In-vivo gp100-specific Cytotoxic CD8+ T Cell Killing Assay.

Cytotoxic CD8+ T lymphocytes (CTLs) represent a crucial component of the adaptive immune system and play a prominent role in the anti-tumor immune responses of both mice and humans. Cytotoxic CD8+ T cells are responsible for the lysis of cells expressing peptides associated with MHC class I molecules and derived from infection with a pathogen or from mutated antigens. In order to quantify in vivo this antigen-specific CD8+ T cell killing activity, we use the in vivo killing assay (IVKA). Here, we describe the protocol for the lysis of cells expressing a CD8+ T cell melanoma epitope of the hgp10025-33 protein (KVPRNQDWL). C57BL/6 recipient mice, receive first target cells, prepared from naive congenic (CD45.1) C57BL/6 spleen cells pulsed with the hgp10025-33 peptide and labeled with CFSE and of non-pulsed control cells labeled with Brilliant violet. One day later, the spleen cells of recipient mice are isolated and analyzed by FACS to measure the amount of CFSE cells and Brillant Violet (BV) cells. The percentage of lysis is calculated by the difference between CFSE versus BV. Measuring the ability of antigen-specific CD8+ T cells to lyse their antigen in vivo is very important to evaluate the adaptive cytotoxic response induced against a pathogen or a tumor antigen.

Identification of a novel population of highly cytotoxic c-Met-expressing CD8+ T lymphocytes.

CD8+ cytotoxic T lymphocytes (CTLs) are critical mediators of anti-tumor immunity, and controlling the mechanisms that govern CTL functions could be crucial for enhancing patient outcome. Previously, we reported that hepatocyte growth factor (HGF) limits effective murine CTL responses via antigen-presenting cells. Here, we show that a fraction of murine effector CTLs expresses the HGF receptor c-Met (c-Met+ CTLs). Phenotypic and functional analysis of c-Met+ CTLs reveals that they display enhanced cytolytic capacities compared to their c-Met- CTL counterparts. Furthermore, HGF directly restrains the cytolytic function of c-Met+ CTLs in cell-mediated cytotoxicity reactions in vitro and in vivo and abrogates T-cell responses against metastatic melanoma in vivo Finally, we establish in three murine tumor settings and in human melanoma tissues that c-Met+ CTLs are a naturally occurring CD8+ T-cell population. Together, our findings suggest that the HGF/c-Met pathway could be exploited to control CD8+ T-cell-mediated anti-tumor immunity.

Activation of human B cells negatively regulates TGF-ß1 production.

BACKGROUND: Accumulating evidence indicate that B cells can exhibit pro- or anti-inflammatory activities. Similar to interleukin (IL)-10-competent B cells, we recently showed that transforming growth factor (TGF)-ß1-producing regulatory B cells limit the induction of autoimmune neuroinflammation in mice, making them potentially important in maintaining peripheral immune tolerance in central nervous system inflammatory demyelinating disorders such as multiple sclerosis. METHODS: In this study, we compared B cell production of TGF-ß1 and IL-10, the two most studied regulatory cytokines, and the pro-inflammatory B cell-derived IL-6 and tumor necrosis factor cytokines under basal conditions and following polyclonal stimulation with dual B cell receptor (BCR) cross-linking and Toll-like receptor (TLR)9 engagement. RESULTS: We showed that resting TGF-ß1-producing B cells fall within both the naïve (CD27-) and memory (CD27+) B cell compartments. We found no spontaneous B cell-derived IL-10, IL-6 or tumor necrosis factor (TNF) production. Human B cell activation with anti-Ig antibodies plus CPG-B leads to only modest IL-10 production by memory CD19+CD27+ B cells while expression levels of IL-6 and TNF by both naive and memory B cells were strongly induced. Remarkably, stimulated B cells showed significantly reduced capacity to produce TGF-ß1. CONCLUSIONS: These findings indicate that B cell activation may facilitate the development of excessive immune responses and autoimmunity by restricting B cell-derived TGF-ß1 production by resting B cells and favoring in turns the proinflammatory actions of activated cytokine-producing B cells.

