Modulation of lymphocyte motility by beta-endorphin and met-enkephalin

The opioid peptides, beta-endorphin and met-enkephalin, have been shown to affect the immune system, resulting in either enhancement or suppression of immune response. However, the mechanism of the immunomodulatory effects and the immune cells that are affected by the opioid peptides are unclear. Early studies have provided evidence for their influence on granulocytes, monocytes and lymphocytes. In this study, the effect of beta-endorphin and met-enkephalin on the motility of lymphocyte subpopulations was examined. beta-Endorphin, depending on the concentration tested, has only slight enhancing or inhibitory effect on the motility of spleen lymphocytes; interestingly, met-enkephalin inhibited their motility. However, both beta-endorphin and met-enkephalin have a similar effect on the motility of separated spleen T and B lymphocytes, in that the motility of B lymphocytes was enhanced by both opioid peptides whereas the motility of T lymphocytes was inhibited. In contrast to the inhibitory effect of beta-endorphin and met-enkephalin on mature spleen T lymphocytes, the motility of thymocytes was enhanced by both opioid peptides. The results from this study suggest that the interaction of beta-endorphin and met-enkephalin with lymphocytes is complex and intricate.     

Augmentation of concanavalin A-induced immunosuppression by indomethacin

These studies show that, in BALB/C mice, when antibody synthesis against sheep red blood cells is suppressed by concanavalin A, treatment with indomethacin (4-8 mg/kg per os) will augment this suppression. Two other nonsteroidal anti-inflammatory drugs, flufenamic acid and meclofenamic acid (50 mg/kg), also have this effect, whereas phenylbutazone was inactive at this dose. The augmentation of concanavalin A-induced immunosuppression by indomethacin could not be demonstrated on the response to the T-independent antigen polyvinypyrrolidone. In contrast to indomethacin, which inhibits cyclooxygenase, neither nordihydroguaiaretic acid, an inhibitor of the lipoxygenase pathway, nor eicosa-5,8,11,14-tetraynoic acid, an inhibitor of both the cyclooxygenase and the lipoxygenase pathways, had this augmenting effect. Therefore, we do not have strong evidence that the absence of a prostaglandin is responsible for the effect of indomethacin. However, inhibition of the pathway leading to prostaglandin synthesis causes an increase in arachidonic acid metabolism via the lipoxygenase pathway. A product of this pathway, such as a leukotriene, may have immunosuppressive effects in this model. Evidence for the enhancement of a suppressor cell population is provided by an in vitro coculture assay. Cells treated with concanavalin A and indomethacin had more suppressive activity than cells treated with concanavalin A or indomethacin alone.     

Inhibition of concanavalin A-induced mice hepatitis by coumarin derivatives

The effects of coumarin derivatives, osthole, imperatorin, Pd-Ia, Pd-II and Pd-III, on mice concanavalin A (Con A) (0.2 mg/mouse, i.v.)-induced hepatitis were studied. At the dose of 200 mg/kg (i.p.), these coumarins inhibited more than 90% of the Con A-induced elevation of plasma alanine aminotransferase activity, but glycyrrhizin (200 mg/kg, i.p.) caused only 45% inhibition. At the dose of 100 mg/kg (i.p.), osthole produced the strongest inhibition among these coumarins. The inhibitory activity of osthole is lost when its 7-methoxy group is replaced by a 7-hydroxy group to form osthenol. The present results showed that coumarin derivatives inhibited Con A-induced hepatitis, with osthole being the most inhibitory.     

Prevention by rolipram of concanavalin A-induced T-cell-dependent hepatitis in mice

Rolipram is a type IV phosphodiesterase inhibitor endowed with powerful immunomodulatory properties. In this study, we evaluated the effects of this drug on the development of the T-cell-mediated hepatitis inducible in mice by concanavalin A. The results indicated that prophylactic treatment with either 5 or 10 mg/kg rolipram injected intraperitoneally 24 h and 1 h prior to intravenous (i.v.) challenge with 20 mg/kg concanavalin A successfully ameliorated serological and histological signs of liver damage, so that the treated mice showed lower transaminase levels in the plasma and milder mononuclear cell infiltration of the liver as compared to vehicle-treated controls. Moreover, this effect was associated with profound modifications of circulating levels of cytokines released after concanavalin A injection, with the blood levels of interferon-gamma and tumor necrosis factor-alpha being significantly lower and those of interleukin-10 higher than those of the control mice. In particular, the increased blood levels of interleukin-10 might play an important role in the anti-hepatitic effects of rolipram as coadministering this compound with anti-interleukin-10 monoclonal antibody significantly reduced its anti-inflammatory action. These results suggest that rolipram may be useful in the clinical setting for the treatment of cell-mediated immunoinflammatory diseases such as immunoinflammatory hepatitis.     

