In vitro effects of H1-antihistamines on histamine and PGD2 release from mast cells of human lung, tonsil, and skin

Mast cells from different anatomic sites differ in cytochemistry and response to various secretory stimuli. We have investigated whether responsiveness to the second-generation H₁-receptor antagonists, which are important first-line drugs for the relief of symptoms in patients with chronic urticaria and allergic rhinoconjunctivitis, also differs according to the site of origin of mast cells. The effects of terfenadine, ketotifen, and cetirizine were therefore examined in relation to the IgE-dependent release of histamine and prostaglandin D₂ (PGD₂) from dispersed human lung, tonsil, and skin mast cells. Terfenadine had a biphasic effect on lung and skin mast cells: at low concentrations, a concentration-dependent inhibition of histamine release from lung and skin mast cells was observed, whereas at higher concentrations the drug stimulated mediator release. Even at a high concentration, terfenadine inhibited mediator release from tonsil mast cells. Ketotifen had low potency as an inhibitor of mediator release from lung and tonsil mast cells. In skin mast cells, no inhibition of mediator release was observed below 1.0 μM, and above that concentration it induced mediator release. Cetirizine, a much less lipophilic drug than the others tested, did not induce mediator release from mast cells even at concentrations up to 100 μM. This drug showed concentration-dependent inhibition of IgE-dependent mediator release from lung and tonsil mast cells only. Our results show that human mast cells are heterogeneous with respect to modulation of mediator release by these H₁-antihistamines. In particular, differences were observed between skin mast cells and those dispersed from lung and tonsils.     

Comparison of the central nervous system effects produce by six H1-receptor antagonists

Background A comprehensive comparative study of the central nervous system (CNS) properties of newer H₁-receptor antagonists is needed. Objective Our objective was to investigate the central nervous system eifects of u single manufacturer's recommended dose of six H₁-receptor antagonists, using appropriate controls. Methods Fifteen healthy subjects received astemizole 10mg, cetirizine 10mg, ketotifen 2mg, loratadine 10 mg, terfenadine 60mg, diphenhydramine 50mg or placebo. Before and 2-1.5 h after dosing, cognitive function was assessed using the P300-event-related potential, somnolence was assessed using a subjective score, and histamine skin tests were performed. Results In rank order from least to greatest effect on the P300 latency, the medications were: terfenadine, placebo, cetirizine, ketotifen, loratadine, astemizole and diphenhydramine. Only diphenhydramine increased the P300 latency significantly compared with baseline and placebo. Subjective somnolence was significantly greater than baseline and placebo after cetirizine, ketotifen and diphenhydramine. All the H₁-receptor antagonists suppressed the histamine-induced weal significantly compared with baseline. Conclusions The H₁-receptor antagonists tested affected cognitive functioning and somnolence to different extents, although all produced satisfactory peripheral H₁-blockade.     

Differential effects of new-generation H1-receptor antagonists in pruritic dermatoses

In search of an improved treatment of pruritic dermatoses, we have studied azelastine, a novel H₁-receptor antagonist, during a 2-week treatment period, using a double-blind, placebo-controlled design. The potent H₁-antagonist cetirizine was used for comparison. Symptoms were recorded daily by the patients on a diary card, using a 4-point scale. The same parameters and adverse events were evaluated at weekly intervals, and global improvement was evaluated at the end of treatment. In all 230 evaluable patients with moderate to severe itching, azelastine caused an overall significant improvement in comparison to placebo (F=0.02), with significance also for pruritus (P=0.01 after 1 week and P=0.02 after 2 weeks). Both drugs reduced itching more effectively in urticaria than in atopic eczema. Azelastine was superior to cetirizine in reducing pruritus, whereas cetirizine caused a more marked reduction of whealing. Both drugs rarely caused fatigue and dry mouth, but taste perversion occurred only in azelastine-treated patients (9.7%) and headaches only with cetirizine (10.4%). Therefore, the two H, blockers exert differential effects on pruritus verses whealing and a distinctive adverse events pattern. TTie data also underline the low efficacy of antihistamines in atopic eczema, compared to urticaria.     

Influence of H1-receptor antagonists on adhesion molecules and cellular traffic

Bagnasco M, Canonica GW. Influence of H₁-receptor antagonists on adhesion molecules and cellular traffic.     

