Different effects of psychotropic drugs on delayed hypersensitivity responses in mice.

The influence of certain psychotropic drugs on the delayed-type hypersensitivity (DTH) response to sheep red blood cells in BALB/c mice was studied. The effects on the overall response and the induction and elicitation phases were evaluated, using two alternative dosage schedules for each agent. It was found that diazepam had no effect on the DTH reaction but the administration of imipramine, haloperidol, chlorpromazine or meprobramate all resulted in depression of the response, impairing both the induction or elicitation phases. The results indicate that psychotropic drugs may produce in vivo depression of cell-mediated immunity by different mechanisms.

Suppression of humoral and cellular immunity in normal mice by diazepam.

Diazepam, a benzodiazepine derivative useful in the treatment of anxiety disorders, was found to depress both primary antibody to sheep red blood cells and delayed-type hypersensitivity responses in normal mice. This immunodepressant activity warrants further investigation owing to its potential consequences on human health and the putative involvement of specific receptors on immunocompetent cells.

The popliteal lymph node response to streptozotocin is under type 1, MHC class-I restricted, CD8(+) T-cell control.

The popliteal lymph node (PLN) assay has been proposed to predict the 'autoimmunogenic' potential of xenobiotics. A better understanding of the processes involved in PLN responses is needed to establish the value of this assay for preclinical safety evaluation. In order to determine whether PLN responses involve CD4(+) or CD8(+) T-cells, the effects of streptozotocin (STZ), a prototypic immunotoxic compound, were analyzed after injection into the hind footpad of C57 BL/6 mice and major histocompatibility complex (MHC) class I or II deficient mice. The involvement of type 1 or type 2 cell control on the production of cytokine mRNAs was analyzed in lymph node cells by quantitative RT-PCR, together with the analysis of a wide range of cytokine mRNAs after STZ injection (IL-1alpha, IL-1beta, TNF-alpha, IFN-gamma, IL-2, IL-2 receptor, IL-4, IL-5, IL-6, IL-10 and IL-12). We have found that mice depleted in CD8(+) T-cells did not respond to STZ, whereas mice depleted in CD4(+) T-cells exhibited the expected positive PLN responses, with increased weight and cellularity indices. STZ induced a low production of interleukin (IL)-2 mRNAs, a mild increase in IL-1alpha and IL-6 mRNAs production, and a dramatic increase in IFN-gamma, IL-1beta, TNF-alpha, IL-12 and IL-2 receptor mRNAs, which correlated with positive PLN responses. No effects on IL-4, IL-5 and IL-10 mRNAs synthesis were noted. In CD8(+) T-cell deficient mice, there was no production of IFN-gamma or IL-6 mRNAs. These results suggest that PLN responses to STZ are under the control of type 1, MHC class-I-restricted, CD8(+) T-cells. This is in accordance to the known physiopathology of STZ-induced diabetes. Additional studies are necessary to establish the mechanism of CD8+ T-cell activation.

Enhancement of antibody response and delayed-type hypersensitivity by thalidomide in mice.

Renewed interest in thalidomide stemmed from the suggestion by several authors that it might prove useful against immune-related diseases. Despite interesting preliminary findings, little is actually known regarding its immunoenhancing properties in vivo. Male and female Swiss mice, aged 6-8 wk, were given 0.5 or 25 mg/kg/day of thalidomide orally for 4 wk. Various immunological parameters were then assessed. When compared to controls, mice exposed to the higher dose presented with a significantly increased thymus weight index, whereas spleen weight index remained unchanged. Anti-sheep erythrocyte plaque-forming cells and hemagglutinin titers were increased by 44% and 29%, respectively. Contact hypersensitivity to picryl chloride was enhanced by 62%. Finally, colloidal carbon clearance was not altered. Interestingly, the lower dose (5 mg/kg/day) exerted no effect whatsoever.

Induction of delayed-type hypersensitivity to sulfamethoxazole in mice: role of metabolites.