Protective effects of soyasapogenol A on liver injury mediated by immune response in a concanavalin A-induced hepatitis model

The present study was carried out to analyze the effects of soyasapogenol A on the liver injury mediated by the immune response in concanavalin A-induced hepatitis in mice. Soyasapogenol A reduced the number of infiltrating inflammatory cells in the liver and significantly lowered the elevated level of plasma tumor necrosis factor-alpha (TNF-alpha) 2 h after concanavalin A treatment, and then markedly reduced the elevated plasma alanine aminotransferase activity and decreased the number of apoptotic bodies in the liver parenchymal cells but not in the sinusoidal cells at 24 h. Since the effect of soyasapogenol A on the elevated plasma TNF-alpha level was not appreciable compared to the preventive effect of soyasapogenol A on the elevated plasma alanine aminotransferase level, these results suggest that soyasapogenol A directly prevents apoptosis of hepatocytes, and secondly, inhibits the elevation of plasma TNF-alpha, which consequently resulted in the prevention of liver damage in the concanavalin A-induced hepatitis model.     

Protective effect of andrographolide against concanavalin A-induced liver injury

This study was designed to investigate the hepatic protective effect and the molecular mechanisms of andrographolide in concanavalin A-induced liver injury model. Results showed that andrographolide (Ag) attenuated concanavalin A (Con-A)-induced liver injury and inhibited hepatocyte apoptosis. Further results showed that oxidative stress response genes were significantly elevated during the pathogenesis induced by Con-A. Meanwhile, gadolinium chloride and N-acetyl-l-cysteine (NAC) treatment, which inactivates Kupffer cells or reduces reactive oxygen species, respectively, prevented the liver injury. So the messenger RNA levels of the oxidative response genes mentioned above were detected, and the following results showed that Ag treatment reduced their expression. Besides, serum lactate dehydrogenase and myeloperoxidase activity was significantly reduced by Ag. Finally, Ag treatment did not further reduce serum tumor necrosis factor-α production compared with NAC treatment alone. Thus, our results indicate that Ag prevents Con-A-induced liver injury and reduced the hepatic oxidative stress response. The hepatic protective effect of Ag indicates that Ag supplementation may be beneficial in the treatment of immune-mediated liver injury.     

Separate purinoceptors mediate enhancement by adenosine of concanavalin A-induced mediator release and the cyclic AMP response in rat mast cells

Adenosine caused a concentration-related enhancement of concanavalin A (Con A)-induced 5-hydroxytryptamine (5-HT) release from rat purified peritoneal mast cells. This was accompanied by an enhancement and prolongation of the cyclic AMP response to Con A. The cyclic AMP response but not enhancement of 5-HT release was blocked by 8-phenyltheophylline suggesting the two events to be unrelated. The effects of AMP and ADP, adenosine analogues and adenosine uptake inhibitors suggest the enhancement of 5-HT release to be mediated by a P1-cell surface purinoceptor which does not show the characteristics of either A1 or A2 subtypes.     

Modulatory effect of isoprinosine on lymphocyte proliferative response under immunodepressive conditions

In the present work the effect of Isoprinosine on the mitogenic responses of T and B lymphocytes has been studied. We have found that Isoprinosine can enhance in vitro the response to Concanavalin A. This enhancement was more apparent in cell cultures showing an initially low blastogenic response. In low responses artificially induced by treatments in vivo with cyclophosphamide, our results indicate that Isoprinosine, administered in vivo, does not enhance the response to Con A of treated mice. However, addition of Isoprinosine (75 micrograms/ml) to cultures of spleen cells from mice previously treated with cyclophosphamide enhanced the suppressed response up to normal levels. Neither in vivo nor in vitro Isoprinosine treatments increased the response of lymphocytes to lipopolysaccharide, but usually inhibited the blastogenesis of B cells.     

Modulation of mesolimbic dopaminergic projections by beta-endorphin in the rat

Intracerebroventricular injection of beta-endorphin stimulated the metabolism of dopamine in a dose-dependent, opiate antagonist-reversible manner. Local injections into the nucleus accumbens also caused similar increases, indicating that the actions of this peptide on mesolimbic dopaminergic projections were occurring at opioid receptor sites within the nucleus accumbens. Tolerance experiments suggested that epsilon opioid receptors may be involved in mediating these effects in the n. accumbens, unlike in the striatum.     