MHC class II-dependent B cell APC function is required for induction of CNS autoimmunity independent of myelin-specific antibodies.

Whether B cells serve as antigen-presenting cells (APCs) for activation of pathogenic T cells in the multiple sclerosis model experimental autoimmune encephalomyelitis (EAE) is unclear. To evaluate their role as APCs, we engineered mice selectively deficient in MHC II on B cells (B-MHC II(-/-)), and to distinguish this function from antibody production, we created transgenic (Tg) mice that express the myelin oligodendrocyte glycoprotein (MOG)-specific B cell receptor (BCR; IgH(MOG-mem)) but cannot secrete antibodies. B-MHC II(-/-) mice were resistant to EAE induced by recombinant human MOG (rhMOG), a T cell- and B cell-dependent autoantigen, and exhibited diminished Th1 and Th17 responses, suggesting a role for B cell APC function. In comparison, selective B cell IL-6 deficiency reduced EAE susceptibility and Th17 responses alone. Administration of MOG-specific antibodies only partially restored EAE susceptibility in B-MHC II(-/-) mice. In the absence of antibodies, IgH(MOG-mem) mice, but not mice expressing a BCR of irrelevant specificity, were fully susceptible to acute rhMOG-induced EAE, also demonstrating the importance of BCR specificity. Spontaneous opticospinal EAE and meningeal follicle-like structures were observed in IgH(MOG-mem) mice crossed with MOG-specific TCR Tg mice. Thus, B cells provide a critical cellular function in pathogenesis of central nervous system autoimmunity independent of their humoral involvement, findings which may be relevant to B cell-targeted therapies.

The neurotrophic hepatocyte growth factor attenuates CD8+ cytotoxic T-lymphocyte activity.

BACKGROUND: Accumulating evidence suggests a deleterious role for CD8+ T cells in multiple sclerosis (MS) pathogenesis. We have recently reported that hepatocyte growth factor (HGF), a potent neuroprotective factor, limits CD4+ T cell-mediated autoimmune neuroinflammation by promoting tolerogenic dendritic cells (DCs) and subsequently regulatory T cells. Whether HGF modulates cell-mediated immunity driven by MHC class I-restricted CD8+ T cells remains to be determined. METHODS: Here we examined whether HGF regulates antigen-specific CD8+ T cell responses using an established model of murine cytotoxic T lymphocyte (CTL)-mediated killing. RESULTS: We found that HGF treatment of gp100-pulsed DCs reduced the activation of gp100-specific T cell receptor (Pmel-1) CD8+ T cells and subsequent MHC class I-restricted CTL-mediated cytolysis of gp100-pulsed target cells. The levels of perforin, granzyme B, IFN-γ, and the degranulation marker CD107a as well as Fas ligand were decreased among CD8+ T cells, suggestive of a dual inhibitory effect of HGF on the perforin/granzyme B- and Fas-based lytic pathways in cell-mediated cytotoxicity. Treatment of CD8+ T cells with concanamycin A, a potent inhibitor of the perforin-mediated cytotoxic pathway, abrogated CTL cytotoxicity indicating that blockade of the perforin-dependent killing is a major mechanism by which HGF diminished cytolysis of gp100-pulsed target cells. Moreover, HGF suppressed the generation of effector memory CTLs. CONCLUSIONS: Our findings indicate that HGF treatment limits both the generation and activity of effector CTL from naïve CD8+ T cells. Complementary to its impact on CD4+ T-cell CNS autoimmunity and myelin repair, our findings further suggest that HGF treatment could be exploited to control CD8+ T-cell-mediated, MHC I-restricted autoimmune dysfunctions such as MS.

Cerebrospinal fluid interleukin-6 in central nervous system inflammatory diseases.