Ebastine inhibits T cell migration, production of Th2-type cytokines and proinflammatory cytokines

BACKGROUND: Cytokine imbalance and cellular migration to inflammatory sites are critical components of allergic diseases. Redirecting cytokine imbalance and inhibiting cell migration therefore represent important therapeutic strategies for the treatment of these disorders. OBJECTIVES: To study the in vitro effect of ebastine, a novel non-sedating H1 receptor antagonist, on cytokine secretion and migration of activated T cells, as well as production of pro-inflammatory cytokines by macrophages. METHODS: Peripheral T cells obtained from healthy volunteers were cultured in wells coated with the combination of anti-CD3 monoclonal antibody (mAb) and anti-CD26 mAb, anti-CD3 mAb and anti-CD28 mAb, or anti-CD3 mAb with PMA, in the presence or absence of ebastine. T cell proliferation and the production of cytokines were measured by [3H]thymidine incorporation assay and ELISA, respectively. In addition, transendothelial migration of T cells and production of pro-inflammatory cytokines by macrophages were examined. RESULTS: Ebastine inhibited T cell proliferation and the production of IL-4, IL-5, IL-6, and TNF-alpha by T cells under each co-stimulatory condition tested, whereas it exhibited no effect on the production of IL-2 or IFN-gamma. In addition, T cell migration and the production of such pro-inflammatory cytokines as TNF-alpha and IL-6 by macrophages were inhibited by ebastine. CONCLUSIONS: These results indicate that ebastine has a specific inhibitory effect on Th2-type cytokine production. Moreover, ebastine inhibited T cell migration and pro-inflammatory cytokine production by T cells and macrophages, suggesting that ebastine might be useful for the treatment of T cell-mediated allergic inflammatory disorders, including asthma, atopic dermatitis, and Th2-type autoimmune diseases.     

Effects of H1-receptor antagonists on nasal obstruction in atopic patients

The aim of this study is to investigate whether H₁-receptor antagonists, besides their effect on nasal itching and sneezing, also have a measurable effect on nasal obstruction caused by allergen challenge. The antihistamine used was astemizole (10 mg) versus placebo, in a double-blind, cross-over, randomized study of two groups. Between the two sessions there was a wash-out period of at least 4 weeks. Seven patients of both sexes, with proven allergy to grass pollen, underwent a specific long-term provocation with grass pollen in the Vienna challenge chamber. Using a physiological method of challenge and a sensitive method for evaluating nasal function, we were able to prove H₁-receptor antagonist influence on nasal airway obstruction. The main parameters obtained arc nasal flow and nasal resistance at 75, 150 and 300 Pa, evaluated by active anterior rhinomanometry. We also investigated subjective symptom scores (0–3) of nasal, eye. and lung symptoms. It can be shown that the nasal flow under astemizole treatment is statistically significantly higher than the nasal flow under placebo treatment ( P = 0.034). This is in accordance with the findings in subjective nasal itching and sneezing.     

Histamine H1-receptor antagonists with immunomodulating activities: potential use for modulating T helper type 1 (Th1)/Th2 cytokine imbalance and inflammatory responses in allergic diseases

Being a first-line treatment for hypersensitivity allergic disease, histamine H1-receptor antagonists possess anti-inflammatory activity in addition to being H1-receptor antagonists. While it is not purely a histamine-related condition, hypersensitivity allergic disease is associated with an increase in the number of T helper type 2 (Th2) cells and Th2 cytokines, and a decrease in the number of Th1 cells and Th1 cytokines. Suppression of Th2-type cytokine production in addition to H1-receptor blockade may therefore represent a successful therapeutic strategy for the treatment of hypersensitivity allergic diseases. H1-receptor antagonists have been reported to modulate immune cascade at various points by acting on T cell-related inflammatory molecules, including adhesion molecules, chemokines and inflammatory cytokines. These effects of H1-receptor antagonists may be optimized for the treatment of allergic diseases. Besides their ability to regulate inflammatory molecules, some H1-receptor antagonists have been reported to down-regulate Th2 cytokine production. In particular, it has been shown that several H1-receptor antagonists specifically inhibit the production of Th2, but not Th1, cytokines. Accumulating evidence indicates a crucial role for Th1/Th2 cytokine imbalance on the development of allergic diseases. Accordingly, the use of H1-receptor antagonist with Th2 cytokine inhibitory activity to modulate Th1/Th2 cytokine imbalance might be a favourable strategy for the treatment of hypersensitivity allergic diseases. Furthermore, the identification of H1-receptor antagonists which possess immunoregulatory activities in addition to their anti-histamine activity will provide an important insight into the development of novel immunoregulatory drugs.     

