Combined effects of tumor necrosis factor-alpha, prostaglandin E2, and corticosterone on induced Ia expression on murine macrophages

Ia expression is an important marker of macrophage functional capacity. IFN-gamma induces Ia expression on perhaps all murine macrophages, whereas IL-4, granulocyte-macrophage CSF, and CSF-1 induce Ia on restricted sets of macrophages. Inhibitors of expression include PGE2, glucocorticoids, and IFN-beta. TNF has been found to augment Ia expression on several macrophage lineage cell lines but to inhibit expression on murine peritoneal macrophages. Our study shows that TNF can have opposite effects on Ia expression (induced by IFN-gamma) on thioglycollate-elicited peritoneal macrophages, depending on the length of time cells are treated and on the presence of other modulators. In particular, TNF augmented early expression induced by IFN-gamma but inhibited later expression. And although TNF synergized with PGE2 to markedly inhibit Ia induction on these cells, it partially antagonized the inhibition by corticosterone and IFN-beta. TNF and PGE2 also synergized to inhibit Ia expression induced on bone marrow-derived and splenic macrophages by either IFN-gamma or IL-4. In contrast to their effect on Ia expression, TNF and PGE2 had opposite effects on expression of gamma 2a FcR in macrophages. TNF blocked the increase in FcR expression due to any combination of PGE2, IFN-gamma, and IFN-beta. However, TNF and PGE2 both increased expression of gamma 2a FcR on WEHI-3 cells. If the different effects of TNF reflect the differentiation states of macrophages, its effects on Ia and FcR expression may vary with the progression of an immune response.     

Monoclonal antibody-mediated inhibition of interferon-gamma-induced macrophage antiviral resistance and surface antigen expression

A monoclonal antibody (MAb) with specificity for murine interferon-gamma (IFN-gamma) was used as a probe for studying the effect of recombinant IFN-gamma (rIFN-gamma) on antiviral activity, Fc receptor expression, and Ia antigen induction in macrophages. Cultures of C3H/HeJ peritoneal exudate macrophages were used to allow direct comparison of all three functions in the same target cell system. Our data provide two major findings: the efficacy of the MAb is very different depending on whether murine fibroblasts or macrophages are used as the target cell in the antiviral assay, i.e., greater than 20 to 100 times more MAb was required to block antiviral activity in macrophage cultures; and 10 to 50 times more MAb was required to inhibit Fc receptor vs Ia antigen expression in response to rIFN-gamma. These latter findings confirm and extend previous observations, which indicate that the induction pathways of two important differentiation markers by IFN-gamma may be dissociable.     

Regulation of macrophage activation by IL-3. I. IL-3 functions as a macrophage-activating factor with unique properties, inducing Ia and lymphocyte function-associated antigen-1 but not cytotoxicity

Here we report that IL-3 (also referred to as multi-CSF because of its colony-stimulating activity on a variety of hemopoietic cell lineages) can function as a macrophage-activating factor (MAF). IL-3 was able to regulate the expression of class II MHC Ag and the cellular interaction molecule lymphocyte function-associated Ag-1 on the surface of murine peritoneal exudate cells. The kinetics of IL-3-induced Ia expression appeared to be distinct from that induced by either IFN-gamma, IL-4, or granulocyte-macrophage-CSF. IL-3 was also distinguished from these factors by the finding that it did not induce macrophage tumoricidal activity. In addition to its inherent MAF activities, IL-3 also showed a marked synergy with low doses of LPS (0.05 to 0.5 ng/ml) as well as IFN-gamma in Ia induction. When lymphocyte function-associated Ag-1 expression was evaluated, the effects of these stimuli appeared to be only additive. Although LPS has been shown to inhibit IFN-gamma-induced Ia expression, in our experiments this property of LPS is manifest only when present at doses greater than or equal to 50 ng/ml. At lower concentrations, LPS potentiated both IL-3- and IFN-gamma-induced class II MHC Ag expression. Data presented here also suggest that the synergistic interactions between low doses of LPS and IL-3 are not mediated by known LPS-inducible cytokines of macrophage origin, because rIL-1, TNF-alpha, or IL-6 did not enhance the response to IL-3. Because IL-3 can also participate in the regulation of IL-1 expression, it appears that IL-3 can function as a MAF which selectively regulates the accessory cell characteristics required for Ag presentation, as opposed to the cytolytic functions of the macrophage.     