BACKGROUND: Interleukin (IL)-6 is recognised as an important cytokine involved in inflammatory diseases of the central nervous system (CNS). OBJECTIVE: To perform a large retrospective study designed to test cerebrospinal fluid (CSF) IL-6 levels in the context of neurological diseases, and evaluate its usefulness as a biomarker to help discriminate multiple sclerosis (MS) from other inflammatory neurological diseases (OIND). PATIENTS AND METHODS: We analyzed 374 CSF samples for IL-6 using a quantitative enzyme-linked immunosorbent assay. Groups tested were composed of demyelinating diseases of the CNS (DD, n = 117), including relapsing-remitting MS (RRMS, n = 65), primary progressive MS (PPMS, n = 11), clinically isolated syndrome (CIS, n = 11), optic neuritis (ON, n = 30); idiopathic transverse myelitis (ITM, n = 10); other inflammatory neurological diseases (OIND, n = 35); and non-inflammatory neurological diseases (NIND, n = 212). Differences between groups were analysed using Kruskal-Wallis test and Mann-Whitney U-test. RESULTS: CSF IL-6 levels exceeded the positivity cut-off of 10 pg/ml in 18 (51.4%) of the 35 OIND samples, but in only three (3.9%) of the 76 MS samples collected. CSF IL-6 was negative for all NIND samples tested (0/212). IL-6 cut-off of 10 pg/ml offers 96% sensitivity to exclude MS. CONCLUSION: CSF IL-6 may help to differentiate MS from its major differential diagnosis group, OIND.

Interferon-ß induces hepatocyte growth factor in monocytes of multiple sclerosis patients.

Interferon-ß is a first-line therapy used to prevent relapses in relapsing-remitting multiple sclerosis. The clinical benefit of interferon-ß in relapsing-remitting multiple sclerosis is attributed to its immunomodulatory effects on inflammatory mediators and T cell reactivity. Here, we evaluated the production of hepatocyte growth factor, a neuroprotective and neuroinflammation-suppressive mediator, by peripheral blood mononuclear cells collected from interferon-ß--treated relapsing-remitting multiple sclerosis patients, relapsing remitting multiple sclerosis patients not treated with interferon-ß, and healthy volunteers. Using intracellular flow cytometry analysis, increased production of hepatocyte growth factor was observed in circulating CD14(+) monocytes from patients undergoing long-term treatment with interferon-ß versus untreated patients. Complementary in vitro studies confirmed that treatment with interferon-ß induced rapid and transient transcription of the hepatocyte growth factor gene in CD14(+) monocytes and that intracellular and secreted monocytic hepatocyte growth factor protein levels were markedly stimulated by interferon-ß treatment. Additional exploration revealed that "pro-inflammatory" (CD14(+)CD16(+)) monocytes produced similar levels of hepatocyte growth factor in response to interferon-ß as "classical" (CD14(+)CD16(-)) monocytes, and that CD14(+) monocytes but not CD4(+) T cells express the hepatocyte growth factor receptor c-Met. Our findings suggest that interferon-ß may mediate some of its therapeutic effects in relapsing-remitting multiple sclerosis through the induction of hepatocyte growth factor by blood monocytes by coupling immune regulation and neuroprotection.

IgG glycan hydrolysis by EndoS inhibits experimental autoimmune encephalomyelitis.

Studies in experimental autoimmune encephalomyelitis (EAE), a mouse model of multiple sclerosis, have shown that B cells markedly influence the course of the disease, although whether their effects are protective or pathological is a matter of debate. EndoS hydrolysis of the IgG glycan has profound effects on IgG effector functions, such as complement activation and Fc receptor binding, suggesting that the enzyme could be used as an immunomodulatory therapeutic agent against IgG-mediated diseases. We demonstrate here that EndoS has a protective effect in myelin oligodendrocyte glycoprotein peptide amino acid 35-55 (MOG(35-55))-induced EAE, a chronic neuroinflammatory demyelinating disorder of the central nervous system (CNS) in which humoral immune responses are thought to play only a minor role. EndoS treatment in chronic MOG(35-55)-EAE did not impair encephalitogenic T cell priming and recruitment into the CNS of mice, consistent with a primary role of EndoS in controlling IgG effector functions. In contrast, reduced EAE severity coincided with poor serum complement activation and deposition within the spinal cord, suggesting that EndoS treatment impairs B cell effector function. These results identify EndoS as a potential therapeutic agent against antibody-mediated CNS autoimmune disorders.