Although cutaneous adverse drug reactions (ADRs) are relatively frequent and potentially severe, their mechanisms are poorly understood and no validated predictive experimental model is available. Sulfamethoxazole (SMX) is commonly used to treat infections in HIV-positive patients and severe cutaneous ADRs have been described. This study was undertaken to test whether sensitization to SMX could be achieved in mice using a combination of in vivo and in vitro endpoints. No delayed-type hypersensitivity (DTH) response could be evidenced following SMX injection in the back and subsequent challenge into the footpad or onto the ear. Pretreatment with the enzymatic inducers phenobarbitone and betanaphtoflavone, or depletion in CD4(+) T-lymphocytes were not successful either. In contrast, the injection of SMX/S9 mix in the back and challenge with SMX/S9 mix induced a significant increase in footpad thickness. A significant proliferation of spleen cells from SMX- or SMX/S9 mix-treated mice was evidenced following incubation with SMX/S9 mix, but not SMX alone. This study provides indirect evidence that SMX metabolites are involved and confirms previous in vitro results obtained with lymphocytes from patients with a history of SMX-induced ADRs cultured with murine microsomes. Further investigations using other drugs known to induce similar ADRs are warranted to establish the predictive value of this murine model.

In vitro assessment of phagocytosis: Interspecies comparison of chemiluminescence response.

Phagocytosis is a major component of the host's defences against pathogens. Particulate or soluble stimuli trigger the intracellular respiratory burst in activated phagocytes which can be measured in vitro by the luminol-enhanced chemiluminescent response. In this study, the phagocytic capacity of peripheral blood leucocytes was assayed using chemiluminescence and preliminary in vitro activation by phorbol myristate acetate, opsonized zymosan or latex beads. Leucocytes from rats, mini-pigs, dogs, monkeys and humans were preincubated for 2 hr with either doxycycline (0, 2 and 50 mug/ml) (n = 5) or lead acetate (0, 2 and 20 mug/ml) (n = 5), both compounds impairing phagocytosis. Whatever the species and the activator used, a similar dose-dependent decrease in chemiluminescence response was observed with either doxycycline or lead acetate, showing that the results can be extrapolated between species and to humans. The chemiluminescence assay is proposed as a tool for assessment of the safety of drugs and chemicals.

Immunotoxicity screening of drugs and chemicals: value of contact hypersensitivity to picryl chloride in the mouse.

Contact hypersensitivity to picryl chloride in the mouse is proposed as a valuable tool for the in vivo immunotoxicological testing of new compounds. Reproducibility was found to be excellent. Results obtained in the present work with chlorpromazine, cyclophosphamide, diazepam, haloperidol, hydrocortisone, promethazine and lead acetate, nickel chloride and selenium were similar to those previously reported regarding the effects of these compounds on cell-mediated immunity.

Positive responses to imipramine in the popliteal lymph node assay are due to primary irritation.

The popliteal lymph node (PLN) assay has long been proposed as a tool to detect immunotoxicants with the potential to induce systemic autoimmunity. A major problem hampering the further validation of this assay is the need to rule out irritants that cause false-positive PLN responses. The anti-depressant, imipramine, has not been reported to induce systemic autoimmune reactions in treated patients, but has been repeatedly found positive in the PLN assay, suggesting that this is a false-positive response. To test this hypothesis, the effects of imipramine were compared to those of 50% ethanol in C57B1/6 mice. Footpad edema was evidenced in the few days after injection of both ethanol and imipramine. T-cell depletion using monoclonal antibodies against either CD4+ or CD8+ T-lymphocytes prior to the PLN assay did not influence the responses to either ethanol or imipramine. Cytokine (TNFalpha, IL-1alpha, IL-1beta, IL-2R, IL-6, IL-12 and IFN-gamma) fingerprinting of the PLNs after injection of ethanol and imipramine evidenced the same pattern of responses. These results indicate a closely similar pattern of responses following the footpad injection of either imipramine or ethanol. The conclusion can be drawn that imipramine induces positive responses in the PLN assay via primary (nonspecific) irritation.

Neutrophil functions in lead-exposed workers.