Fumigaclavine C improves concanavalin A-induced liver injury in mice mainly via inhibiting TNF-alpha production and lymphocyte adhesion to extracellular matrices

Fumigaclavine C, an alkaloidal metabolite, was produced by Aspergillus fumigatus (strain No. CY018). This study examined the effect of this compound on concanavalin A (Con A)-induced liver injury in mice, a T cell-dependent model of liver damage. Con A administration resulted in severe liver injury, T lymphocyte activation and a strong increment in spleen cell adhesion, as well as in tumour necrosis factor-alpha (TNF-alpha) production. Against this liver injury, the intraperitoneal administration of fumigaclavine C dose-dependently inhibited the elevation in transaminase activity, TNF-alpha production in serum and the histological changes, including inflammatory infiltration, hepatocyte necrosis and degeneration and Kupffer cell hyperplasia. In addition, this compound in-vitro also inhibited the proliferation of spleen cells induced by Con A, and reduced their IL-2 and TNF-alpha production. Moreover, the intraperitoneal administration of fumigaclavine C inhibited the potential of spleen cells isolated from the liver-injured mice to adhere to fibronectin, laminin and type IV collagen. These results suggest that the improvement of this T cell-mediated liver injury by fumigaclavine C may be related to the inhibition of lymphocyte activation, proliferation and adhesion to extracellular matrices as well as the reduction in TNF-alpha production.     

Astilbin prevents concanavalin A-induced liver injury by reducing TNF-alpha production and T lymphocytes adhesion

The aim of this study was to evaluate the effect of astilbin on concanavalin A (Con A)-induced hepatitis, a T cell-dependent model of liver injury. Con A administration resulted in a severe liver injury in mice, with a strong increment in spleen cell adhesion and liver infiltration of T cells, as well as in tumour necrosis factor (TNF)-alpha production. Against this liver injury, astilbin significantly inhibited the elevation in transaminase activity, reduced the TNF-alpha production, and improved the histological changes, including inflammatory infiltration, hepatocyte necrosis and degeneration and Kupffer cell hyperplasia. In addition, astilbin inhibited the adhesion of spleen cells and purified T lymphocytes isolated from the liver-injured mice to fibronectin, laminin and type IV collagen.Moreover, the adhesion of human Jurkat T cells to endothelial cell line ECV-304 was also inhibited by astilbin. These results suggest that the improvement of the T cell-mediated liver injury by astilbinmay be related to the reduction in TNF-alpha production and in T cell adhesion to extracellular matrices and endothelial cells.     

Chlorogenic acid ameliorated concanavalin A-induced hepatitis by suppression of Toll-like receptor 4 signaling in mice

Chlorogenic acid (CGA), one of the most abundant dietary polyphenolic compounds, has been reported to exhibit anti-inflammatory ability. However, the hepatoprotective effects and molecular mechanisms of CGA on concanavalin A (Con A)-induced hepatitis have not been explored. In the present study, we found that pretreatment with CGA dose-dependently inhibited the elevation of plasma aminotransferases and alleviated hepatic pathological damage as well as hepatocyte apoptosis in Con A-exposed mice. Additionally, CGA markedly suppressed the production of serum tumor necrosis factor (TNF)-α and interferon (IFN)-γ, alleviated the infiltration of hepatic macrophages, neutrophils, and activated CD4⁺ T lymphocytes in Con A-primed mice. Moreover, CGA downregulated Con A-induced hepatic expression of adhesion molecules (ICAM-1, VCAM-1 and ELAM-1) mRNA and protein, and inhibited Con A-activated Toll-like receptor (TLR) 4 signal molecules including TLR4, p-IRAK1, p-IκB, and p-p38. Finally, our results also showed that CGA exhibited a therapeutic effect, which CGA posttreatment improved hepatic damage at 1, 3, and 6 h after Con A. Taken together, these data suggested that CGA could effectively prevent mice from Con A-induced hepatitis, which might be through suppressing the activation of TLR4 signaling, downregulating the expression of adhesion molecules, and alleviating the infiltration and activation of hepatic leukocytes and the production of pro-inflammatory cytokines.     

Effect of cyclosporin A on the anti-CD3 antibody- and concanavalin A-induced activation and membrane potential of human T lymphocytes

The immunosuppressive effect of cyclosporin A (CsA) was studied on the anti-CD3 antibody (anti-CD3)- and concanavalin A (Con A)-induced activation of human T lymphocytes. A similar suppressive effect was observed in the Con A- and anti-CD3-induced proliferation as measured by the amount of [3H]thymidine incorporated on the third day of culture. When the cells were washed 30 min after activation with anti-CD3, the response was diminished and the CsA sensitivity was increased. In contrast, addition of interleukin-2 (IL-2) resulted in a significantly increased response and a decreased sensitivity. In cultures activated with anti-CD3, CsA treatment resulted in suppression of proliferation after 3 days and an enhancement after 5 days. This latter phenomenon was not seen in cultures treated with Con A. Removal of macrophages (MPH) abolished the proliferative response to anti-CD3. The addition of 10% MPH restored the response, while pretreatment of MPH with CsA diminished this ability. There were differences in the activation kinetics elicited by these two agents as reflected by the changes in membrane potential and the rate of IL-2 receptor (Tac) expression. The changes in the membrane potential (in the absence of CsA) seemed to be parallel with Tac expression and thymidine incorporation. CsA caused a hyperpolarization of the cell membrane. Both Con A and anti-CD3 brought about a strong depolarization which was blocked by the presence of CsA.     