Antibody response in MOG(35-55) induced EAE.

Neurological deficit in experimental autoimmune encephalomyelitis (EAE) and multiple sclerosis is widely considered to be a consequence of synergistic T and B cell responses to central nervous system (CNS) antigens. We show that mice immunized with encephalitogenic myelin oligodendrocyte glycoprotein (MOG(35-55)) peptide develop significant serum levels of anti-MOG antibodies in parallel with disease progression. Furthermore, EAE mice developed antibodies against DNA and RNA, a serological hallmark observed in autoimmune diseases such as systemic lupus erythematosus. The presence of anti-nucleic responsive B cells and antibodies during EAE may highlight a previously unappreciated mechanism in the pathogenesis of CNS autoimmunity.

A novel MEK2/PI3Kδ pathway controls the expression of IL-1 receptor antagonist in IFN-ß-activated human monocytes.

IFN-ß and sIL-1Ra play crucial roles in the regulation of innate immunity and inflammation. IFN-ß, which is widely used to improve the course of relapsing, remitting multiple sclerosis, induces the production of sIL-1Ra in human monocytes through mechanisms that remain largely unknown. In this study, we identified PI3Kδ and MEK2 as key elements that control sIL-1Ra production in isolated human monocytes activated by IFN-ß. Blockade of MEK2, but not of MEK1, by inhibitors and siRNA prevented IFN-ß-induced PI3Kδ recruitment to the membrane, Akt phosphorylation, and sIL-1Ra production, suggesting that MEK2 acted upstream of PI3Kδ. Furthermore, ERK1/2, the only identified substrates of MEK1/2 to date, are dispensable for sIL-1Ra production in response to IFN-ß stimulation. Upon IFN-ß activation, MEK2 and PI3Kδ are translocated to monocyte membranes. These data suggest that MEK1 and MEK2 display different, nonredundant functions in IFN-ß signaling. That neither MEK1 nor ERK1/2 play a part in this mechanism is also an unexpected finding that gives rise to a better understanding of the MAPK signaling network. Together, these findings demonstrate that IFN-ß triggers an atypical MEK2/PI3Kδ signaling cascade to regulate sIL-1Ra expression in monocytes. The premise that MEK1 and MEK2 play a part in the induction of the proinflammatory cytokine, IL-1ß in human monocytes provides a rationale for an alternative, IFN-ß-mediated pathway to induce/enhance sIL-1Ra production and thus, to dampen inflammation.

Glatiramer acetate triggers PI3Kδ/Akt and MEK/ERK pathways to induce IL-1 receptor antagonist in human monocytes.

Glatiramer acetate (GA), an immunomodulator used in multiple sclerosis (MS) therapy, induces the production of secreted IL-1 receptor antagonist (sIL-1Ra), a natural inhibitor of IL-1ß, in human monocytes, and in turn enhances sIL-1Ra circulating levels in MS patients. GA is a mixture of peptides with random Glu, Lys, Ala, and Tyr sequences of high polarity and hydrophilic nature that is unlikely to cross the blood-brain barrier. In contrast, sIL-1Ra crosses the blood-brain barrier and, in turn, may mediate GA anti-inflammatory activities within the CNS by counteracting IL-1ß activities. Here we identify intracellular signaling pathways induced by GA that control sIL-1Ra expression in human monocytes. By using kinase knockdown and specific inhibitors, we demonstrate that GA induces sIL-1Ra production via the activation of PI3Kδ, Akt, MEK1/2, and ERK1/2, demonstrating that both PI3Kδ/Akt and MEK/ERK pathways rule sIL-1Ra expression in human monocytes. The pathways act in parallel upstream glycogen synthase kinase-3α/ß (GSK3α/ß), the knockdown of which enhances sIL-1Ra production. Together, our findings demonstrate the existence of signal transduction triggered by GA, further highlighting the mechanisms of action of this drug in MS.