Neutrophil functions were studied in 38 lead-exposed workers compared to 34 controls. Both groups were matched according to age, sex, drinking and smoking habits, ethnic origin and drug intake. Blood lead levels were found to be seven times higher in exposed workers than in controls. Phagocytosis assayed by chemiluminescence was found to be slightly but not significantly altered in exposed workers. In contrast, chemotaxis using the agarose technique was significantly depressed. These results are in agreement with previous in-vitro findings. A further assessment of clinical consequences is warranted.

Influence of macrolide antibiotics on the chemiluminescence of zymosan-activated human neutrophils.

No statistically significant changes in the chemiluminescence response of the whole blood were noted following in vitro treatment with 0, 2, 10, 25, or 50 micrograms/ml of erythromycin, josamycin, miomycin, roxithromycin, and spiramycin. These results suggest that these macrolide antibiotics are unlikely to impair the phagocytizing abilities of human neutrophils, in agreement with previous findings indicating their lack of influence upon neutrophil chemotaxis.

Miconazole influence on both cellular immune response and hepatic drug metabolism in mice.

Miconazole was found to exert a biphasic influence on both cellular immune response and hepatic drug metabolism in adult Swiss mice. Indeed, at the dose of 2.5 or 12.5 mg/kg/twice daily via intraperitoneal route, miconazole depressed delayed-type hypersensitivity (DTH) to sheep red blood cells and hepatic drug metabolism as determined from barbiturate sleeping time, following one-day treatment. By contrast, at the same dose level, miconazole enhanced DTH and hepatic drug metabolism after a five-day administration schedule. Primary humoral response and colloidal carbon clearance were not affected. Although the underlying mechanism remains to be fully elucidated, these findings clearly suggest a close relation between modulation of the cellular immune response and activity of liver drug metabolizing enzymes.

Effect of serotonin on the chemiluminescence response of rat peripheral blood leucocytes.

Serotonin (5-HT) has been shown to exert various immunomodulatory effects. In this study, the effects of 5-HT, 5-hydroxy-DL-tryptophan (5-HTP) and DL-p-chlorophenylalanine (PCPA) on the chemiluminescence (CL) responses of rat peripheral blood leucocytes (PBL) activated by phorbol myristate acetate (PMA), opsonized zymosan or latex beads were assessed. The CL responses were measured following in vitro treatment with 0.01-100 microM 5-HT, and either 1 h after the last i.p. administration of 5-HT (0.05, 0.5, 1, 2.5, 5 or 10 mg/kg for 4 days), 5-HTP (25 or 100 mg/kg for 4 days) or PCPA (200 mg/kg for 4 days, n = 5), or 48 h after a single 200 mg/kg PCPA injection. A concentration-dependent decrease in CL responses was noted following in vitro 5-HT treatment. In vivo treatment of rats with 5-HT produced a reverse bell-curve inhibiting effects on the CL response with a maximal inhibition in rats receiving 1 mg/kg/day 5-HT and a weaker response of PMA-activated PBL. In vivo treatment with high-dose 5-HTP increased CL response of opsonized zymosan-activated PBL, while low-dose 5-HTP decreased CL response of opsonized zymosan and latex beads-activated PBL. No effect was observed in PMA-activated PBL from rats treated with 5-HTP. By contrast, in vivo treatment with PCPA increased CL responses induced by PMA or latex beads, whereas CL responses using opsonized zymosan were not significantly affected. These results suggest that 5-HT modulates the CL response of rat leucocytes to particulate stimuli.

Effects of cimetidine and ranitidine on delayed-type hypersensitivity.

The effects of two selective histamine H2 receptor antagonists on delayed-type hypersensitivity to sheep red blood cells were studied in Balb/c mice. While cimetidine significantly enhanced DTH response when given from immunization to challenge via the intraperitoneal route (10-30 mg/kg twice daily), ranitidine (5-20 mg/kg) did not produce any effect. Both drugs proved ineffective on acquisition as well as expression of DTH. These results suggest that most immunopharmacological effects of cimetidine may not be mediated through histamine H2 receptors, but may possibly be mediated through its imidazole moeity.

