Listeria monocytogenes activation of human peripheral blood lymphocytes: induction of non-major histocompatibility complex-restricted cytotoxic activity and cytokine production.

Gram-negative bacteria have been shown to activate human natural killer (NK) cells. In this report, we show that the gram-positive bacterium Listeria monocytogenes can also activate human NK cells with regard to non-major histocompatibility complex (MHC)-restricted killing and the production of cytokines. Overnight incubation of peripheral blood mononuclear (PBM) cells or enriched NK cell populations with live or Formalin-fixed L. monocytogenes resulted in high levels of non-MHC-restricted cytotoxic activity. Listeria-stimulated non-MHC-restricted cytotoxic activity could be achieved with pathogenic as well as nonpathogenic Listeria strains. PBM cells also produced tumor necrosis factor alpha and different interferons (IFNs) after incubation with Listeria strains. Optimal cytokine production appeared to be dependent on nylon wool- and plastic-adherent cells. Different IFNs were produced by Listeria-stimulated PBM cells obtained from different donors. IFN-gamma was always produced but was sometimes associated with IFN-alpha and/or IFN-beta. Interleukin-2 (IL-2) activity was never detected in culture supernatants obtained from Listeria-stimulated PBM cell cultures. However, IL-2 appeared to be produced by Listeria-stimulated PBM cells, since antibody to IL-2 inhibited Listeria-stimulated NK cell cytotoxic activity. Listeria activation of NK cell cytotoxic activity was also dependent on tumor necrosis factor alpha production. Antibody to IFN-gamma, IFN-beta, or IFN-alpha had no effect on Listeria-stimulated NK cell cytotoxic activity. These results demonstrate that NK cells can be activated by Listeria strains and add further evidence that NK cells may play an important role in host defense against bacterial infections.     

Production of Interferon-β by Murine T-Cell Lines Induced by 10-Carboxymethyl-9-Acridanone

Besides the established T-cell property of producing gamma interferon (IFN-gamma), murine T cells additionally possess the ability to produce IFN-alpha and IFN-beta when appropriate inducers such as 10-carboxymethyl-9-acridanone (CMA) or Newcastle disease virus (NDV) are used. Interleukin 2 (IL-2)-dependent murine T-cell lines, but not purified resting splenic T cells, responded to CMA and NDV with production of IFN-alpha, beta. The IFN production by these T cells was not restricted to a special subset, since T cells expressing the Lyt 1+2- and the Lyt 1-2+ phenotype responded to these inducers with IFN production. After prolonged passaging of the T-cell lines in IL-2-containing medium, the ability to respond to CMA with production of antiviral activity was sustained longer than the ability for concanavalin A-induced IFN-gamma production. Whereas the NDV-induced T-cell supernates contained both IFN-alpha and IFN-beta, the induction with CMA resulted exclusively in the synthesis of IFN-beta by the T-cell lines.     

Gamma interferon is produced by human natural killer cells but not T cells during Staphylococcus aureus stimulation.

Gamma interferon (IFN-gamma) production from cultured human peripheral blood mononuclear cells was studied during stimulation with Staphylococcus aureus Cowan I or S. aureus Wood. IFN-gamma was specifically produced from CD16+ natural killer (NK) cells under stimulation by S. aureus Cowan I or Wood because these strains (i) induced IFN-gamma production exclusively from CD3-, CD4- CD8-, and CD16+ cells and (ii) induced CD69 and interleukin 2 (IL-2) receptor alpha expression on CD16+ cells without simultaneously augmenting CD71 or IL-2 receptor alpha on T cells. The effects of biological agents on the induction of S. aureus-induced IFN-gamma production paralleled those of S. aureus-induced CD69 expression on CD16+ cells: IL-2, IFN-alpha, and indomethacin augmented the S. aureus-induced IFN-gamma production, whereas IL-4, transforming growth factor beta 1, prostaglandin E2, and dexamethasone inhibited it. However, IFN-alpha was unique in that it did not induce IFN-gamma production from NK cells while it simultaneously augmented CD69 expression on NK cells, suggesting a unique pathway in the activation of NK cells. Thus, we may conclude that S. aureus-induced IFN-gamma production appears to faithfully represent NK cell function within peripheral blood mononuclear cells.     

Stimulation of human gamma interferon production by diterpene esters.