Glatiramer acetate increases IL-1 receptor antagonist but decreases T cell-induced IL-1beta in human monocytes and multiple sclerosis.

Mechanisms of action as well as cellular targets of glatiramer acetate (GA) in multiple sclerosis (MS) are still not entirely understood. IL-1beta is present in CNS-infiltrating macrophages and microglial cells and is an important mediator of inflammation in experimental autoimmune encephalitis (EAE), the MS animal model. A natural inhibitor of IL-1beta, the secreted form of IL-1 receptor antagonist (sIL-1Ra) improves EAE disease course. In this study we examined the effects of GA on the IL-1 system. In vivo, GA treatment enhanced sIL-1Ra blood levels in both EAE mice and patients with MS, whereas IL-1beta levels remained undetectable. In vitro, GA per se induced the transcription and production of sIL-1Ra in isolated human monocytes. Furthermore, in T cell contact-activated monocytes, a mechanism relevant to chronic inflammation, GA strongly diminished the expression of IL-1beta and enhanced that of sIL-1Ra. This contrasts with the effect of GA in monocytes activated upon acute inflammatory conditions. Indeed, in LPS-activated monocytes, IL-1beta and sIL-1Ra production were increased in the presence of GA. These results demonstrate that, in chronic inflammatory conditions, GA enhances circulating sIL-1Ra levels and directly affects monocytes by triggering a bias toward a less inflammatory profile, increasing sIL-1Ra while diminishing IL-1beta production. This study sheds light on a mechanism that is likely to participate in the therapeutic effects of GA in MS.

Differential regulation of cytokine production by PI3Kdelta in human monocytes upon acute and chronic inflammatory conditions.

Deregulated production of cytokines, including IL-1beta, IL-6 and TNF plays an important role in chronic inflammation. Relevant to this condition, direct cellular contact with stimulated T cells is a potent inducer of cytokine production in human monocytes/macrophages. We previously demonstrated that PI3Ks regulate differential production of IL-1beta and its specific inhibitor secreted IL-1 receptor antagonist (sIL-1Ra) by human monocytes. Here we show that in contrast with PI3Kalpha, beta and gamma, PI3Kdelta accounts for most of the PI3K-dependent signaling ruling the production of IL-1beta, IL-6, TNF and sIL-1Ra in monocytes activated by cellular contact with stimulated T cells (mimicked by CHAPS-solubilized membranes of stimulated T cells, CE sHUT) and lipopolysaccharides (LPS); the latter stimuli being relevant to chronic/sterile and acute/infectious inflammation, respectively. Interestingly, PI3Kdelta activity dampened the production of pro-inflammatory cytokines in LPS-activated monocytes, but induced it in CE sHUT-activated cells. In both CE sHUT- and LPS-activated monocytes PI3Kdelta regulated cytokine transcript expression through the phosphorylation/inactivation of glycogen synthase kinase-3beta (GSK3beta). The blockade of GSK3beta displayed inverse effects to those of PI3Kdelta blockade. Thus, by displaying opposite functions in conditions mimicking chronic/sterile and acute/infectious inflammation, i.e., by repressing pro-inflammatory cytokine expression in LPS-activated monocytes but inducing such mediators in T cell contact-activated monocytes, PI3Kdelta represents a potential therapeutic target specific to chronic/sterile inflammatory conditions.

Stimulated T cells generate microparticles, which mimic cellular contact activation of human monocytes: differential regulation of pro- and anti-inflammatory cytokine production by high-density lipoproteins.

