Characterization of IL-4 receptor components expressed on monocytes and monocyte-derived macrophages: variation associated with differential signaling by IL-4.

The anti-inflammatory effects of IL-4 on activated monocytes differ from those on monocyte-derived macrophages (MDMac). While IL-4 suppresses LPS-induced IL-1beta , IL-12, IL-10 and TNFalpha production by monocytes, IL-4 suppresses only IL-1beta and IL-12 production by MDMac. The U937 and Mono Mac 6 cell lines have similar cytokine responses to IL-4 as monocytes and MDMac, respectively. The IL-4Ralpha and IL-2Rgamma (gammac) chains are well-characterized components of the IL-4 receptor. Cross-linking studies with 125I-IL-4 revealed that for monocytes and U937 cells, the binding of IL-4 to the receptor components was approximately 1:1 for IL-4Ralpha:gammac. In contrast, for MDMac and Mono Mac 6 cells that have a relative reduction in gammac surface expression, the binding of IL-4 to IL-4Ralpha:gammac was approximately 3:1. Furthermore, IL-4 induced IL-4Ralpha chain phosphorylation more rapidly in MDMac and Mono Mac 6 cells than in monocytes and U937 cells. This study identifies a correlation between altered 125I-IL-4 cross-linking to IL-4Ralpha:gammac, IL-4-induced signaling and regulation of pro-inflammatory cytokine production by IL-4.

Differential responses of human monocytes and macrophages to IL-4 and IL-13.

The primary interleukin-4 (IL-4) receptor complex on monocytes (type I IL-4 receptor) includes the 140-kDa alpha chain (IL-4R alpha) and the IL-2 receptor gamma chain, gamma(c), which heterodimerize for intracellular signaling, resulting in suppression of lipopolysaccharide (LPS)-inducible inflammatory mediator production. The activity of IL-13 on human monocytes is very similar to that of IL-4 because the predominant signaling chain (IL-4R alpha) is common to both receptors. In fact, IL-4R alpha with IL-13R alpha1 is designated both as an IL-13 receptor and the type II IL-4 receptor. When the anti-inflammatory activities of IL-4 and IL-13 were investigated on synovial fluid macrophages and compared with the responses by monocytes isolated from the patients at the same time as joint drainage, the response profiles differed with some responses similar in the two cell populations, others reduced on the inflammatory cells. Similar differences were recorded in the response profiles to IL-4 and IL-13 by monocytes and monocytes cultured for 7 days in macrophage colony-stimulating factor (M-CSF) or granulocyte-macrophage CSF (GM-CSF) (monocyte-derived macrophages, MDMac). MDMac have reduced gamma(c) mRNA levels and reduced expression of the functional 64-kDa gamma(c). There was a similar loss of IL-13R alpha1 mRNA on monocyte differentiation. In turn, there was a significant reduction in the ability of IL-4 and IL-13 to activate STAT6. These findings suggest that different functional responses to IL-4 and IL-13 by human monocytes and macrophages may result from reduced expression of gamma(c) and IL-13R alpha1.

Diminished responses to IL-13 by human monocytes differentiated in vitro: role of the IL-13Ralpha1 chain and STAT6.

The primary IL-13 receptor complex on human monocytes is believed to be a heterodimer comprised of the IL-4R alpha chain and the IL-2R gamma chain (gamma(c))-like molecule, IL-13R alpha1. mRNA levels for IL-13R alpha1, but not IL-4R alpha, were markedly decreased in in vitro monocyte-derived macrophages (MDMac), and with increasing time of monocytes in culture correlated with the loss of IL-13 regulation of lipopolysaccharide-induced TNF-alpha production. Analysis of cell lines Daudi and THP-1 that differentially express gamma(c) and IL-13R alpha1 showed that IL-13 can activate STAT6 in IL-13R alpha1-positive THP-1 cells but not in gamma(c)-positive, IL-13R alpha1-negative Daudi cells. IL-13 activation of STAT6 was reduced in MDMac which was associated with diminished IL-13-induced expression of CD23 and MHC class II. However, with reduced IL-13R alpha1 expression and low nuclear STAT6 activity, some IL-13-induced responses were unaltered in magnitude in MDMac. In the absence of functional IL-13R alpha1 and gamma(c), IL-13 must signal through an alternative receptor complex on MDMac. Experiments with a blocking antibody to IL-4R alpha showed that this chain remains an essential component of the IL-13 receptor complex on MDMac.