The diterpene ester 12-O-tetradecanoylphorbol-13-acetate (TPA) and several structurally related compounds were tested for their ability to stimulate interferon (IFN) production in primary cultures of human leukocytes. In cultures of Ficoll-Hypaque-purified mononuclear cells, TPA treatment alone induced only low levels of IFN, but TPA pretreatment of cells caused significant enhancement of IFN yields produced with phytohemagglutinin or several other T cell mitogens. In cultures of unprocessed cells derived from plateletpheresis residues or buffy coats, TPA treatment alone induced high levels of IFN and costimulation with TPA and phytohemagglutinin produced some further enhancement of IFN production. Phorbol 12,13-dibutyrate was comparable to TPA in its ability to enhance phytohemagglutinin-induced IFN production. Several other phorbol ester analogs were also active, but maximal stimulation occurred only at higher drug concentrations. Mezerein, a structurally related diterpene ester, was at least as active as TPA in stimulating IFN production in either Ficoll-Hypaque-purified or unprocessed cells. IFN produced after stimulation with TPA or mezerein, singly or in combination with phytohemagglutinin, had several properties characteristic of IFN-gamma, e.g., it was largely inactivated by dialysis at pH 2, or after exposure to sodium dodecyl sulfate, whereas it was not neutralized by antibody to IFN-alpha and IFN-beta. The stimulatory effect of diterpene esters has proved helpful in producing IFN-gamma for physicochemical analysis and other studies.     

The human intracellular Mx-homologous protein is specifically induced by type I interferons

The murine Mx-1 protein is one of the best biochemically and functionally characterized interferon (IFN)-induced proteins that is necessary, and sufficient, for providing resistance to murine cells against viral influenza infection. Recently an intracellular human protein homologous to the murine Mx-1 protein has been identified by means of a specific monoclonal antibody. The restricted induction of this intracellular protein in human mononuclear cells (MNC) by various cytokines was investigated. MNC from 26 of 28 healthy people and 35 of 36 cancer patients before IFN-alpha therapy had no detectable Mx-homologous protein. Incubation of human MNC with IFN-alpha and IFN-beta for 24 h at different concentrations led to a dose-dependent induction of the Mx-homologous protein. All IFN-alpha or IFN-beta preparations tested were equally effective in eliciting this intracellular protein. IFN-gamma induced only 1% of the Mx amount elicited by type-1 IFN compared on a weight basis. Neither interleukin (IL) 1 nor IL3, IL4, IL5, IL6, tumor necrosis factor-alpha/beta, granulocyte colony-stimulating factor (CSF) or granulocyte macrophage-CSF at any of the concentrations tested were capable of eliciting any detectable amount of the Mx homolog, while IL2 was a poor Mx-homologous protein inducer. In the presence of high-titered IFN-alpha antisera both IL2 and IFN-gamma were unable to stimulate this protein, proving that IFN-gamma and IL2 indirectly induce the Mx homolog via IFN-alpha. Therefore, the human Mx-homologous protein is a strictly by type I IFN-regulated protein in human peripheral blood lymphocytes.     

Lymphokines produced by herpesvirus-transformed marmoset monkey lymphoid cell lines. I. Characterization of a constitutively produced interferon

Conditioned media from cultures of marmoset monkey T-lymphoid cell lines transformed by Herpesvirus saimiri or Herpesvirus ateles were found to contain interferon (IFN) activity. Titers between individual cell lines varied by a factor of 100; large amounts (up to 10(5) units/ml, assayed on human cells) were produced in one of the cell lines. IFN production was enhanced by the diterpene tumor promoters, TPA and mezerein, but not by classical T-cell mitogens. The IFN resembles human IFN-gamma by the following criteria: lability at pH 2, stability against 2-mercaptoethanol, cross-species activity, shape of dose-response curves, and molecular weight determined by size-exclusion chromatography (50,000-55,000). Its activity was not inhibited, however, by antiserum against human IFN-gamma or antisera against human IFN-alpha or IFN-beta.     

