David M. Mosser

The Role of IL-10 in Promoting Disease Progression in Leishmaniasis

To determine the role of IL-10 in cutaneous leishmaniasis, we examined lesion development following Leishmania major infection of genetically susceptible BALB/c mice lacking IL-10. Whereas normal BALB/c mice developed progressive nonhealing lesions with numerous parasites within them, IL-10−/− BALB/c mice controlled disease progression, and had relatively small lesions with 1000-fold fewer parasites within them by the fifth week of infection. We also examined a mechanism whereby Leishmania induced the production of IL-10 from macrophages. We show that surface IgG on Leishmania amastigotes allows them to ligate Fcγ receptors on inflammatory macrophages to preferentially induce the production of high amounts of IL-10. The IL-10 produced by infected macrophages prevented macrophage activation and diminished their production of IL-12 and TNF-α. In vitro survival assays confirmed the importance of IL-10 in preventing parasite killing by activated macrophages. Pretreatment of monolayers with either rIL-10 or supernatants from amastigote-infected macrophages resulted in a dramatic enhancement in parasite intracellular survival. These studies indicate that amastigotes of Leishmania use an unusual and unexpected virulence factor, host IgG. This IgG allows amastigotes to exploit the antiinflammatory effects of FcγR ligation to induce the production of IL-10, which renders macrophages refractory to the activating effects of IFN-γ.

Reversing Lipopolysaccharide Toxicity by Ligating the Macrophage Fcγ Receptors

Our laboratory has previously demonstrated that the ligation of phagocytic receptors on macrophages can influence cytokine production. In this study, we examine the cytokine responses to multiple inflammatory stimuli following FcγR ligation. Macrophages were stimulated in vitro with LPS, lipoteichoic acid, CD40 ligand, or low molecular mass hyaluronic acid. All of these stimuli were proinflammatory in character, inducing the production of high levels of IL-12, but only modest amounts of IL-10. The coligation of FcγR along with these stimuli resulted in an anti-inflammatory profile, abrogating IL-12 production and inducing high levels of IL-10. The modulation of these two cytokines occurred by two independent mechanisms. Whereas the abrogation of IL-12 biosynthesis was a property shared by several macrophage receptors, the induction of IL-10 was specific to the FcγR. The biological relevance of these observations was examined in murine models of endotoxemia, in which FcγR ligation induced the rapid production of IL-10 and prevented IL-12 synthesis. Mice could be passively immunized with Abs to LPS to reverse inflammatory cytokine production, and the transfer of macrophages whose FcγR had been ligated could rescue mice from lethal endotoxemia. Thus, the ligation of the macrophage FcγR can be exploited to prevent inappropriate inflammatory cytokine responses.

Suppression of IL-12 Transcription in Macrophages Following Fcγ Receptor Ligation

Ligating FcγR on macrophages results in suppression of IL-12 production. We show that FcγR ligation selectively down-regulates IL-12 p40 and p35 gene expression at the level of transcription. The region responsive to this inhibition maps to the Ets site of the p40 promoter. PU.1, IFN consensus sequence binding protein, and c-Rel form a complex on this element upon macrophage activation. Receptor ligation abolishes the binding of this PU.1-containing activation complex, and abrogates p40 transcription. A dominant-negative construct of PU.1 diminishes IL-12 p40 promoter activity and endogenous IL-12 p40 protein secretion. Thus, the specificity of IL-12 down-regulation following receptor ligation lies in the inhibition of binding of a PU.1-containing complex to the Ets site of the IL-12 promoter. These findings provide evidence demonstrating for the first time the importance of PU.1 in the transcriptional regulation of IL-12 gene expression.

Leishmania parasites and their ploys to disrupt macrophage activation.

Leishmania are intracellular protozoan parasites of macrophages. At the cellular level, the disease leishmaniasis involves the invasion of tissue macrophages by the parasite, the avoidance of cellular killing mechanisms, and the subsequent intracellular replication of parasites, with the eventual spread of the organisms to adjacent macrophages. This paper describes the process by which Leishmania organisms invade macrophages, with an overview of some of the molecules involved in this process; the mechanisms available to macrophages that have the potential to restrict the growth of Leishmania within them; and the ways that Leishmania and Leishmania-derived molecules can modulate macrophage functions and circumvent leukocyte antimicrobial responses.

Stimulatory and inhibitory signals originatingfrom the macrophage Fcγ receptors

The macrophage receptors for the Fc portion of immunoglobulin G (FcγR) have long been known to mediate a variety of effector functions that are vital to the adaptive immune response. Recent studies, however, have begun to stress potential regulatory roles that these receptors can play in modulating immune and inflammatory responses. In this article we discuss the activating and inhibitory properties of the individual macrophage FcγR and the conditions under which these heterologous responses can occur.