CD137 ligand prevents the development of T-helper type 2 cell-mediated allergic asthma by interferon-γ-producing CD8+ T cells

BACKGROUND: Allergic asthma is a T-helper type 2 (Th2) cell-mediated chronic disease that is characterized by airway hyperreactivity (AHR) and chronic eosinophilic airway inflammation. Several studies suggest co-stimulatory molecules like CD137 as potential targets for therapeutic interventions in allergic airway disease. Recently, we could show in a murine asthma model that administration of an agonistic antibody against the receptor of the co-stimulatory molecule CD137 prevented and even reversed an already-established asthma phenotype. OBJECTIVE: The purpose of this study was to analyse the effect of stimulation of the CD137 ligand by a monoclonal antibody (CD137L mAb). METHODS: To induce an asthma-like phenotype, BALB/c mice were sensitized to ovalbumin (OVA), followed by an intrapulmonary allergen challenge. Anti-CD137L or control mAb were applied 1 day before OVA immunization or after the asthma phenotype was already established. RESULTS: Stimulation of the CD137L instead of the receptor by CD137L mAb prevents the development of an asthma-like phenotype but does not reverse established disease. While the receptor-mediated effect is partly mediated by anergy of CD4(+) T cells and partly by induction of IFN-gamma-producing CD8(+) T cells, the effect of the CD137L mAb is completely dependent on IFN-gamma-producing CD8(+) T cells: blockade of IFN-gamma and depletion of CD8(+) T cells fully abrogated the observed protective effect. In vitro experiments showed that the anti-CD137L mAb ligates directly to CD8(+) T cells and induces the generation of IFN-gamma by this cell population. CONCLUSION: Our results demonstrate that anti-CD137L mAb prevents disease development via IFN-gamma-producing CD8(+) T cells but is inferior to stimulation of the receptor that reverses established disease by a mechanism including CD4(+) T cell anergy.     

Role of antigen-presenting cells in the polarized development of helper T cell subsets: evidence for differential cytokine production by Th0 cells in response to antigen presentation by B cells and macrophages

Immune challenges can elicit polarized responses skewed towards the development of T helper type 1 (Th1) or Th2 T cell subsets. To determine if distinct antigen-presenting cells (APC) populations might selectively influence Th subset development, we studied the role of two key APC populations, B cells and macrophages, in the differentiation of effector Th populations from naive precursor Th in vitro. Antigen (Ag)-specific, naive CD4⁺ T cells were enriched from a mouse strain, AND, bearing a transgenic α/β T cell receptor (TCR) encoding reactivity with pigeon cytochrome c peptide 88-104. Peptide Ag was used throughout these studies so that differences in the uptake and processing by the two APC populations would not influence the results. Both APC populations, activated B cells and bone marrow-derived macrophages, supported the development of effector Th having the capacity to secrete high levels of cytokines when restimulated. Regardless of APC population present during effector development, exogenous interferon-γ (IFN-γ) and interleukin-4 (IL-4) had dominant effects on Th subset development. Thus, with both APC populations, effector Th generated in the presence of IFN-γ acquired a Th1-type cytokine profile, Th generated with IL-4 acquired a Th2-type cytokine profile, and Th generated without IFN-γ or IL-4 acquired a Th0-type cytokine profile. B cells and macrophages also had equivalent APC function in the restimulation of Th1 and Th2-like effectors, since only minor differences in cytokine production were noted for these effector populations when restimulated with the two APC populations. However, in 8 of 19 experiments, the Th0-like effector population generated in the presence of IL-2 differentially responded to restimulation with B cells and macrophages, secreting significantly more IFN-γ when restimulated with B cells, and significantly more IL-4 when restimulated with macrophages. We also found that Th effector populations recultured in IFN-γ or IL-4 assumed a more Th1 or Th2-like phenotype, respectively, regardless of their initial cytokine profile. We conclude that through a subtle capacity to skew cytokine production by a Th0 subset, different APC may selectively influence Th subset development under conditions of prolonged or chronic stimulation in an autocrine fashion.     

