Influence of serum from atopic children on T lymphocytes.

The relative and absolute number of peripheral T cells was found to be depressed in atopic children. Sera from atopic children had slightly less stimulatory effects on lymphocyte DNA synthesis induced by PHA, Con A and PPD, than sera from nonatopic children. This finding indicates an occurrence of inhibitory factor(s) in atopic serum. Sera from some severely ill atopic patients almost completely abolished mitogen-induced lymphocyte DNA synthesis. Inhibition by atopic serum appeared to be an early event in lymphocyte mitogenesis and not due to CRP, IgE or factors binding to mitogens. Lymphocytes from atopic children were no more sensitive to suppressive influences of atopic serum factors than were lymphocytes from adult blood donors. Normal serum enhanced mitogen-induced DNA synthesis more in lymphocytes from adult blood donors than in those from atopic children. The results indicate that, although the T cell defect in atopy may be partly caused by serum factors, occurring during clinical allergic disease, the main reason for the defect is probably an altered reactivity of the T lymphocytes in atopic individuals.     

Stimulatory and inhibitory actions of VIP and cyclic AMP on cytoplasmic Ca2+ signal generation in pancreatic acinar cells

In pancreatic acinar cells the effects of vasoactive intestinal polypeptide (VIP) and dibutyryl cyclic AMP (dbcAMP) alone and during stimulation with acetylcholine (ACh) or internally applied inositol trisphosphate (InsP₃) were investigated utilizing the patch-clamp whole-cell current recording configuration to assess changes in cytoplasmic Ca²⁺ concentration by measurement of Ca²⁺ dependent Cl^– current. VIP (1 nM) and dbcAMP (0.5 mM) each evoked repetitive pulses of Ca²⁺-dependent Cl^– current. A high VIP concentration (100 nM) acutely and reversibly inhibited the responses evoked by 1 nM VIP and also acutely inhibited ACh-evoked responses. DbcAMP (2 mM) also reversibly inhibited the ACh-evoked responses. Neither VIP nor dbcAMP were able to inhibit InsP₃-evoked repetitive Ca²⁺ pulses. VIP in a low concentration can generate cytosolic Ca²⁺ oscillations via cyclic AMP, but high VIP concentrations inhibit both ACh and VIP-evoked Ca²⁺ signals and this effect is mediated by high intracellular cyclic AMP levels.     

T lymphocytes in atopic children.

The number of circulating T and B cells and the sensitivity of lymphocytes to stimulation with phytohemagglutinin (PHA), concanavalin A (Con A) and pokeweek mitogen (PWM) was studied in 233 atopic children. The number of T lymphocytes was found to be decreased in cases of rhinoconjunctivitis, asthma as well as atopic eczema. Levels of B lymphocytes were normal. Sensitivity to stimulation with PHA and to a lesser degree, Con A, was significantly decreased whereas stimulation with PWM was unaffected. The severity of the atopic eczema was inversely correlated to T cell numbers. Several lines of evidence indicated that the abnormalities observed were intrinsically associated with the atopic conditions and not evoked by corticosteroid treatment. The results are compatible with the hypothesis that atopy is associated with a defect of a subpopulation of T cells. The possibility that this subpopulation has a suppressor function on reagin formation is discussed.     

The antagonistic action of cyclic GMP and cyclic AMP on proliferation of B and T lymphocytes.

The effect of c-AMP, c-GMP and both substances together on (3H)-thymidine incorporation into nuclear DNA was investigated using spleen cells of normal and athymic nude mice. c-GMP induces DNA synthesis in both normal and nude spleen cell populations. c-AMP inhibited the stimulatory activity of c-GMP as well as the phytohaemagglutinin (PHA) and lipopolysaccharide (LPS) response of spleen cells. The inhibitory activity of c-AMP on the PHA and LPS responses can be reversed by c-GMP. The possible role of cyclic nucleotides in the regulation of cell proliferation and of the immune response is discussed.     

Effects of platelet activating factor on cyclic AMP accumulation in human lymphocytes.

The in vitro effects of platelet activating factor (PAF) on intracellular adenosine 3',5'-cyclic adenosine monophosphate (cAMP) accumulation were assessed in human lymphocytes in ten healthy volunteers and in eight asthmatic patients. Platelet activating factor to increased intracellular cAMP in both groups. In addition, PAF also enhanced the effects of isoproterenol and theophylline on intracellular cAMP production in these two groups.     

The effects of cyclic AMP on leucocyte inhibitory factor (LIF) production and on the inhibition of leucocyte migration.