Splenic macrophage function in early syphilitic infection is complex. Stimulation versus down-regulation

Macrophages are important regulatory cells that can both stimulate and down-regulate various immune functions. During syphilitic infection, these cells phagocytize, kill, and lyse Treponema pallidum. They also modulate early T cell activation by decreasing IL-2 production through secretion of PG. This report focuses on additional complexities of macrophage regulation. Non-adherent splenic cells were stimulated with Con A to induce IFN-gamma synthesis. High levels were detected in preparations from normal rabbits and much lower levels in preparations from infected rabbits. The organisms also readily stimulated IL-1 synthesis by adherent spleen preparations from normal but not from infected rabbits. When indomethacin was added to these latter preparations, this IL-1 defect was reversed, implicating PG in this down-regulation. Spleen cells were obtained from normal rabbits and from rabbits infected testicularly for 9 to 12 days. Infection elevated basal levels of class II Ia Ag on adherent cells. In addition, macrophage Ia expression was increased during 4 days of in vitro incubation with treponemes. Non-adherent spleen cells from infected animals inhibited two different macrophage functions. First, culture filtrates obtained after 48 h of incubation contained a soluble factor that subsequently decreased LPS-induced IL-1 synthesis. Second, when macrophages were co-incubated with non-adherent cells, treponemal stimulation of macrophage Ia expression was inhibited; this inhibition was reversed by indomethacin implicating prostaglandins in this down-regulation. In further experiments an exogenous source of IFN-gamma was incubated with adherent cells from infected rabbits. This stimulated macrophage function as shown by increased IL-1 synthesis and Ia expression and decreased PGE2 secretion. Results are discussed in terms of the complexities of immunoregulation by macrophages during syphilitic infection.     

Induction of macrophage Ia expression by lipopolysaccharide and Listeria monocytogenes in congenitally athymic nude mice

Experiments were performed to analyze the mechanism by which lipopolysaccharide (LPS) modulates the expression of Ia by murine peritoneal macrophages in vivo. We investigated the effect of LPS on Ia expression in T cell deficient mice by using the congenitally athymic nude mouse model. Injection (i.p) of LPS into athymic (nu/nu) mice resulted in a dramatic increase in the expression and biosynthesis of Ia by peritoneal macrophages 7 days after injection. The magnitude and kinetics of this induction were equivalent to increases observed after LPS injection of euthymic (nu/+) mice. Viable Listeria monocytogenes also increased Ia expression in athymic mice, but in contrast to the induction observed in euthymic mice at 3 and 7 days after injection, increased Ia expression was not seen until 7 days. Ia induction by either LPS or L. monocytogenes in athymic mice was not due to the presence or development of mature T cell function as defined by assays for T cell mitogenesis and interleukin 2 production. We conclude that increased macrophage Ia expression by LPS and L. monocytogenes in vivo can occur in the absence of mature functioning T cells.     

Reversal of defective IL-6 production in lipopolysaccharide-tolerant mice by phorbol myristate acetate

The development of LPS tolerance has been suggested to be mediated by an inhibition of cytokine synthesis. Here we have studied serum IL-6 and TNF levels in mice after LPS administration. Repeated administration of LPS (35 micrograms daily for 4 days) to mice induced a refractoriness (tolerance) to subsequent administrations of LPS in terms of induction of circulating IL-6 and TNF. To investigate the mechanism by which LPS down-regulates its own induction of cytokine synthesis and the relationship between IL-6 and TNF production, we attempted to revert the inhibition of IL-6 and TNF production using agents like PMA or IFN-gamma, previously reported to activate macrophage production of cytokines. Pretreatment with PMA (4 micrograms, 10 min before LPS) partially restored IL-6 production in LPS-tolerant mice given 2 micrograms LPS. On the other hand, PMA did not restore TNF induction in LPS-tolerant mice, even when administered with high doses of LPS (up to 200 micrograms). A similar reversal of LPS resistance to IL-6, but not TNF, induction by PMA was observed in genetically LPS-resistant C3H/HeJ mice. IFN-gamma also restored, although to a lesser extent than PMA, IL-6 production. However, unlike PMA, IFN-gamma could also partially restore TNF production in LPS-tolerant mice, although only when LPS was administered at high doses. By contrast with PMA, IFN-gamma was clearly more active in restoring TNF synthesis than that of IL-6. Similar results were obtained in genetically LPS-unresponsive C3H/HeJ mice. These data suggest that different mechanisms are implicated in the inhibition of IL-6 and TNF synthesis in LPS-tolerant mice and that part of this inhibition can be overcome by PMA or IFN-gamma.     

