Inhibitors of serotonin synthesis and antagonists of serotonin 1A receptors inhibit T lymphocyte function in vitro and cell-mediated immunity in vivo.

The serotonin (5HT1A) receptor subtype is one member of the 5HT1 receptor family and is constitutively expressed on Jurkat cells and is elevated on human T lymphocytes after mitogenic activation. Published reports show that human T lymphocytes and monocytes also release 5HT after stimulation with PHA or IFN-gamma. In lymphocytes and the central nervous system, the 5HT1A receptor is coupled to regulation of adenylate cyclase. The 5HT1A receptor agonists inhibit activation of adenylate cyclase. The purpose of the experiments reported here was to investigate further the role 5HT and the 5HT1A receptor may play in the regulation of human and murine T cell activity. For this purpose, human PBMC or murine spleen cells were used for experimental purposes rather than Jurkat cells. The results show that inhibition of 5HT synthesis inhibits IL-2-stimulated human T cell proliferation and that addition of 5-hydroxytryptophan, a precursor of 5HT, reverses inhibition of T cell proliferation. The 5HT1 receptor antagonist, metitepine, and the 5HT1A selective antagonist, pindobind-5HT1A, also block T cell proliferation. Inhibition by metitepine is reversed by 5HT and by the selective 5HT1A receptor agonist, 8-hydroxy-2-(di-n-propylamino) (8-OH-DPAT). Selective 5HT1A receptor antagonists cause elevation of cAMP in human T cells. In a murine model, selective 5HT1A receptor antagonists inhibit contact sensitivity responses but not Ab responses to oxazalone in vivo. Inhibition is reversed by 8-OH-DPAT. In addition, production of Th1 cytokines, such as IL-2 and IFN-gamma, by Ag-stimulated, immune murine spleen cells is inhibited by 5HT1A receptor antagonists in vitro but not by 5HT1C/2 receptor antagonists.

A synthetic mycobacterial heat shock peptide prevents adjuvant arthritis but not proteoglycan-induced synovitis in the rat.

Intradermal immunization of female Lewis rats with 100 micrograms of a nine-amino acid synthetic peptide corresponding to the arthritic T cell-reactive epitope of mycobacterial heat shock protein, three weeks prior to induction of adjuvant arthritis, produced inhibition of day 16 ankle swelling and histologic score. Intraarticular injection of 10 micrograms of bovine articular cartilage proteoglycan monomer emulsified in heavy mineral oil into normal Lewis rat stifle joints produced several hallmarks of chronic synovitis at day 16. Pre-treatment with nonapeptide did not inhibit proteoglycan-induced synovitis. These results indicate that tolerance to the critical epitope of heat shock protein does not abrogate the ability of proteoglycan to induce synovitis in rats.

Role of prostaglandins in the behavioral changes induced by murine interleukin 1 alpha in the rat.

Continuous infusion of murine recombinant interleukin 1 alpha (rIL-1 alpha) produces weight loss, appetite suppression, reduction in horizontal locomotor activity (crossovers) and vertical locomotor activity (rears), and an increase in drinking behavior in the rat. The role of prostaglandins (PG) in the elicitation of these effects was studied. Infusion of rIL-1 alpha produced a transient increase in serum (PGs) which peaked at 24 to 48 h. This increase was completely inhibited by piroxicam. However, inhibition of circulating PG by piroxicam did not block the reductions in appetite, crossover, and rears induced by rIL-1 alpha; it restored normal drinking behavior and only partially restored body weight. Continuous intraperitoneal infusion of PGE2 at 24 micrograms/day exposed the animals to serum levels of PGE2 comparable to those produced by infusion with rIL-1 alpha. Yet, at the point of maximum weight loss induced by rIL-1 alpha (72 h), PGE2 infusion resulted in only a quarter of the weight loss. Compared with rIL-1 alpha, continuously infused PGE2 produced significantly smaller reductions in appetite, crossovers, and rears, and had no effect on drinking behavior. From these observations, we conclude that the rIL-1 alpha-induced increase in drinking behavior was fully dependent on products of the cyclooxygenase pathway, but not necessarily PGE2. However, because of the failure of piroxicam to fully reverse rIL-1 alpha effects on eating, mobility, and weight loss, there must also be a significant PG-independent component to account for the full range of rIL-1 alpha effects.

Effects of continuously administered murine interleukin-1 alpha: tolerance development and granuloma formation.

Continuous infusion of murine recombinant interleukin-1 alpha (rIL-1 alpha) into rats by using intraperitoneally implanted osmotic pumps led to marked decreases in body weight, liver enzymes (serum glutamic oxalacetic transaminase, serum glutamic pyruvic transaminase, and sorbitol dehydrogenase), appetite, and mobility and increases in drinking, blood urea nitrogen, and total peripheral blood leukocytes within 3 days. Granuloma formation was found in the local area of rIL-1 alpha release. As early as day 3, a focal infiltrate of polymorphonuclear leukocytes, mononuclear leukocytes, and plasma cells filled the area; by day 6, extensive fibrosis was found. A loss of rIL-1 alpha-induced changes, with the exception of granuloma formation, occurred by day 10. A marked decrease in the response to rIL-1 alpha was also observed when animals were challenged by implantation of new pumps containing rIL-1 alpha, with monitoring of body weight, or by subcutaneous injection of rIL-1 alpha, with monitoring of serum colony-stimulating factor production. We propose that, even in the continuous presence of interleukin-1, replacement of the acute responses to interleukin-1 by restoration of more normal physiology may be advantageous upon acquisition of specific immunity.

