultures after 48 h as compared to that of the control cultures. A similar but not significant trend was also seen in the CAPD cultures. Thus, it is suggested that a deficient production of IL-2 may partly explain the reduced lymphocyte response of uremic lymphocytes in vitro.     

Response of mononuclear cells to recombinant bovine interleukin‐1β during the non‐lactating period

Interleukin‐1 has been shown to play a critical role in the initial stages of lymphocyte proliferation. Recombinant bovine interleukin‐1 may be useful as an immunoenhancer of bovine mammary gland mononuclear cells to enhance resistance of the mammary gland to infection during the non‐lactating period. The objective of this study was to determine if recombinant bovine interleukin‐1β influenced proliferation of blood and mammary gland mononuclear cells obtained during the non‐lactating period. Bovine mononuclear cells were isolated from five primiparous Holstein cows at 14–18 and 28–32 days of involution, and 7–13 days prior to parturition. Proliferation of mononuclear cells in response to recombinant bovine interleukin‐1β was evaluated in the presence and absence of suboptimal concentrations of concanavalin A and pokeweed mitogen. Varying concentrations (0.78–25 ng ml^?1) of recombinant bovine interleukin‐1β did not influence blood or mammary gland mononuclear cell proliferation in the presence or absence of mitogens at any of the time periods studied. Data suggest that exogenous recombinant bovine interleukin‐1β may not be effective as an immunoenhancer of existent mammary gland mononuclear cell populations.     

Anti-CD27 monoclonal antibodies identify two functionally distinct subpopulations within the CD4+ T cell subset

Anti-CD27 monoclonal antibodies react with a cell surface molecule expressed on medullary thymocytes and a large subpopulation (75%) of peripheral blood T lymphocytes. This study was undertaken to analyze the functional capacities of CD27+ and CD27- subpopulations within the CD4+ subset. In addition, we investigated whether CD27 subpopulations belong to two mutually exclusive T cell sublineages. Proliferation upon lectin stimulation by either phytohemagglutinin or pokeweed mitogen (PWM) was found to be consistently higher in CD4+CD27+ cells compared to CD4+CD27- cells. In contrast, CD27+ and CD27- cells did not differ in anti-CD3, soluble antigen or interleukin (IL)2-induced proliferation. In PWM-driven B cell differentiation, CD4+CD27+ cells provided helper activity on IgM production, while CD4+CD27- cells did not. The differences observed between CD4+CD27+ and CD4+CD27- cells in both proliferation and T helper activity on IgM production can, at least in part, be explained by the inadequate production of IL2 by CD27- cells upon lectin stimulation. In contrast, CD27+ and CD27- subpopulations did not differ in the production of interferon-gamma. CD27- cells could be induced to express the CD27 antigen through stimulation of these cells with immobilized anti-CD3 antibodies. After 3 days of culture, approximately 50% of the cells had CD27 membrane expression, whereas after 6 days 80% of the cells express the antigen. We conclude that anti-CD27 antibodies identify two functionally distinct T cell populations within the CD4+ subset. These subpopulations apparently do not belong to two separate T cell sublineages, but may reflect differences in the activation state of CD27+ vs. CD27- peripheral blood T lymphocytes.     

Involvement of fibronectin and its receptor in human lymphocyte proliferation.

Adhesion between lymphocytes and antigen-presenting cells is necessary for the development of certain immune reactions. We have previously shown that fibronectin (FN) added to mixed lymphocyte cultures (MLC) can restore a decreased lymphocyte proliferation in immunocompromised individuals. Using highly purified cell populations from peripheral blood for depletion and adding back experiments we show here that exogenous FN enhanced proliferation only when allogeneic monocytes were co-cultured with responder lymphocytes. Although lymphocyte proliferation in MLC was augmented by FN, there was no preferential proliferation of any particular major lymphocyte subpopulation in cultures supplemented with FN as compared to control cultures lacking its addition. Antibody against the FN receptor (FN-R) of the beta 1 integrin family, as well as Arg-Gly-Asp containing peptide, could inhibit alloantigen-induced lymphocyte proliferation in a concentration-dependent manner. Anti-CD3-induced proliferation was inhibited by anti-FN-R antibody but not Arg-Gly-Asp peptide whereas no inhibition was seen with the phytohemagglutinin (PHA)-induced lymphocyte proliferation. This study presents further evidence that FN and its receptor (alpha 5 beta 1) are involved in the augmentation of T-cell responsiveness to proliferative stimuli.     