Expression of a 120,000 dalton protein during tumoricidal activation in murine peritoneal macrophages

Modulation of protein expression during interferon-gamma (IFN-gamma)-lipopolysaccharide (LPS)-mediated macrophage tumoricidal activation has been examined by metabolic radiolabeling of various murine peritoneal macrophage populations with [35S]methionine followed by SDS-PAGE analysis. Although both IFN-gamma and LPS are capable of stimulating the expression of several proteins when used independently, combined treatment induced the enhanced or de novo expression of a 120,000 dalton polypeptide. The expression of this protein was synergistically regulated by both IFN-gamma and LPS in a manner strongly reminiscent of the functional synergism that these two agents exhibit with respect to induction of tumoricidal activity. p120 expression could be seen first at approximately 3 hr after the addition of both agents, reached optimal expression by 6 hr, and maintained elevated synthesis for up to 24 hr. This time course corresponds closely to that seen for the acquisition of tumoricidal competence. Macrophages elicited in the primed state of activity in vivo with methyl vinyl ether co-polymer II (MVE-II) did not express p120, but could be induced to do so when treated with low doses of LPS. Under similar conditions, MVE-II-elicited cells also acquire tumoricidal activity. Macrophages obtained from mice chronically infected with bacillus Calmette-Guerin constitutively expressed both p120 and cytolytic activity. If such macrophages were cultured for 24 hr, the expression of both events decayed and was lost, but could be restored by treatment with low doses of LPS. Thus the data support a strong correlation between the expression by macrophages of a novel 120,000 dalton protein and the expression of tumor cytotoxicity.     

Regulation of tumor necrosis factor (TNF) expression: interferon-gamma enhances the accumulation of mRNA for TNF induced by lipopolysaccharide in murine peritoneal macrophages

The secretion of tumor necrosis factor (TNF) by macrophages is initiated by lipopolysaccharide (LPS); considerable evidence indicates that such secretion can be potentiated by interferon-gamma (IFN-gamma). The present studies show that accumulation of mRNA for tumor necrosis factor, which represents an important regulatory focus for controlling secretion of TNF, is enhanced by physiologic doses of IFN-gamma (20 units/ml of purified recombinant IFN-gamma). mRNA for TNF induced by LPS, which was maximal 2 hr after LPS was applied to the cells, was enhanced 5- to 8-fold by IFN-gamma as determined by Northern blot analysis. Interferon did not change the kinetics of accumulation but did change the dose effects of LPS in that increasing amounts of LPS led to increasing amounts of TNF mRNA in IFN-gamma-treated macrophages. IFN-gamma itself, however, did not induce expression of TNF mRNA. These studies document that IFN-gamma potentiates the cytoplasmic accumulation of mRNA for TNF induced in murine peritoneal macrophages by LPS.     

IL-18-binding protein protects against lipopolysaccharide- induced lethality and prevents the development of Fas/Fas ligand-mediated models of liver disease in mice.

IL-18-binding protein (IL-18BP) is a natural IL-18 inhibitor. Human IL-18BP isoform a was produced as fusion construct with human IgG1 Fc and assessed for binding and neutralizing IL-18. IL-18BP-Fc binds human, mouse, and rat IL-18 with high affinity (K(D) 0.3-5 nM) in a BIAcore-based assay. In vitro, IL-18BP-Fc blocks IL-18 (100 ng/ml)-induced IFN-gamma production by KG1 cells (EC(50) = 0.3 microg/ml). In mice challenged with an LD(90) of LPS (15 mg/kg), IL-18BP-Fc (5 mg/kg) administered 10 min before LPS blocks IFN-gamma production and protects against lethality. IL-18BP-Fc administered 10 min before LPS blocks IFN-gamma production induced by LPS (5 mg/kg) with ED(50) of 0.005 mg/kg. Furthermore, IL-18BP-Fc (5 mg/kg) abrogates LPS (5 mg/kg)-induced IFN-gamma production even when administered 6 days before LPS but shows no effect when administered 9 or 12 days before LPS. Given 10 min before LPS challenge to mice primed 12 days in advance with heat-killed Propionibacterium acnes, IL-18BP-Fc prevents LPS-induced liver damage and IFN-gamma and Fas ligand expression. Given at the moment of priming with P. acnes, IL-18BP-Fc decreases P. acnes-induced granuloma formation, macrophage-inflammatory protein-1alpha and macrophage-inflammatory protein-2 production and prevents sensitization to LPS. IL-18BP-Fc also prevents Con A-induced liver damage and IFN-gamma and Fas ligand expression as well as liver damage induced by Pseudomonas aeruginosa exotoxin A or by anti-Fas agonistic Ab. In conclusion, IL-18BP can be engineered and produced in recombinant form to generate an IL-18 inhibitor, IL-18BP-Fc, endowed with remarkable in vitro and in vivo properties of binding and neutralizing IL-18.     