Inhibitory effect of statins on inflammatory cytokine production from human bronchial epithelial cells

Statins are 3-hydroxy-3-methylglutaryl-co-enzyme A reductase inhibitors of cholesterol biosynthesis, and have been reported to exert pleiotropic effects on cellular signalling and cellular functions involved in inflammation. Recent reports have demonstrated that previous statin therapy reduced the risk of pneumonia or increased survival in patients with community-acquired pneumonia. However, the precise mechanisms responsible for these effects are unclear. In the present study, we examined the effects of statins on cytokine production from lipopolysaccharide (LPS)-stimulated human bronchial epithelial cells (BEAS-2B). Interleukin (IL)-6 and IL-8 mRNA expression and protein secretion in LPS-stimulated cells were inhibited significantly by the lipophilic statin pitavastatin and the hydrophilic statin pravastatin. As these inhibitory effects of statin were negated by adding mevalonate, the anti-inflammatory effects of statins appear to be exerted via the mevalonic cascade. In addition, the activation levels of Ras homologue gene family A (RhoA) in BEAS-2B cells cultured with pitavastatin were significantly lower than those without the statin. These results suggest that statins have anti-inflammatory effects by reducing cytokine production through inhibition of the mevalonic cascade followed by RhoA activation in the lung.     

Effect of non-operative management (NOM) of splenic rupture versus splenectomy on the distribution of peripheral blood lymphocyte populations and cytokine production by T cells

Post-traumatic splenectomy is associated with increased postoperative morbidity and mortality and long-term impairment of humoral and cellular immunity. Alternatives to surgery have been developed to minimize or avoid the immediate and/or long-term complications of splenectomy. Herein we investigated the long-term effect of non-operative management (NOM) of the traumatic rupture of the spleen on the distribution of peripheral blood (PB) lymphocyte populations and cytokine production by T cells. PB samples were drawn from six NOM patients, 13 age-matched adults who had undergone splenectomy after trauma (SP patients) and 31 age-matched controls. Cellular phenotypes and the intracellular production of interferon (IFN)-gamma, interleukin (IL)-2, IL-4 and IL-10 cytokines in T cells were determined in whole blood +/- mitogens by flow cytometry. NOM patients did not show any changes in the absolute numbers of lymphocytes or the distribution of their subsets, compared to the controls. In contrast, SP patients showed a sustained increase in the percentage and/or absolute numbers of lymphocytes, CD8 T cells, activated CD8 T cells, natural killer (NK) T cells, NK cells and gammadelta T cells, and a reduction in naive CD4 T cells. The constitutive or induced cytokine production by T cells of the NOM group was similar to the control group, whereas SP patients had increased percentages of constitutive IL-2- and IFN-gamma-producing CD8 T cells and IFN-gamma-producing CD4 T cells. Our findings indicate collectively that the healing process in NOM does not affect the architecture of the spleen to such an extent that it would lead to long-term alterations of the proportions of PB lymphocytes or the T cell cytokine profiles.     

DC-SIGN, but not sDC-SIGN, can modulate IL-2 production from PMA- and anti-CD3-stimulated primary human CD4 T cells

Dendritic cell (DC)-specific intercellular cell adhesion molecule-3 (ICAM-3)-grabbing non-integrin (DC-SIGN) is expressed on the surface of DCs and specialized macrophages and can support T cell proliferation. Antibody-mediated co-ligation of CD3 and ICAM-3, the ligand for both DC-SIGN and leukocyte function-associated antigen-1, leads to T cell activation. Therefore, we tested to see whether DC-SIGN or a splice variant of dendritic cell-specific intercellular cell adhesion molecule-3-grabbing non-integrin (sDC-SIGN) can co-stimulate primary human T cells. The sDC-SIGN lacking the transmembrane domain encoded by exon 3 localizes to the cytoplasm of cells and is not secreted. Both B7 and DC-SIGN co-stimulated phorbol myristate acetate-stimulated CD4+ cells as compared with controls. However, unlike B7, both DC-SIGN and sDC-SIGN failed to co-stimulate CD4+ T cells treated with sub-optimal amounts of anti-CD3 (2 microg ml(-1)) as defined by a lack of CD69 and CD25 up-regulation, cell division and cytokine secretion. Instead, DC-SIGN, and not sDC-SIGN, induced a small but consistent down-regulation of IL-2 production by these CD4+ T cells. In contrast, DC-SIGN in the presence of 30 mug ml(-1) of anti-CD3 modestly up-regulated cytokine production as compared with control. These results suggest that DC-SIGN can differentially modulate T cell stimulation.     