The effects of continuous administration of murine interleukin-1 alpha in the rat.

Recombinant murine IL-1 alpha was administered continuously to rats by means of osmotic pumps implanted intraperitoneally. Continuous infusion of rIL-1 alpha in a range between 0.12 and 12.0 micrograms/day for four days was found to produce concentration-dependent weight loss. Behavioral parameters were continuously monitored and recorded at the 3.0 micrograms/day concentration in electronically-monitored activity cages during Days 2 through 5 of rIL-1 alpha administration. Parameters were separated into those affected during the dark phase (active period) or the light phase (resting period). Eating activity was found to be significantly reduced during each dark period through day 5, when compared with either untreated or PBS vehicle-infused animals. During the fourth and fifth days of infusion, however, eating behavior in animals infused with rIL-1 alpha began to increase toward control level in the latter, but not the earlier, half of the dark period. In contrast, drinking behavior was found to be significantly elevated only during the light periods. Continuous infusion of rIL-1 alpha also produced significant reductions in both horizontal locomotor activity (crossovers) and vertical locomotor activity (rears). However, in contrast to the trend toward a return of normal eating behavior, locomotor activity remained decreased through the fifth day of rIL-1 alpha infusion. These results suggest changes that could be produced by IL-1 in chronic inflammatory disease and infection.

Inhibition of C5a-induced basophil degranulation by disodium cromoglycate.

Injection of purified porcine C5a into 24-hr basophil-rich cutaneous basophil hypersensitivity sites in the dose range 10(-12)-10(-10) moles/site produced cutaneous basophil anaphylaxis (CBA). The H1 antihistamine antagonist mepyramine, given orally (3.0-30 mg/kg), inhibited the vasopermeability, but not the basophil degranulation, characteristic of CBA. The antiallergy agent disodium cromoglycate (DSCG), administered intravenously (3.0-30 mg/kg), inhibited vasopermeability and basophil degranulation. DSCG inhibition of mast cell degranulation was not important in the inhibition of CBA, since intact mast cells were found to be depleted at basophil-rich sites and absent at C5a-induced CBA sites from animals treated with DSCG. C5a at 10(-11) moles/site also induced vasopermeability and mast cell degranulation in normal guinea pig skin. Vasopermeability, but not mast cell degranulation, was inhibited by mepyramine at 30 mg/kg p.o. However, DSCG at 10 mg/kg i.v. failed to inhibit either the vasopermeability or the mast cell degranulation of this reaction. These results indicate that C5a induces the degranulation of both basophils and mast cells in the guinea pig, and that C5a-induced degranulation of basophils, but not mast cells, is inhibited by DSCG.

Effect of disodium cromoglycate on cutaneous basophil anaphylaxis.

Cutaneous basophil anaphylaxis (CBA) was elicited by intradermal rechallenge of cutaneous basophil hypersensitivity (CBH) sites in guinea pigs sensitized 7 days previously with keyhole limpet hemocyanin (KLH). The antiallergy agent disodium cromoglycate (DSCG), administered i.v. immediately before rechallenge, inhibited the increased vasopermeability (measured by tissue dye uptake) and basophil degranulation (measured by light microscopic counts of intact basophils) characteristic of the CBA reaction. The antihistamine mepyramine, administered orally, inhibited vasopermeability but not basophil degranulation. The component contributed by DSCG inhibition of mast cell degranulation to the overall inhibition of the reaction was found to be minimal, since intact mast cells were found to be depleted at CBH sites and totally absent at CBA sites from animals treated with DSCG. Electron microscopic examination of basophils at CBA sites from DSCG-treated animals revealed the presence of ruffled perigranular membranes and enlarged perigranular spaces, but both the formation of degranulation sacs and the subsequent fusion of granule sac membranes with the plasma membrane were inhibited. DSCG also inhibited the vasopermeability and basophil degranulation of the CBA reaction elicited by KLH at day 14 and by C5a at day 7. When a basophil-enriched leucocyte preparation from KLH-sensitized guinea pigs was studied in vitro, DSCG inhibited both antigen-induced and C5a-induced basophil degranulation at 10(-5) and 10(-4) M. DSCG failed to inhibit the vasopermeability and the mast cell degranulation produced by either intradermal C5a or intradermal compound 48/80. These results indicate that anaphylactic degranulation of basophils, but not mast cells, is inhibited by DSCG in the guinea pig. This inhibition appears to take place independent of stimulus at an early stage of granule membrane fusion.