The effect of drugs known to increase intracellular levels of cyllic AMP were studied in the leucocyte migration ihibition system. It was found that cyclic AMP, dibutyryl cyclic AMP, theophyline, and prostaglandins E1 and E2 inhibited the production of leucocyte inhibiting factor by HA pulsed lymphocytes Inhibition only occured when the drugs were present during or after the PHA pulse. In addition it was found that these drugs enhanced the migration of polymorphonuclear leucocytes (PMN), in this system. Electrophoretic mobility of PMN cells was not altered by these drug indicating that the effect is not due to changes in membrane charge. However, granulocyte adhesion was reduced in the presence of these drug suggesting that adhesion is of primary importance in the migration of polymorphonuclear leucocytes out of capillary tubes. The findings show that cyclic AMP is important in modulating both cell-mediated and inflammatory responses.     

The effects of uremic serum and 3'-5' cyclic AMP on blastogenesis of normal lymphocytes

Phytohemagglutinin (PHA) induced lymphocyte blast transformation is impaired both in uremic lymphocytes and in normal lymphocytes exposed to uremic serum. Cyclic AMP is known to inhibit blast transformation in normal and uremic lymphocytes. This investigation was undertaken to assess quantitatively the effects of uremic serum and cyclic AMP on blastogenesis of normal lymphocytes. Uremic serum or cyclic AMP significantly inhibited blast transformation of normal lymphocytes. These effects were statistically similar and cumulative. We conclude that the inhibition imposed by uremic serum on normal lymphocyte blastogenesis is predominantly mediated by a mechanism(s) different from cyclic AMP.     

Pertussis toxin blocks the inhibitory effects of calcitonin on cyclic AMP accumulation in stimulated cultured human monocytes

Surface stimulation of fresh or cultured human mononuclear cells by latex particles causes an increase in the accumulation of cyclic AMP that is inhibited by preincubation with calcitonin (CT). Preincubation of cultured monocytes with 500 ng/ml pertussis toxin totally blocks the inhibitory effects of CT at low concentrations of this hormone. The effects of pertussis toxin are dose-related and eliminated by boiling the toxin. Similar preincubations with cholera toxin have no significant effects on subsequent inhibition of surface-stimulated cyclic AMP by CT. Membranes prepared from cultured human monocytes contain a 41,000-dalton protein that is ADP-ribosylated by pertussis toxin and may be the inhibitory guanine nucleotide regulatory protein (Ni) mediating this inhibition.     

Spontaneous expression of IL-4 mRNA in lymphocytes from children with atopic dermatitis.

Normal lymphocytes do not generally produce or secrete lymphokines in the resting or unstimulated state and only express or release cytokines following activation. Recently, the spontaneous production of intracellular interferon-gamma (IFN-gamma) and spontaneous secretion of IL-6 has been documented in patients with atopic dermatitis. These findings indicated that lymphocytes had been previously activated in vivo. Such in vivo activation may also be associated with spontaneous production of IL-4. As measurement of IL-4 secretion by immunoassay is complicated by poor sensitivity, and only provides information on the net amount of cytokine present after secretion, adsorption, consumption and degradation have occurred, IL-4 mRNA expression in peripheral blood lymphocytes from children with atopic dermatitis and controls was examined by polymerase chain reaction (PCR)-assisted mRNA amplification. Spontaneous expression of IL-4 mRNA was detected in four of eight patients with severe atopic dermatitis. Following stimulation in vitro, seven of eight atopic patients demonstrated detectable IL-4 mRNA. In comparison, no spontaneous expression of IL-4 mRNA was found in controls, and only six of 10 controls expressed IL-4 mRNA in stimulated cultures. The spontaneous expression of IL-4 mRNA in unstimulated cultures from children with atopic dermatitis supports the possibility that previous in vivo activation has occurred, and suggests that IL-4 production is increased in vivo in atopic dermatitis. This in vivo activation together with the constitutive expression of IL-4 mRNA are likely to contribute to the spontaneous in vitro production of IgE in atopic patients.     

Stimulatory effect of lymphocytes from Chagas'' patients on spontaneously beating rat atria.