Interferon-induced indoleamine 2,3-dioxygenase activity in human mononuclear phagocytes

Interferon (IFN)-induced tryptophan degradation, catalyzed by indoleamine 2,3-dioxygenase (IDO), has been shown to mediate antimicrobial activity in epithelial cells. IDO activity has also been augmented in peripheral blood mononuclear cells (PBMC) treated with IFN or interleukin-2 (IL-2). The effector cells in this population have now been further characterized. PBMCs were isolated from normal donors, separated into monocyte and lymphocyte populations by plastic adherence, treated with IFN or IL-2, and cultivated in medium supplemented with [3H]tryptophan. Culture supernatants were collected after a 48-h incubation and fractionated by high-performance liquid chromatography; radioactivity was determined in fractions corresponding to tryptophan and its metabolites. IFN-gamma and IFN-beta induced IDO activity only in monocytes (plastic-adherent, nonspecific esterase-positive PBMCs). The induction of IDO activity by IL-2 required both monocytes and lymphocytes. Interaction was required between these populations for induction of IDO by IL-2, due to production of IFN-gamma by T lymphocytes, with subsequent IFN-gamma-mediated induction of IDO in monocytes. A number of myeloid cell lines as well as monocyte-derived macrophages were also tested for their ability to be induced to degrade tryptophan in response to IFN treatment. Monocyte-derived macrophages were found to retain their capacity to be induced by IFN-gamma and IFN-beta to degrade tryptophan after differentiation, and to possess seven times more IDO activity per cell than IFN-induced monocytes. However, the presence of lipopolysaccharide (LPS) in the culture medium was required for the maximum induction of IDO activity by IFN-beta. Furthermore, higher concentrations of LPS were sufficient to induce IDO activity in macrophages in the absence of exogenous IFN.     

Type I interferons (IFN-alpha and -beta) suppress cytotoxin (tumor necrosis factor-alpha and lymphotoxin) production by mitogen-stimulated human peripheral blood mononuclear cell.

We studied the effect of the different types of interferons on the production of cytotoxin by human peripheral blood mononuclear cells (PBMCs) stimulated with the mitogen phytohemagglutinin (PHA). Maximum secreted levels of cytotoxin were observed at day 3 in culture and consisted of both tumor necrosis factor alpha (TNF-alpha) and lymphotoxin as determined by specific antibodies. Type I interferons (IFN-alpha and IFN-beta) consistently suppressed cytotoxin production. Both TNF-alpha and lymphotoxin were significantly suppressed. Mean suppression by IFN-alpha and IFN-beta (1000 U/ml) was 56 and 66%, respectively, in PBMCs from 18 different donors. The suppressive effects of IFN-alpha and IFN-beta on cytotoxin production were dose responsive over a range of 10 to 1000 U/ml. Type II interferon (IFN-gamma) did not have consistent significant effects. Pretreatment with IFN-alpha or IFN-beta for 24 or 48 h prior to PHA stimulation also resulted in significant suppression. Supplementation with interleukin-2 (10 U/ml) or IFN-gamma (1000 U/ml) did not overcome cytotoxin suppression by IFN-alpha or IFN-beta. Cytotoxin suppression by IFN-alpha and IFN-beta together appeared to be noninteractive. Suppression appeared not to be due to blockade of the cytotoxin release, since both cell-associated cytotoxin and secreted cytotoxin were suppressed to the same level. These results demonstrated that cytotoxin and lymphotoxin production by PHA-stimulated PBMCs could be down-regulated by type I interferons and that there is a substantial difference between the action of type I interferons and type II interferons (IFN-gamma) in modulating the biosynthesis of cytotoxins.     

Production of a monoclonal antibody to human interferon-alpha (IFN-alpha) and its use to identify IFN-alpha-producing cells in virus infection in vivo

A monoclonal antibody to human interferon-alpha (IFN-alpha) was produced using affinity-purified IFN-alpha, that reacted with recombinant human IFN-alpha 2, but not with IFN-alpha 1, IFN-alpha M1 or IFN-beta. Indirect immunofluorescence using this monoclonal (designated 6C3) and anti-IFN-alpha polyclonal antibodies identified cells expressing IFN-alpha. After Sendai virus induction of normal human buffy-coat cells the proportion of monocytes and lymphocytes expressing IFN-alpha rose progressively from 0% to 50% and 34% respectively, preceding peak IFN-alpha titres in the culture supernatants. Around 80-90% of polymorphs were IFN-alpha-positive using both antisera, with or without IFN induction, although very little IFN bioactivity was released to the supernatant of polymorph cultures after IFN induction. Sections of hepatitis B virus infected human liver tissue showed foci of IFN-alpha-positive infiltrating mononuclear cells and (to a lesser extent) fibroblasts in patients who had active cirrhosis and evidence of virus replication. These findings suggest that polymorphs constitutively express IFN-alpha 2 related antigenic activity, whose biological activity is at present unknown; and demonstrates the identification of IFN-alpha-expressing cells in sections of tissue undergoing natural virus infection.     