CD4+CD25+ regulatory T cells: II. Origin, disease models and clinical aspects

Autoimmune diseases afflict approximately 5% of the population and reflect a failure in the immune system to discriminate between self and non-self resulting in the breakdown of self-tolerance. Regulatory CD4⁺CD25⁺ T cells (T_reg cells) have been shown to play an important role in the maintenance of immune homeostasis and self-tolerance by counteracting the development and effector functions of potentially autoreactive T cells. We have in the previous APMIS review described the phenotype and physiology of T_reg cells. The present overview deals with the thymic origin of T_reg cells and their role in disease models such as autoimmune gastritis and inflammatory bowel disease. Finally, we will consider some aspects of the therapeutic potential of T_reg cells.     

Regulation of histamine release from human basophil leucocytes: role of H1, H2 and H3 receptors

A novel class of histamine receptors (H3), controlling histamine synthesis and release, was described in rat and human brain and peripheral nerve endings. The present study was undertaken to evaluate whether H3 receptors contribute to the regulation of histamine release from human basophils. Basophil leucocytes were incubated with a H3 antagonist (thioperamide; concentrations ranging from 1 nM to 10 microM) or with a H3 ((R)alpha methyl-histamine; concentrations ranging from 1 to 100 mM), and subsequently were stimulated with optimal doses of anti-IgE and formyl-methionyl-leucyl-phenyl-alanine (f-met peptide). No significant modifications of histamine release were observed after incubation either with the H3 agonist or with the H3 antagonist. By contrast, a H2 antagonist (cimetidine; concentrations ranging from 1 to 100 microM) exerted a dose-dependent enhancing effect on anti-IgE- and, to a lesser extent, on f-met peptide-induced histamine release. A H1 antihistamine (chlorpheniramine; concentrations ranging from 100 nM to 1 microM), at the highest concentration employed, displayed an inhibitory activity on IgE-dependent and IgE-independent histamine release. Exogenous histamine was shown to exert a dose-dependent inhibitory effect on two-staged anti-IgE-induced histamine release. Taken as a whole, these results suggest that H3 receptors are not involved in the regulation of histamine release from human basophils; by contrast, H2 receptors participate in controlling histamine release from human basophils, as previously demonstrated by other authors.     

Different cytokine production and activation marker profiles in circulating cutaneous-lymphocyte-associated antigen+ T cells from patients with acute or chronic atopic dermatitis

BACKGROUND: Atopic dermatitis (AD) is an inflammatory skin disease whose lesions can have two stages: acute and chronic. In skin biopsies a biphasic pattern of cytokine expression has been shown, Th2 in acute lesions and Th1 in chronic AD lesions. OBJECTIVE: We investigated the expression of an activation marker and a homing receptor, as well as cytokine production, in different peripheral blood T cell subpopulations from AD patients with chronic (Group A) and acute lesions (Group B) and controls. METHODS: We evaluated 26 adult AD patients (12 Group A, 14 Group B) and 14 non-atopic controls. IgE was measured by immunoassay. CD4, CD8, cutaneous-lymphocyte-associated antigen (CLA) and human leucocyte antigen (HLA)-DR expression, and cytokine production (IL-2, IL-13, IFN-gamma, TNF-alpha, IL-10, IL-4) were analysed in mononuclear cells by flow cytometry. RESULTS: In Group B there was a significant increase in eosinophil levels and a non-significant increase in IgE. In Group A we found an increase in CLA(+)CD4(+) cells (8.19+/-1.84) compared with controls (4.83+/-0.53) (P<0.05) and CD4(+)HLA-DR(+) cells in the CLA(+) subpopulation (45.54+/-15.40) compared with controls (30.49+/-6.07) (P<0.05). In the CLA(+)CD4(+) subpopulation, there was a significant increase in IL-4, IL-13 and TNF-alpha production in Group B (12.46+/-7.7, 11.26+/-5.97, 43.92+/-15.55) compared with controls (5.34+/-3.50, 4.54+/-1.78, 19.29+/-9.97) with no differences in Group A. CONCLUSION: Greater immunological differences were detected in peripheral blood from patients with acute compared with chronic lesions, especially in the circulating T cell-subset with skin tropism that preferentially responded to cutaneous allergens. This is the first demonstration of phenotypic changes in circulating CLA(+) T cells between AD patients with acute and chronic lesions.     

Interaction of IL-1 beta, IL-6 and tumour necrosis factor-alpha (TNF-alpha) in human T cells activated by murine antigens.