The inhibition of T-lymphocyte proliferation by fatty acids is via an eicosanoid-independent mechanism.

Eicosanoids, in particular prostaglandin E2 (PGE2), are potent inhibitors of a number of immune responses, including lymphocyte proliferation. We have previously shown that fatty acids, especially polyunsaturated fatty acids (PUFA), inhibit mitogen-stimulated proliferation of lymphocytes. One mechanism by which fatty acids could exert their inhibitory effect is via modulation of eicosanoid synthesis. This possibility was investigated in the present study. PGE2 concentrations in the medium taken from lymphocytes cultured in the presence of a range of different fatty acids did not correlate with the inhibitory effects of the fatty acids upon lymphocyte proliferation. Although PGE2 at concentrations above 10 nM caused inhibition of lymphocyte proliferation, PGE2 at the concentration measured in lymphocyte culture medium (0.3-4 nM) was not inhibitory. PGE3 did not inhibit lymphocyte proliferation, except at high concentrations (greater than 250 nM). The maximal inhibition of proliferation caused by PGE2 or PGE3 was less than the inhibition caused by each of the fatty acids except myristic or palmitic acids. Inclusion of inhibitors of phospholipase A2, cyclo-oxygenase or lipoxygenase in the culture medium did not prevent the fatty acids from exerting their inhibitory effect on lymphocyte proliferation. The eicosanoids present in lymph node cell cultures originate from macrophages rather than lymphocytes. Depletion of macrophages from the cell preparation by adherence did not prevent fatty acids from inhibiting proliferation. Proliferation of thoracic duct lymphocytes, which are devoid of macrophages, is inhibited by fatty acids to a similar extent as proliferation of lymph node lymphocytes. These observations provide convincing evidence that the inhibition of lymphocyte proliferation by fatty acids is independent of the production of eicosanoids. Therefore, other mechanisms must be investigated if the effect of fatty acids upon lymphocyte proliferation is to be understood at a biochemical level.     

Inhibition of human peripheral blood mononuclear cell proliferative response by glycosphingolipids from metacestodes of Echinococcus multilocularis.

The effect on human peripheral blood mononuclear cells (PBMCs) of neutral glycosphingolipids extracted from metacestodes of the parasite Echinococcus multilocularis was investigated. Neutral glycosphingolipids inhibited [3H]thymidine uptake by human PBMCs upon stimulation by mitogens such as phytohemagglutinin A and pokeweed mitogen or by allogeneic Burkitt B cells. This effect was dose dependent and was related to a decrease in interleukin 2 (IL-2) synthesis, the expression of IL-2 receptors (CD25) being unmodified. Addition of exogenous recombinant IL-2 restored the cell proliferation. Partial inhibition of immunoglobulin G (IgG), IgA, and IgM synthesis was observed in the supernatant of cell culture in association with the inhibitory effect. Identification of active subfractions contained in the neutral glycosphingolipid fraction was also studied in relation to cell viability. The free ceramide fraction had an inhibitory effect, in part related to cell lysis, particularly at high concentration, while the monogalactosylceramides had a paradoxical effect: as an activator at low concentrations and as an inhibitor at high concentrations, with limited cell survival. The immunogenic neutral glycosphingolipids containing at least two carbohydrate residues, all having a structure based on Gal beta 1-->6Gal, were inhibitors of PBMC proliferation and showed good cell survival. These results suggest that parasite neutral glycosphingolipids may play an immunologically relevant role in alveolar hydatid disease.