Imbalance in cytokine homeostasis plays an important part in the pathogenesis of chronic inflammatory diseases such as multiple sclerosis and rheumatoid arthritis. We demonstrated that T cells might exert a pathological effect through direct cellular contact with human monocytes/macrophages, inducing a massive up-regulation of the prototypical proinflammatory cytokines IL-1beta and TNF. This mechanism that might be implicated in chronic inflammation is specifically inhibited by high-density lipoproteins (HDL). Like many other stimuli, besides proinflammatory cytokines, the contact-mediated activation of monocytes induces the production of cytokine inhibitors such as the secreted form of the IL-1 receptor antagonist (sIL-1Ra). The present study demonstrates that stimulated T cells generate microparticles (MP) that induce the production of TNF, IL-1beta, and sIL-1Ra in human monocytes; the production of TNF and IL-1beta but not that of sIL-1Ra is inhibited in the presence of HDL. The results were similar when monocytes were stimulated by whole membranes of T cells or soluble extracts of the latter. This suggests that MP carry similar monocyte-activating factors to cells from which they originate. Thus, by releasing MP, T cells might convey surface molecules similar to those involved in the activation of monocytes by cellular contact. By extension, MP might affect the activity of cells, which are usually not in direct contact with T cells at the inflammatory site. Furthermore, this study demonstrates that HDL exert an anti-inflammatory effect in nonseptic activation of human monocytes, not only by inhibiting the production of IL-1beta and TNF but also, by leaving sIL-1Ra production unchanged.

Opposite regulation of IL-1beta and secreted IL-1 receptor antagonist production by phosphatidylinositide-3 kinases in human monocytes activated by lipopolysaccharides or contact with T cells.

The unbalanced production of IL-1beta and its natural, specific inhibitor, the secreted IL-1R antagonist (sIL-1Ra), plays an important role in chronic/sterile inflammation. Relevant to this condition is direct cellular contact with stimulated T cells which is a potent inducer of cytokine production in human monocytes/macrophages. We previously demonstrated that activation of PI3Ks is a prerequisite of the transcription of the sIL-1Ra gene in human monocytes activated by IFN-beta. In this study, we addressed the question of PI3K involvement in the production of IL-1beta and sIL-1Ra in monocytes activated by cellular contact with stimulated T cells (mimicked by CHAPS-solubilized membranes of stimulated T cells (CE(sHUT))), and a crude preparation of LPS, to compare stimuli relevant to chronic/sterile and acute/infectious inflammation, respectively. In monocytes activated by either CE(sHUT) or LPS, the inhibition of PI3Ks abrogated sIL-1Ra transcript expression and sIL-1Ra production, demonstrating that PI3Ks control the induction of sIL-1Ra gene transcription. In contrast, PI3K inhibition increased the production of IL-1beta protein in both CE(sHUT)- and LPS-activated monocytes, the enhancement being drastically higher in the former. This was not due to changes in IL-1beta mRNA steady-state levels or transcript stability, but to the involvement of PI3Ks in the repression of IL-1beta secretion. The downstream PI3K effector, Akt, was implicated in this process. The present results demonstrate that PI3Ks are involved in the inhibition of IL-1beta secretion and in the induction of sIL-1Ra production in human blood monocytes by controlling different mechanisms in conditions mimicking chronic/sterile (CE(sHUT)) and acute/infectious (LPS) inflammation.

From cellular receptors to transduction-transcription pathways for cytokines: at which level should the inhibition be targeted in inflammation?

An imbalance in cytokine homeostasis is considered to play a major part in the pathogenesis of immuno-inflammatory diseases. Since the identification and cloning of cytokines and their receptors, therapeutic approaches have been developed with the purpose of impeding the interaction between the ligand (cytokine) and its specific receptor, or interactions that involve the use of anti-inflammatory cytokines to switch off inflammation. Although some diseases have been treated successfully with cytokines or anticytokines (i.e., anti-TNF, and to a lesser extent recombinant IL-1 receptor antagonist, in rheumatoid arthritis; IFN-beta in multiple sclerosis), the fact remains that these therapies do not abrogate the concomitant use of steroids or immunosuppressive drugs, and that a significant percentage of patients do not respond to such therapies; these are important limitations. The identification of signalling pathways preferentially used in inflammatory conditions has boosted approaches that target these intracellular mechanisms. This review examines the different therapeutic approaches that may be considered for the treatment of immuno-inflammatory diseases, and discusses the advantages and disadvantages of targeting extracellular or intracellular mechanisms.