We used a mixed leucocyte culture between human T cells and irradiated murine splenocytes which allowed us to distinguish between cytokine production from the responder and stimulator cells by the use of species-specific assays for mRNA up-regulation. Using this model of T cell activation by antigen, we studied the effects of human antigen-presenting cell-derived cytokines IL-1 beta, IL-6 and TNF-alpha on the activation of human T cell subsets. We show in this system that exogenously added IL-1 beta, IL-6 and TNF-alpha induces IL-2 receptor (R) up-regulation and IL-2 production, and proliferation by both CD4+ and CD8+ cells. The addition of IL-1 beta induces IL-6 mRNA, and anti-IL-1 antibodies or an IL-1R antagonist protein completely suppresses IL-6 and TNF-alpha supported proliferation. Similarly, addition of IL-6 or TNF-alpha induces up-regulation of IL-1 beta mRNA. However, anti-IL-6 and anti-IL-6R antibodies only partially block proliferation supported by IL-1 beta. These findings suggest that IL-6 and TNF-alpha will induce IL-2R up-regulation/IL-2 secretion via the induction of IL-1 beta production.     

Phenotype, cytokine production and cytolytic capacity of fresh (uncultured) tumour-infiltrating T lymphocytes in human renal cell carcinoma

We investigated the phenotype and functional capacities of tumour-infiltrating lymphocytes (TIL), freshly isolated from primary renal cell carcinoma (RCC) specimens (n = 20). Three-colour flow cytometry immunophenotyping revealed that RCC TIL consist mainly of CD3⁺ T cells, with a clear predominance of CD4⁻CD8⁺ over CD4⁺ CD8⁻ T cells, and a marked population of CD4⁺ CD8⁺ T cells. Natural killer (NK) cells were also strongly represented (> 25% in 15 of 20 tumour samples), while B cells constituted a minor TIL subset (< 5% in 18 of 20 tumour samples). More importantly, the T and NK cells within the tumour displayed a significantly higher expression of the early activation marker CD69 than their counterparts in adjacent normal renal tissue and in peripheral blood. Expression of CD54 and of HLA-DR was also elevated on CD3⁺ TIL, and HLA-DR expression was further vigorously up-regulated following ex vivo stimulation with anti-CD3, all suggesting enhanced immune activity within the tumour microenvironment. CD3⁺ CD4⁺ TIL displayed a normal capacity to up-regulate CD25 expression and to secrete both Th1-type (IL-2, tumour necrosis factor-alpha (TNF-α) and interferon-gamma (IFN-γ)) and Th2-type (IL-4, IL-5 and IL-10) cytokines upon triggering with anti-CD3. Furthermore, cytokine production was susceptible to modulation by CD28 costimulation. CD3⁺ CD8⁺ TIL, on the other hand, consistently demonstrated a poor up-regulation of CD25 upon triggering with anti-CD3, and displayed poor ex vivo cytolytic activity in an anti-CD3-redirected 4-h cytotoxicity assay against murine P815 cells. Collectively, our findings indicate that the CD3⁺ CD4⁺ TIL in RCC have normal functional capacities, whereas the proportionally major CD3⁺ CD8⁺ TIL are functionally impaired. The relevance of these findings to the in vivo local immune response in RCC is discussed.     

Pretreatment of activated human CD8 T cells with IL-12 leads to enhanced TCR-induced signaling and cytokine production

During the immune response to pathogens and autoantigens, CD8T cells are exposed to numerous inflammatory agents including the cytokine IL-12. Previous studies have focused on how IL-12 regulates T cell functions when present during or after the activation of the T cell receptor (TCR). However, recent studies suggest that prior exposure to IL-12 also alters the TCR responsiveness of murine T cells. Whether similar phenomena occur in human activated CD8T cells and the mechanisms mediating these effects remain unexplored. In this study, we observed that pretreatment of human activated CD8T cells with IL-12 results in increased cytokine mRNA and protein production following subsequent TCR challenge. The potentiation of TCR-mediated cytokine release was transient and required low doses of IL-12 for at least 24 h. Mechanistically, prior exposure to IL-12 increased the TCR induced activation of select MAPKs and AKT without altering the activation of more proximal TCR signaling molecules, suggesting that the IL-12 mediated changes in TCR signaling are responsible for the increased production of cytokines. Our data suggest that prior treatment with IL-12 potentiates human CD8T cell responses at sites of infection and inflammation, expanding our understanding of the function of this clinically important cytokine.