Interferon-beta. A potential autocrine regulator of human vascular smooth muscle cell growth.

Positive and negative signals regulate the proliferation in vitro of vascular smooth muscle cells (SMC), a principle cell type in the blood vessel wall. Immune interferon (IFN-gamma, a type II IFN) retards the growth of human SMC, but the effect of type I IFN (IFN-alpha or beta) is unknown. Furthermore, the capacity of SMC to produce IFN is uncharacterized. If type I IFN alters SMC growth and is produced by this cell type, an autocrine inhibitory loop could operate in vascular growth control. To test this possibility, we compared the effects of IFN-alpha, beta, and gamma on the growth of SMC stimulated by platelet-derived growth factor, interleukin-1 or tumor necrosis factor alpha. IFN-beta and IFN-gamma, but not IFN-alpha, consistently retarded growth of SMC cultures (measured by net DNA accumulation and cell number). We investigated whether SMC could produce IFN-beta, a mediator characteristically produced by fibroblasts. Vascular SMC treated with poly(I):poly(C) or tumor necrosis factor-alpha expressed IFN-beta mRNA. SMC treated with poly(I):poly(C) or Newcastle Disease virus elaborated biologically active IFN-beta as well. Our results establish that IFN-beta inhibits human vascular SMC growth and that these cells can express the IFN-beta gene. These findings show that human vascular SMC have the capacity of producing a potential autocrine growth regulator.     

IFN-alpha inhibits IL-3 priming of human basophil cytokine secretion but not leukotriene C4 and histamine release

BACKGROUND: Innate immune responses play a critical role in determining the course of acquired immunity, including that associated with allergic disease. Type I interferons, which are generated early in these reactions, are important soluble factors that prime for TH1-like activity. OBJECTIVE: Because human basophils secrete IL-4 and IL-13 in response to both IgE-dependent and IgE-independent stimuli, we tested whether IFN-alpha, a major type I IFN, affects the production of these TH2 cytokines and/or mediator release from these cells. METHODS: Basophils isolated from blood were treated with IFN-alpha in the presence and absence of IL-3 priming before stimulating through the IgE receptor to release histamine, leukotriene C4, and IL-4. Effects of IFN-alpha on IL-3-mediated IL-13 secretion and basophil survival were also tested. IFN-alpha receptor expression was determined by RT-PCR. RESULTS: IFN-alpha specifically inhibited the effects IL-3 has on basophil cytokine secretion. Enhanced secretion of IL-4 resulting from IL-3 priming was significantly inhibited in cells concurrently cultured with IFN-alpha. This effect was specific for cytokine generation, because histamine and leukotriene C4 were unaffected. Furthermore, IFN-alpha blocked IL-13 secretion directly induced by IL-3. Although IFN-beta also possessed some inhibitory activity, IFN-gamma (a type II IFN) had no effect on basophil cytokine secretion. Basophils constitutively expressed mRNA for the type I IFN receptor, and IFN-alpha did not affect basophil viability with regard to inhibition of cytokine secretion. CONCLUSIONS: These results support the belief that early innate immune responses resulting in IFN-alpha production negatively regulate allergic responses by also inhibiting priming of basophil cytokine release.     

Autoregulatory role of interleukin-10 in hepatitis C patients treated with IFN-alpha.

Interferon-alpha2 (IFN-alpha2) is used as standard treatment of patients with chronic hepatitis C (cHCV), but little is known about the immunomodulatory effects of this cytokine in vivo. We have studied immunologic parameters in freshly isolated peripheral blood mononuclear cells (PBMC) of 26 patients with cHCV 12 h before and 12 h after the first s.c. injection of 5-6 MU IFN-alpha2. In PBMC obtained after IFN injection, a substantial increase in IL-10 production after antigen-specific and nonspecific stimulation was observed, whereas IFN-gamma production and proliferation were significantly diminished compared with PBMC obtained before IFN injection. Patients were stratified according to single nucleotide polymorphisms (SNPs) in the interleukin-10 (IL-10) promoter, which have been associated with the response to IFN therapy. Induction of IL-10 and suppression of IFN-gamma levels were more prominent in patients with genotype CC at position -592 (n = 15) compared with patients with genotype AA/AC (n = 11). In conclusion, our data indicate that IFN-alpha2 therapy can potently enhance IL-10 and suppress IFN-gamma production of PBMC, which is, at least partially, dependent on an SNP in the IL-10 promoter. This suggests an autoregulatory role of IL-10 in IFN therapy.     