The aim of this work was to study the effect of lymphocytes from individuals infected with Trypanosoma cruzi (Chagas' patients) on the contractile behaviour of living heart tissue. Chagas' lymphocytes (ChL) reacted with isolated rat atria preparations increasing the isometric development tension (IDT) and frequency of contractions (FC) in a dose-dependent manner. The maximal stimulatory effect was reached after 30-40 min of contract. In contrast, normal lymphocytes (NL) did not alter the basal IDT and FC values. beta-adrenergic antagonists, anti-histamine agents and inhibitors of the synthesis and action of arachidonic acid (AA) products were used to study the mechanisms of the reaction. (-)-propranolol (10(-7)M) and pyrilamine (10(-6)M) had no effect ruling out the participation of beta-adrenergic agonists or histamine. However, indomethacin (10(-6)M) and acetylsalicylic acid (1.8 X 10(-4)M) enhanced the effect of ChL. Inhibitors of the lipoxygenase pathway (5,8,11,14-eicosatetraynoic acid, 10(-7)M; nordihydroguairetic acid, 10(-5)M) and FPL55712, an antagonist of one of its terminal products: the slow reacting substance of anaphilaxis (SRS-A), abolished the reaction. Therefore, a fundamental role for SRS-A in the production of the stimulatory effect is postulated. Lymphocytes of the T cell lineage (E rosette forming cells, ERFC) are the effector cells involved in this reaction, whereas non-rosetting ChL depressed IDT. T ascertain if effector cells could be replaced by soluble factors, ChL were reacted with homogenates of rat atria and the cell free supernatants were added to beating rat atria. Positive ino- and chronotropic effects were obtained, indicating that soluble factors generated during the reaction can substitute for the intact effector cells. On the other hand if the effector cells were purified from Chagas' patients that had been treated 1 month to 6 years before the assay with trypanocidal drugs (3-methyl-4-(5'-nitrofurfurylidene-amino)-tetrahydro-4H-1, 4-tiazine-1, 1-dioxide, nifurtimox or N-benzyl-2-nitro-imidazolacetamide, benznidazole) only depressor effects were found. The depressor inotropic action of lymphocytes from treated patients (tr-ChL) was abolished with indomethacin and acetyl salicylic acid indicating that products of the cyclooxygenase pathway of AA were involved. While this work provides additional evidence for the hypothesis that lymphocytes from T. cruzi infected patients may react with heart tissue and alter its contractile behaviour, the results should not be extrapolated to the in vivo situation.(ABSTRACT TRUNCATED AT 400 WORDS)     

Cyclic AMP and cyclic GMP phosphodiesterase activities in Hodgkin's disease lymphocytes

Cyclic nucleotide phosphodiesterase (PDE) activities were studied in peripheral blood monocyte-depleted lymphocytes and enriched T-lymphocyte suspensions from thirteen patients with previously untreated Hodgkin's disease (HD) and fourteen age and sex matched healthy volunteers. Monocyte-depleted lymphocytes from HD patients showed PDE-activities which were two times higher than in their normal counterpart cells. The mean cAMP-PDE activity present in enriched HD T-lymphocyte suspensions was four times higher than in control T-lymphocytes, and the mean cGMP-PDE associated with HD T-lymphocytes was three times higher than in the controls. The hydrolytic activities present in both monocyte-depleted and T-lymphocyte enriched cells suspensions remained unchanged in absence or in the presence of calmodulin and calcium. Since depressed cAMP and cGMP resting levels have been observed in HD lymphocytes and lymphocyte subpopulations, our results suggest that the elevated PDE activities are, at least in part, responsible for the alterations in lymphocyte cyclic nucleotide levels.     

The effect of nerve growth factor on DNA synthesis, cyclic AMP and cyclic GMP accumulation by mouse spleen lymphocytes.

Nerve growth factor (NGF), a trophic neuropeptide, is known to stimulate development, and to be important in the maintenance and survival of sympathetic and sensory neurons. Considering the presence of specific receptors on the surface of spleen cells, the effect of 2.5s nerve growth factor on 3H-thymidine uptake, cAMP and cGMP accumulation in mouse spleen lymphocytes has been studied. It was found that NGF added in vitro at the concentrations between 4 x 10(-7) and 4 x 10(-8) M significantly inhibited the incorporation of 3H-thymidine into lymphocytes DNA and increased cAMP levels in a dose-dependent manner but had no effect on cGMP levels. The maximal stimulation of cAMP synthesis occurred between 5 and 30 min after the NGF addition to the culture medium. When NGF was administered in vivo a significant dose-dependent inhibition of the lymphocytes proliferation was observed. These results indicate that an early increase of cAMP concentration is responsible for the antiproliferative action of NGF on mouse spleen lymphocytes and suggest that NGF could play an important role in the regulation of immune system function.     

Control of immune responses by cyclic AMP and lymphocytes that adhere to histamine columns.