Interleukin-4 regulation of human monocyte and macrophage interleukin-10 and interleukin-12 production. Role of a functional interleukin-2 receptor gamma-chain.

Interleukin-4 (IL-4) is the prototypic type 2 immunoregulatory cytokine that can suppress the production of many monocyte and macrophage pro-inflammatory mediators. In this study we investigated the regulation by IL-4 of IL-12 and IL-10 production. While IL-4 suppressed lipopolysaccharide (LPS)-induced IL-12 and IL-10 production by human peripheral blood monocytes, IL-4 suppressed LPS-induced IL-12, but not IL-10, production by synovial fluid mononuclear cells from patients with rheumatoid arthritis. IL-4 also suppressed IL-12, but not IL-10 production, by LPS-stimulated in vitro monocyte-derived macrophages. Similarly, IL-4 cannot suppress LPS-induced tumour necrosis factor-alpha (TNF-alpha) production by synovial fluid cells and monocyte-derived macrophages. The failure of IL-4 to regulate IL-10 production is not due to the failure of IL-4 to suppress TNF-alpha, and vice versa. The data suggest that the IL-4 receptor subunit, gammac, is essential for IL-4 regulation of LPS-induced IL-10 production and that a correlation exists between duration of monocyte culture, reduction in gammac mRNA in cultured cells and hyporesponsiveness of monocyte-derived macrophages to IL-4 for regulation of LPS-induced IL-10 production. This study highlights the importance of investigating responses to IL-4, as a potential therapeutic anti-inflammatory agent, by cells isolated from inflammatory sites and not by the more easily accessible blood monocytes. This study emphasizes the involvement of signalling from gammac in IL-4 regulation of LPS-induced IL-10 production by monocytes and macrophages.

Involvement of the IL-2 receptor gamma-chain (gammac) in the control by IL-4 of human monocyte and macrophage proinflammatory mediator production.

IL-4 has potent anti-inflammatory properties on monocytes and suppresses both IL-1beta and TNF-alpha production. Well-characterized components of the IL-4 receptor on monocytes include the 140-kDa alpha-chain and the IL-2R gamma-chain, gammac, which normally dimerize 1:1 for signaling from the receptor. However, mRNA levels for gammac were very low in 7-day-cultured monocytes. As mRNA levels for gammac declined with culture, so too did the ability of IL-4 to down-regulate LPS-induced TNF-alpha production. In contrast, IL-4 consistently down-regulated IL-1beta production by cultured monocytes. Immunoprecipitation and Western blot analyses demonstrated that 7-day-cultured monocytes do not express the functionally active 64-kDa gammac protein. This was associated with decreased STAT6 activation by IL-4. Studies with Abs to gammac and an IL-4 mutant that is unable to bind to gammac showed that IL-4 can suppress IL-1beta but not TNF-alpha production by LPS-stimulated monocytes in the presence of little or no functioning gammac. IL-4 also suppressed IL-1beta but not TNF-alpha production by Mono Mac 6 cells, which express minimal levels of gammac. For gammac-expressing LPS/PMA-activated U937 cells, IL-4 decreased both TNF-alpha and IL-1beta production. These results suggest that functional gammac is not present on in vitro-derived macrophages, and that while some anti-inflammatory responses to IL-4 are lost with this down-regulation of functional gammac, others are retained. We conclude that different functional responses to IL-4 by human monocytes and macrophages are regulated by different IL-4 receptor configurations.

Interleukin (IL)-10, but not IL-4 or IL-13, inhibits cytokine production and growth in juvenile myelomonocytic leukemia cells.