Reduction in antiviral activity of human beta interferon by gallic acid.

Gallic acid (GA) is a common part of the human diet, both in the free form and as a metabolite of tannic acid and propyl gallate. Cell cultures were incubated with mixtures of either GA and beta interferon (IFN-beta) (formerly fibroblast IFN) or medium and IFN-beta. The cells were subsequently challenged with virus. The virus plaque yields were greater in cells incubated with IFN-beta and GA than in cells incubated with IFN-beta and medium, indicating that in the former mixture, IFN-beta had lost antiviral activity. The magnitude of the loss was dependent upon the GA concentration. IFN-alpha and IFN-gamma (formerly leukocyte IFN and immune IFN, respectively) were not similarly affected. The effect of GA on IFN-beta could be reversed with 2-mercaptoethanol, suggesting a possible sulfhydryl involvement. Extensive dialysis of IFN-beta-GA mixtures to remove the GA failed to reverse the reduction in antiviral activity. This suggests that a direct and irreversible interaction between IFN-beta and GA took place, reducing the activity of IFN-beta. The significance of this finding with regard to virus infections of the intestine is discussed.     

Gamma interferon enhances mitogenic responses induced by pokeweed mitogen

Interferon (IFN)-induced suppression of the lectin-stimulated lymphoproliferative response was studied comparatively with human IFN-alpha, IFN-beta and IFN-gamma, using an equal unit of their antiviral activity ranging from 31.25 to 1000 IU/ml. Both IFN-alpha and IFN-beta inhibited phytohemagglutinin (PHA) and pokeweed mitogen (PWM)-stimulated lymphoblastogenic response similarly in a dose-related fashion, but the IFN-gamma effect was far less. Indeed, the PWM-stimulated lymphocyte blastogenesis in the cultures incubated for 7 days was enhanced in the presence of IFN-gamma at a concentration of 62.5 IU/ml. The enhancing effect was found to be highest at the lowest concentration of IFN-gamma examined. The IFN-gamma induced enhancement of lectin-stimulated blastogenesis was found mainly in the PWM cultures incubated for 7 days but less in cultures incubated for 5 days or in PHA cultures incubated for 3 days, suggesting that the observed effect might be caused by the activation of interleukin production.     

Gamma interferon induction in human thymocytes activated by lectins and B cell lines.

Human thymocytes in culture synthesized small quantities of interferon (IFN) when stimulated by the lectins concanavalin A or phytohemagglutinin. IFN production by lectin-activated thymocytes was enhanced in the presence of live B lymphoblastoid cells, irradiated B lymphoblastoid cells, or the conditioned medium from B lymphoblastoid cell cultures. The IFN synthesized in mixed cultures had characteristics of IFN-gamma, whereas the IFN synthesized by B lymphoblastoid cells alone could be identified as IFN-alpha on the basis of its neutralization with specific antisera and stability at pH 2. These findings indicate that human thymocytes in culture synthesize IFN-gamma and that B lymphoblastoid cells and their products considerably stimulate IFN-gamma synthesis by lectin-activated human thymocytes in culture. This stimulation was not diminished in the presence of antibodies to IFN-alpha, indicating that IFN-alpha production by B lymphoblastoid cells was not responsible for the stimulatory effect. Removal of adherent cells from thymocyte suspensions did not abrogate IFN-gamma production.     

Production of human interferon-gamma in serum-free medium.

Interferon (IFN)-gamma was produced with a high yield in cultures of human peripheral mononuclear cells by combined stimulation with OK-432 and staphylococcal enterotoxin B. Human mononuclear cells cultured in serum-free medium produced several times as much IFN as those in RPMI-1640 medium containing 10% fetal bovine serum. A synergistic effect of OK-432 and staphylococcal enterotoxin B on the production of IFN-gamma was demonstrated. Ultrogel AcA54 column chromatography of crude IFN showed a single peak with an apparent molecular weight of 43,000. Our production system for human IFN-gamma offers a feasible approach to preparation of large quantities of purified IFN-gamma for structure studies, antibody production, and clinical applications.     