Mixed lymphocytes from human peripheral blood, murine spleens, lymph nodes or thymus glands have pharmacologically specific receptors for histamine, beta mimetic catecholamines and prostaglandins. When these cells are exposed to the panoply of drugs mentioned above, their intracellular cyclic AMP concentrations increase. The biologic consequences of such an increase were at first elusive. Now we know that the immune potential of some murine spleen cells may be modulated and the release of lysosomal enzymes and histamine from human leukocytes may be inhibited. This paper concentrates on the effects that manipulation of cells with amine receptors has on their immune function. Recent studies have revealed that a subpopulation of splenic suppressor T cells responds to increases in its cyclic AMP content by reversing its suppressive effects on the humoral antibody response. When these T cells are removed from the murine cell population by their differential adherence to insolubilized conjugates of histamine with albumin, the remainder of the cells are more responsive to sheep cell antigen, as tested by transferring the spleen cells together with the antigen into lethally irradiated recipient animals. The suppressor T cells that adhere to the insolubilized conjugates of histamine-albumin (called histamine-rabbit serum albumin-Sepharose, or HRS) are Ia positive, they appear to have receptors for histamine, beta adrenergic amines and prostaglandins of the E series, and when stimulated by these agents their in vivo and in vitro suppressor actions are reversed. The reversal seems quantitatively dependent on cyclic AMP accumulation. Receptors for the amines and prostaglandins are found on the T cell precursors of cell mediated immunity. They develop on some T effector cells in selected models of allogeneic target cell lysis. The receptors also appear to develop on selected B cells once these cells become committed to antibody production. The distribution of receptors on all leukocytes has not been adequately studied nor has their full potential in the immune response been studied in detail.     

Selective regulation of antigen-specific IgE response by cyclic AMP level in murine lymphocytes.

We have reported that prostaglandin E2 (PGE2) is a selective stimulator of the antigen-specific IgE response [6]. Because PGE2 is known to elevate intracellular cAMP, we investigated the regulatory role of cAMP in the production of antigen-specific IgE. Anti-TNP IgE response was induced by stimulating TNP-KLH-primed BALB/c spleen cells with the same antigen in vitro. Addition of 10-100 microM dibutyryl cAMP (DBcAMP) to the lymphocyte culture resulted in a 2-3-fold increase in anti-TNP IgE response without affecting the production of anti-TNP IgG1 or IgM. Forskolin, a stimulator of adenylate cyclase, also specifically augmented the IgE response. In contrast, 2',5'-dideoxyadenosine, an inhibitor of adenylate cyclase, suppressed IgE production in an isotype-specific manner. These results suggest that IgE synthesis can be selectively modulated by intracellular cAMP level. Enhancement of IgE production by DBcAMP was observed, particularly in highly primed spleen cells, suggesting that IgE-committed B cells are subjected to regulation by cAMP.     

Catecholamine effects on cyclic AMP levels and ion secretion in rabbit ileal mucosa.

Effects of catecholamines on cyclic AMP (cAMP) levels and ion fluxes were examined in isolated rabbit ileal mucosa. The base-line cAMP level was unaffected by epinephrine (Epi), norepinephrine (Norepi), and isoproterenol. The theophylline-augmented cAMP level was decreased slightly be Epi in one series of experiments but not in another. Propranolol did not enhance this effect. The increase in cAMP level produced by cholera toxin was almost completely reversed by addition of Epi or Norepi. This reversal was prevented by phenoxybenzamine. Epi also partially reversed the increase in cAMP level produced by prostaglandin E1. Effects of Epi on ion fluxes were determined following addition of secretagogues. Epi significantly decreased theophylline-induced but not cAMP or cholera toxin-induced Cl secretion. A decrease in short-circuit current was nonetheless observed in the latter two instances. The observed discrepancies between alpha-adrenergic effects on cAMP levels and ion fluxes suggest the following possibilities: 1) ion transport-related cAMP is only a small fraction of total mucosal cAMP; 2) cAMP-induced active ion secretion is only slowly reversible, or 3) effects of alpha-adrenergic stimuli on ion transport are not due to inhibition of cAMP accumulation.     

Differences in effect of isoproterenol stimulation on levels of cyclic AMP in human B and T lymphocytes.

The level of cyclic AMP (cAMP) in human lymphocytes in the presence or absence of isoproterenol stimulation was studied in various lymphocyte subpopulations containing different proportions of B and T lymphocytes. Lymphocytes from thymus and peripheral blood (which contain a majority of T cells) were more sensitive to isoproterenol action than lymphocytes from tonsils or adenoids (where B cells are predominant). Using purified B or T lymphocytes from peripheral blood, tonsils or adenoids, we observed an absence of B lymphocyte response to isoproterenol, whereas T lymphocytes, which had a lower basal c-AMP level than B cells, exhibited in all experiments a significant increase in cAMP content after isoproterenol stimulation. The intensity of the response varied in the different T lymphocyte subpopulations.