Modulation of human platelet function by L-canavanine: differential effects of low and high concentrations

L-Canavanine is a naturally occurring L-amino acid that interferes with L-arginine-utilizing enzymes owing to its structural analogy with this L-amino acid. In macrophages and polymorphonuclear leukocytes, which express inducible nitric oxide synthase (iNOS), L-canavanine is able to prevent the L-arginine-derived synthesis of nitric oxide (NO). Its effects on constitutive NOS (cNOS) are far less clear. Because human platelets synthesize NO from L-arginine through a cNOS and because intracellular NO levels modulate platelet function, we have investigated the effects of L-canavanine on parameters potentially influenced by NO, such as platelet levels of 3',5'-cyclic guanosine monophosphate (cGMP) and responses to different aggregating agents. In our experimental conditions, L-canavanine was able to influence the response of human platelets to different aggregating agents such as catecholamines, 5-hydroxytryptamine, and ADP. Low L-canavanine concentrations (10-100 micromol/l) decreased platelet responses, whereas a high concentration (1 mmol/l) was unable to exert antiaggregating effects. In resting platelets, L-canavanine reduced the levels of cGMP, starting from a concentration of 1 mmol/l; furthermore, at the same concentrations, it was able to reduce cGMP levels at the end of the aggregation induced by collagen. In conclusion, L-canavanine exerts differential effects on human platelets in relation to the concentrations: at low levels, it exerts antiaggregating effects by actions independent of NOS inhibition, whereas, at high levels, it inhibits NO synthesis and does not exert antiaggregating effects.     

Crotapotin induced modification of T lymphocyte proliferative response through interference with PGE2 synthesis.

The immunosuppressive property has been demonstrated for the venom of the Crotalus durissus terrificus. Using a simple, novel method for obtaining crotapotin and phospholipase A2 isoforms from venom, it was possible to demonstrate that the addition of crotapotin to cultures of isolated lymphocytes resulted in a significant inhibition of the cellular proliferative response to Concanavalin A. This reduction in blastogenic response of lymphocytes is accompanied by a significant increase in the production of PGE2 by macrophages. This effect on the innate immune response suggests that this compound may modify the subsequent adaptative immune response.     

The opioid specificity of beta-endorphin enhancement of murine lymphocyte proliferation

Beta-endorphin (beta-end) is a potent analgesic peptide which exhibits a variety of pharmacological activities in the central nervous system (CNS) following binding of its N-terminus to specific opioid receptors. Although C-terminal binding sites for this 31-amino-acid peptide have been characterized in CNS tissue, identification of their possible function has been facilitated by studies of beta-end effects on lymphocyte activities. In this communication, we report a detailed analysis of the opioid specificity of the ability of beta-end to enhance T cell mitogen-induced proliferation in unfractionated murine splenocytes. Intact 31-amino-acid beta-end peptides from several species, including human, camel and rat, enhanced concanavalin A-stimulated [3H]thymidine uptake 50-640% in a dose-dependent, naloxone-irreversible fashion. The presence of the C-terminal amino acids was required for the enhancement activity, since met-enkephalin, alpha- and gamma-endorphin, and human beta-end 1-27 were ineffective. Accordingly, the truncated peptides, human beta-end 6-31 and 18-31, were also able to enhance the Con A response. However, human beta-end 18-31 was consistently not as effective as beta-end 6-31 or the intact 31-residue peptide. These data suggest that although the C-terminus contains the primary active sequence, the N-terminus contributes to the overall potency of the effect. In support of this assertion, N-acetylation, which abolishes opioid binding activity, resulted in a reduced magnitude of enhancement. The data suggest that beta-end interacts with a non-opioid receptor which has specificity characteristics strikingly similar to non-opioid receptors characterized in CNS tissue.     

Suppression of the proliferative response of human lymphocytes to cultured allogeneic HeLa cells by zinc

Zinc added to the culture medium caused a dose-related suppression of the proliferative response of human lymphocytes to cultured allogeneic HeLa cells without any significant decrease in cell viability. In contrast to the response to HeLa cells, this metal ion moderately enhanced T lymphocyte response to mitogens such as phytohemagglutinin (PHA), concanavalin A (ConA) and 12-O-tetradecanoylphorbol-13-acetate (TPA). These findings seem to indicate that zinc may be a crucial factor for the modulation of the T lymphocyte function. It can also be considered that the different effect of zinc on the proliferative responses of lymphocytes to allogeneic HeLa cells and to some soluble mitogens might reflect a difference in mechanisms between the two proliferative responses.