Differential induction of IL-1beta and TNF by CD40 ligand or cellular contact with stimulated T cells depends on the maturation stage of human monocytes.

Cellular contact with stimulated T cells potently induces cytokine production in monocytes, a mechanism that is likely to be relevant to chronic inflammation. Although the identity of surface molecules involved in this process remains elusive, CD40 and its ligand, CD40L, are thought to be implicated, considering that they are expressed at the inflammatory site. To ascertain the involvement of CD40L, we compared the activation of three different types of human monocytic cells, i.e., freshly isolated monocytes, monocytes primed with IFN-gamma (IFN-gamma-macrophages), and THP-1 cells. These cells were activated by either membranes isolated from stimulated T cells (HUT-78 or T lymphocytes) to mimic cellular contact, soluble extracts from isolated membranes, or CD40L trimer (CD40LT). The production of TNF and IL-1beta was induced by membranes of stimulated T cells in the three types of target cells, whereas CD40LT induced TNF production in IFN-gamma-macrophages only. Similar results were obtained with soluble extracts of T cell membranes, demonstrating that the difference between membranes and CD40LT was not due to the particulate form of membranes. CD40LT induced neither transcript nor protein of cytokines in monocytes, whereas in IFN-gamma-macrophages, IL-1beta and TNF mRNA were observed, but only TNF was measured in cell supernatants. Finally, anti-CD40L Abs failed to inhibit TNF and IL-1beta production induced in IFN-gamma-macrophages by solubilized membranes, whereas TNF production induced by CD40LT was inhibited. These results demonstrate that CD40L is not required in monocyte activation by direct cellular contact with stimulated T cells, although soluble CD40LT induces the production of TNF in IFN-gamma-macrophages.

Opposite effects of IFN beta on cytokine homeostasis in LPS- and T cell contact-activated human monocytes.

Multiple sclerosis (MS) is an immune-mediated disease improved by interferon-beta (IFNbeta) therapy. IFNbeta may owe its anti-inflammatory property to its ability to induce interleukin-1 receptor antagonist (IL-1Ra) without triggering IL-1beta synthesis in human monocytes. Furthermore, we recently demonstrated that IFNbeta inhibits the production of IL-1beta and tumor necrosis factor-alpha (TNF) in human monocytes activated by cellular contact with stimulated T cells, a mechanism which we suspected of playing an important part in the pathogenesis of chronic inflammatory diseases including MS. Here we compare modulatory effects of IFNbeta on the production of proinflammatory cytokines (IL-1beta, IL-1alpha, TNF, and IL-6) and IL-1Ra in human monocytes stimulated by lipopolysaccharides (LPS) and isolated plasma membranes of stimulated T cells (msHUT), which are likely to reflect monocyte activation in acute and chronic inflammation, respectively. In monocytes activated by either LPS or msHUT, IFNbeta did not modulate the secretion of IL-1alpha and IL-6, but it enhanced the production of IL-1Ra in a dose-dependent manner. However, in monocytes activated by msHUT, the expression of cell-associated and intracellular IL-1alpha was inhibited by IFNbeta, correlating with the inhibition of IL-1alpha transcript. IFNbeta inhibited the expression (mRNA) and production (protein) of IL-1beta and TNF, while enhancing those of IL-1Ra in monocytes activated by msHUT. In contrast, in monocytes activated by LPS, IFNbeta enhanced the expression and production of IL-1beta, TNF, and IL-1Ra, suggesting that it did not display anti-inflammatory properties in these conditions. This study demonstrates that IFNbeta displays opposite effects depending on the type of activation of human monocytes, suggesting that it may affect different pathogenic mechanisms in opposite ways.