Morphine enhances interleukin-12 and the production of other pro-inflammatory cytokines in mouse peritoneal macrophages

In this study we investigated the capacity of morphine to modulate expression of cytokines in peritoneal macrophages. Mice were implanted subcutaneously with a 75‐mg morphine slow‐release pellet, and 48 h later resident peritoneal macrophages were harvested. Control groups received placebo pellets, naltrexone pellets, or morphine plus naltrexone pellets. Adherent cells were stimulated with lipopolysaccharide (LPS: 10 μg/mL) plus interferon‐γ (IFN‐γ: 100 units/mL) to induce cytokine production. After 24 h RNA was extracted for analysis of cytokine mRNA levels by reverse transcriptase‐polymerase chain reaction, or supernatants were collected after 48 h for determination of cytokine production by enzyme‐linked immunosorbent assay (ELISA). Morphine enhanced mRNA expression of interleukin (IL)‐12 p40 and tumor necrosis factor α (TNF‐α) compared with controls, whereas IL‐10 levels were unchanged by drug treatment. ELISA data showed that both IL‐12 p40 and p70 were increased by morphine. The enhancement of IL‐12 at both the mRNA and protein levels was antagonized by naltrexone, indicating that the modulation of this cytokine by morphine is via a classic opioid receptor. These results are particularly interesting in light of our previous observation that 48 h after morphine pellet implantation, the peritoneal cavity is colonized with gram‐negative and other enteric bacteria. The enhancement of IL‐12 by morphine might be related to morphine‐induced sepsis.

Receptor mediated subversion of macrophage cytokine production by intracellular pathogens

Appropriately regulated immune responses depend on the controlled production of cytokines from antigen presenting cells. IL-12 synthesis is tightly regulated by several redundant mechanisms. One mechanism of IL-12 regulation involves the cross-linking of surface receptors on macrophages. This pathway may be exploited by intracellular pathogens of macrophages to inhibit IL-12 production and delay or prevent the development of cell mediated immunity.

Cooperation between CR1 (CD35) and CR3 (CD11b/CD18) in the binding of complement‐opsonized particles

We analyzed the binding of sheep erythrocytes bearing C3b (EC3b) to cells transfected with human complement receptors. EC3b bound avidly to cells expressing CR1 but failed to bind to cells expressing CR3. In the presence of factor I, the binding of EC3b to CR1 was transient. Primary monocytes and cotransfected cells expressing both CR1 and CR3 mediated a stable rosetting of EC3b, even in the prolonged presence of factor I. This stable adhesion was dependent on the presence of CR3, because blocking CR3 with mAb resulted in the factor I‐dependent release of erythrocytes from these cells. A model is proposed in which these two complement receptors cooperate in a unique manner. These results suggest that the stable adhesion of complement‐opsonized particles to cells expressing CR1 and CR3 is actually a dynamic molecular process in which an important function of leukocyte CR1 is to generate the liganda for CR3.

The taming of IL-12: suppressing the production of proinflammatory cytokines.

Interleukin (IL)‐12 is a cytokine that possesses both proinflammatory and immunoregulatory activity. IL‐12, and the interferon‐γ (IFN‐γ) that is induced by IL‐12, play central roles in the development of the Th1‐type immune responses that are required for immunity to intracellular pathogens. Recently a number of these pathogens, including Leishmania, measles virus, and human immunodeficiency virus (HIV), have been shown to subvert the development of cell‐mediated immunity by actively inhibiting the production of IL‐12. Similarly, the ligation of phagocytic receptors on macrophages has also been shown to suppress IL‐12 production. The suppression of IL‐12 production by receptor ligation occurs by at least two distinct mechanisms: one involves a direct inhibition of gene transcription and the other depends on the production of inhibitory cytokines. We review studies in which IL‐12 has been experimentally manipulated, and we compare the mechanisms by which this regulation can occur. Because the IL‐12 that is produced during acute inflammation and chronic autoimmune disorders can lead to exacerbated disease, the development of pharmacological means to suppress IL‐12 production is currently under investigation. This review focuses on the production of IL‐12 by antigen‐presenting cells and the methods by which the down‐regulation of IL‐12 production can be exploited either by pathogens or for therapeutic ends. J. Leukoc. Biol. 65: 543–551; 1999.

Rhodococcus equi: An emerging opportunistic pathogen

Rhodococcus equi is emerging as a cause of pneumonia in immunocompromised people, especially those with AIDS. Like mycobacteria, R. equi is phagocytosed by alveolar macrophages and replicates within them. Recent work is beginning to elucidate the cell and molecular biology of this opportunistic pathogen and the host immune response to it.