Differential effects of human alpha and gamma interferon on mixed lymphocyte culture and on T-cell-mediated cytotoxicity. Inhibition of proliferation but not of IL-2 production by alpha interferons

We compared the effects of natural and recombinant (r) alpha (IFN-alpha) and gamma (IFN-gamma) interferons on the proliferative responses of human peripheral blood mononuclear cells to mitogens and allogeneic cells in mixed lymphocyte culture (MLC) and on the generation of specific T-cell-mediated cytotoxicity. In 14 of 19 donors, natural IFN-gamma and rIFN-gamma had no significant effect on the proliferative responses to mitogens or allogeneic cells in MLC, even at very high IFN-gamma concentrations (10,000 U/ml). In the remaining 5 donors, a statistically significant (p less than 0.001) enhancement by 49 +/- 8% of the proliferative responses was observed. In contrast, natural IFN-alpha and rIFN-alpha 2 significantly inhibited (p less than 0.001) proliferative responses to mitogens and to allogeneic cells, even at concentrations as low as 10 U/ml, in agreement with previous reports. Although natural and recombinant IFN-alpha significantly inhibited these proliferative responses, they did not affect interleukin-2 (IL-2) production in these cultures, suggesting that they inhibit proliferation by a mechanism that does not involve inhibition of IL-2 production. rIFN-gamma did not affect the generation of specific cytotoxicity in MLC, although it was significantly enhanced by natural IFN-alpha and rIFN-alpha 2. Additionally, we compared the ability of human rIFN-alpha subtypes to inhibit proliferative responses to allogeneic cells in MLC. rIFN-alpha 2, rIFN-alpha 4, and rIFN alpha 7 displayed the most potent inhibitory activity of allogeneic responses and were active at concentrations as low as 0.3-0.6 ng/ml.(ABSTRACT TRUNCATED AT 250 WORDS)     

Differential regulation of NK cell proliferation by type I and type II IFN

IFN, produced during viral infections by accessory (type I IFN) or NK cells (type II IFN), play a primary role in the regulation of immune and anti-viral NK cell effector functions. Because IFN have anti-proliferative effects on several cell types, including hematopoietic cells, we asked whether they modulate proliferation of human NK cells, and whether IFN-alpha and IFN-gamma mediate distinct effects on NK cells at different developmental stages. Analysis of proliferation at the single-cell level in human NK cells indicated that both IFN types inhibit IL-4-induced accumulation of immature CD56(-) IL-13(+) NK cells in freshly separated peripheral blood lymphocytes and in cells derived from them after short-term cultures. However, IFN-gamma inhibited specifically the IL-4-dependent proliferation of these cells without affecting the IL-2-dependent one or that of the IL-13(-) cells, whereas IFN-alpha attenuated proliferation of NK cells at any developmental stage (both immature CD56(-)IL-13(+) and mature CD56(+)IL-13(-) IFN-gamma(+) NK cells) and contributed to their monokine-induced differentiation to IFN-gamma-producing cells. Adding to our previous report that IL-13 inhibits accumulation of mature IFN-gamma(+) NK cells, the present data unravel a mechanism by which peripheral immature IL-13(+) and mature IFN-gamma(+) NK cells can negatively regulate each other's accumulation.     

In vitro regulation by muramyl dipeptide of interferon production from peripheral blood mononuclear cells of healthy volunteers

We investigated the effect of N-acetyl-muramyl-L-alanyl-D-isoglutamine (MDP) on interferon (IFN)-alpha and -gamma production by peripheral blood mononuclear cells from healthy subjects. MDP, on its own, was found to lack the ability to induce IFN production. However, this synthetic adjuvant was able to modulate IFN production induced by other stimuli. In cultures from a considerable number of tested donors, MDP enhanced IFN-gamma levels induced by phytohaemagglutinin. This effect was further potentiated after depleting the PBMNC cultures of their adherent cells. In contrast, MDP significantly suppressed the Sendai virus-induced IFN-alpha and this effect was reversed following adherent cell depletion. Identical regulatory effects on IFN production were exerted by the adjuvant active analogue of MDP, namely murabutide. The adjuvant inactive stereoisomer, MDP (DD) exhibited a similar enhancing effect on IFN-gamma but had a significantly lower inhibitory activity on IFN-alpha production. The potential value of this generation of immunomodulators in the treatment of viral infections and in models for studying the regulation of IFN at the molecular level is discussed.