Increased production of interferon-gamma, but not IL-4 mRNA, by streptozotocin in the popliteal lymph node assay.

The popliteal lymph node (PLN) assay has been proposed as a tool to predict systemic autoimmune reactions induced by medicinal products and chemicals, the mechanisms of which are poorly understood. To determine whether PLN responses involved Th1 or Th2 cell control, or both, the effects of streptozotocin (STZ), a prototypic immunotoxic compound, were analysed on the production of interferon-gamma (IFN-gamma) and interleukin-4 (IL-4) mRNA by lymph node cells after injection into the hind footpad of C57 BL/6 mice. Streptozotocin induced a dramatic increase in IFN-gamma mRNA production, which correlated with PLN responses as evidenced by augmented weight and cellularity indices. No effect on IL-4 mRNA synthesis was noted. These results suggest that a Th1 response is involved in the PLN response to STZ.

Effects of variations in time and dose of diazepam injection on delayed hypersensitivity.

Administration of diazepam was suggested to exert immuno-suppressive properties in mice. In the present study, we explored the time and dose-dependence of diazepam effects on delayed hypersensitivity to sheep erythrocytes in Balb/c mice. DTH response was only depressed when diazepam was injected shortly after immunization and this effect on DTH induction was found with doses of 4 and 8 mg/kg ip while lower doses (0, 5, 1 and 2 mg/kg) remained ineffective.

Attenuation by carbocromen of cardiac metabolism alterations due to ischemia.

Carbocromen prevents to some extent, particularly in subendocardial layer, carbohydrate cardiac metabolism alterations induced by the ischemia obtained by intermittent occlusion of left coronary artery.

Dual role of dendritic cells in the induction and down-regulation of antigen-specific cutaneous inflammation.

We have previously reported that contact sensitivity (CS) to dinitrofluorobenzene (DNFB) in C57BL/6 mice was mediated by MHC class I-restricted CD8+ T cells and down-regulated by MHC class II-restricted CD4+ T cells. In this study, we analyzed the contribution of dendritic cells (DC) in the induction of these two T cell subsets endowed with opposite functions. Hapten-pulsed skin- and bone marrow-derived DC, obtained from either normal C57BL/6 mice or from MHC class II (I+ II-) and MHC class I (I- II+)-deficient mice, were tested for their ability to prime normal mice for CS to dinitrofluorobenzene. Expression of MHC class I molecules by transferred DC was mandatory both for the induction of CS and for the generation of hapten-specific CD8+ T cells in lymphoid organs. I+ II- DC were as potent as I+ II+ DC in priming for CS, demonstrating that activation of effector CD8+ T cells can occur independently of CD4+ T cell help. I- II+ DC could not immunize for CS, although they could sensitize for a delayed-type hypersensitivity reaction to protein Ags. Moreover, I- II+ DC injected simultaneously with cutaneous sensitization down-regulated the inflammatory response, suggesting that hapten presentation by MHC class II molecules could prime regulatory CD4+ T cells. These results indicate that DC can present haptenated peptides by both MHC class I and class II molecules and activate Ag-specific CD8+ effector and CD4+ regulatory T cell subsets, concurrently and independently.

Mechanism of measles virus-induced suppression of inflammatory immune responses.

Measles virus (MV) causes profound immunosuppression, resulting in high infant mortality. The mechanisms are poorly understood, largely due to the lack of a suitable animal model. Here, we report that particular MV proteins, in the absence of MV replication, could generate a systemic immunosuppression in mice through two pathways: (1) via MV-nucleoprotein and its receptor FcgammaR on dendritic cells; and (2) via virus envelope glycoproteins and the MV-hemagglutinin cellular receptor, CD46. The effects comprise reduced hypersensitivity responses associated with impaired function of dendritic cells, decreased production of IL-12, and the loss of antigen-specific T cell proliferation. These results introduce a novel model for testing the immunosuppressive potential of anti-measles vaccines and reveal a specific mechanism of MV-induced modulation of inflammatory reactions.