Juvenile myelomonocytic leukemia (JMML) carries a poor prognosis. The endogenous production of cytokines by the JMML cells contributes to their growth and therapeutic resistance. Interleukin (IL)-4, IL-10, and IL-13 inhibit cytokine production in monocytes. We have now studied whether these cytokines can inhibit JMML cell cytokine production, thereby potentially reducing the malignant cell load in this disorder. We found that IL-10, but not IL-4 or IL-13, dose dependently inhibited JMML cell production of the hemopoietic growth factors granulocyte-macrophage colony-stimulating factor, tumor necrosis factor alpha, and IL-1beta. Similarly, IL-10, but not IL-4 or IL-13, suppressed JMML colony formation and cell viability. This was not due to the absence of receptors because we could detect mRNAs for the IL-4 and the IL-13 receptor alpha subunits and the IL-2 common gamma subunit in JMML cells. Furthermore, the receptors were active since both IL-4 and IL-13 up-regulated surface expression of MHC class II and down-regulated CD14 antigens on JMML cells and monocytes. Unlike activated monocytes, the JMML cells did not produce IL-10. It is suggested that the loss of cytokine inhibitory effects of IL-4 and IL-13 could play a role in the pathogenesis of this disorder. On the other hand, the inhibition of cytokine production, growth, and viability of JMML cells by IL-10 suggests that this cytokine may have a therapeutic potential in JMML.

Control of major histocompatibility complex class II expression on human monocytes by interleukin-4: regulatory effect of lipopolysaccharide.

Interleukin-4 (IL-4), like interferon-gamma (IFN-gamma), stimulates monocyte major histocompatibility complex (MHC) class II expression and thus, by increasing antigen presentation, has the potential to increase immune reactivity. In this study, activation of human monocytes by lipopolysaccharide (LPS) prevented concomitant IL-4 stimulation of MHC class II expression. However, this was not a general observation for activated monocytes because although the high levels of MHC class II antigen expressed by monocytes stimulated in vitro with IFN-gamma were not further regulated by IL-4, the stimulatory effects of IL-4 persisted on cells activated with granulocyte macrophage colony-stimulating factor and tumour necrosis factor-alpha for enhanced MHC class II expression. MHC class II expression by monocytes cultured for 7 days with macrophage colony-stimulating factor was regulated by IL-4 and LPS in a manner very similar to that detected for freshly isolated monocytes. The inhibitory effect of LPS was not due to LPS-induced production of IL-10 or regulatory prostanoids. Furthermore, IFN-gamma-increased MHC class II expression was suppressed by LPS, suggesting that the regulation was at the level of MHC class II expression per se. This study suggests that during Gram-negative bacterial infections, IL-4 and IFN-gamma may not be able to signal enhanced MHC class II expression and thus, enhanced immune reactivity.

Regulation of surface and soluble TNF receptor expression on human monocytes and synovial fluid macrophages by IL-4 and IL-10.

By regulating monocyte and macrophage production of IL-1, its receptor, and its receptor antagonist, IL-4 and IL-10 may exert significant anti-inflammatory activity. We determined whether a similar multicomponent process controlling TNF activity was regulated by IL-4 and IL-10 in nonadherent monocytes and synovial fluid macrophages. Previous studies differed in their conclusions. For both the p75 and p55 TNF receptors, mRNA levels, surface receptor expression, and soluble receptor levels were measured for blood monocytes incubated in vitro for 17, 40, or 64 h with IL-4 or IL-10. The predominant TNF receptor on monocytes, the p75 receptor, was down-regulated by IL-4 at the mRNA level. In turn, both surface and soluble receptor levels on LPS-stimulated cells were reduced and the inhibitory effects were maintained for at least 64 h. In contrast, IL-10 increased surface and soluble p75 TNF receptor levels on monocytes for approximately 40 h, which reflected an increase in receptor mRNA. These studies suggest that IL-4 and IL-10 do not directly regulate the cleavage of TNF receptors from monocytes and macrophages. Addition of an Ab to IL-10 suggested that the stimulatory effects of LPS on p75 TNF receptor expression were due, at least in part, to LPS stimulation of IL-10 production and that IL-4 acted, in part, by decreasing IL-10 production. IL-4 was down-regulatory and IL-10 stimulatory for TNF receptor expression by synovial fluid macrophages. By increasing surface receptor levels, IL-10 enhanced the activities of TNF on monocytes for IL-1beta production. By increasing soluble TNF receptor levels, IL-10 may limit only temporarily the activity of other TNF-responsive cells. This study questions the benefit of IL-10 to resolving TNF-associated inflammation.