Somatic cell hybrids between macrophages from Bcgr and Bcgs mice: characterization of MHC class II expression

In order to gain a better understanding of the regulation of MHC class II expression related to the Bcg gene, we have produced macrophage-macrophage somatic cell hybrids by fusing the RAW 309 macrophage cell line derived from BALB/c.Bcgs mice with peritoneal macrophages from Bcgr C3H/HeN mice. The differential screening of the hybrids was based on the differential sensitivity of Ia expression to suppression with cycloheximide. We found that most of the hybrids expressed Ia without further stimulation with rIFN-gamma. Cycloheximide suppressed the expression of Ia by some of the hybrids. Treatment of these cells with rIFN-gamma resulted in a cycloheximide resistant Ia expression of both parental haplotypes. The macrophage hybrids produced IL-1 beta, IL-1 alpha, and TNF-alpha when stimulated with LPS. There was no correlation between the levels of monokines produced and the persistence of Ia expression. The results of this investigation indicate that the product of the Bcg gene contributed by macrophages from C3H/HeN mice will affect the expression of the I-Ad glycoprotein that is normally transiently expressed by the RAW 309 cell line.     

Role of TNF-alpha in the induction of fungicidal activity of mouse peritoneal exudate cells against Cryptococcus neoformans by IL-12 and IL-18

We have recently demonstrated that two IFN-gamma-inducing cytokines, interleukin (IL)-12 and IL-18, synergistically induced the fungicidal activity of mouse peritoneal exudate cells (PEC) against Cryptococcus neoformans through NK cell production of interferon (IFN)-gamma and nitric oxide (NO) synthesis. In the present study, we further dissected these effects by examining the involvement of tumor necrosis factor (TNF)-alpha in the induction of IL-12/IL-18-stimulated PEC fungicidal activity. The addition of neutralizing anti-TNF-alpha mAb significantly suppressed IL-12/IL-18-stimulated PEC anticryptococcal activity. This effect was ascribed to the inhibition of macrophage NO synthesis, but not of IFN-gamma production by NK cells, because the same treatment inhibited the former response, but not the latter one. On the other hand, combined treatment with IL-12 and IL-18 synergistically induced the production of TNF-alpha by PEC and this effect was almost completely abrogated by neutralizing anti-IFN-gamma mAb. The cell type producing TNF-alpha among PEC was mostly macrophage. TNF-alpha significantly promoted macrophage NO production and anticryptococcal activity induced by IFN-gamma, and furthermore anti-TNF-alpha mAb partially inhibited these responses. Considered together, our results indicated that TNF-alpha contributed to the potentiation of IL-12/IL-18-induced PEC fungicidal activity against C. neoformans through enhancement of IFN-gamma-induced production of NO by macrophages, but not through increased production of IFN-gamma by NK cells.     

Recombinant IFN-gamma synergizes with lipopolysaccharide to induce macrophage membrane procoagulants

Fibrin deposition is an important histopathologic feature of inflammation and is mediated, in part, by monocyte/macrophage procoagulants. rIFN gamma acted in synergy with suboptimal levels of bacterial LPS by priming thioglycollate-induced mouse peritoneal exudate cells (TG-PEC) to express high levels of surface procoagulant. TFN-alpha beta, TFN-alpha, IL-1, either alone or in combination with LPS or IFN-gamma, had no effect on macrophage procoagulant activity expression. In contrast to the dramatic increases of macrophage procoagulant activity induced by IFN-gamma/LPS, on exudate macrophages, normal peritoneal macrophages, or peripheral blood monocytes were unresponsive suggesting that the state of activation of the macrophage determines reactivity. IFN-gamma induced a Factor VIIa-like activity detected only after cell disruption. Synergy between LPS and IFN-gamma-induced procoagulants may occur as the result of the assembly of the thromboplastin (induced by LPS), Factor VII/VIIa complex on the macrophage surface. RNA synthesis was required for procoagulant induction. Procoagulant expression may, as for other cytokines involved in inflammatory responses, be regulated by short lived repressor proteins as low dose cycloheximide superinduced procoagulant responses to both LPS and IFN-gamma and caused the extracellular expression of procoagulant in response to IFN-gamma. This study suggests an important role for IFN-gamma in the assembly of components of the extrinsic coagulant cascade on the macrophage surface. The synergy between IFN-gamma and LPS may moderate macrophage-initiated fibrin deposition characteristic of inflammatory responses.     

Lancemaside A inhibits lipopolysaccharide‐induced inflammation by targeting LPS/TLR4 complex

In our previous study, lancemaside A isolated from Codonopsis lanceolata (family Campanulaceae) ameliorated colitis in mice. In this study, the anti‐inflammatory effects of lancemaside A was investigated in lipopolysaccharide (LPS)‐stimulated mice and their peritoneal macrophage cells. Lancemaside A suppressed the production of pro‐inflammatory cytokines, TNF‐α and IL‐1β, in vitro and in vivo. Lancemaside A also down‐regulated inducible nitric oxide synthase (iNOS) and cyclooxygenase‐2 (COX‐2), as well as the inflammatory mediators, nitric oxide (NO), and PGE₂. Lancemaside A also inhibited the expression of IL‐1 receptor‐associated kinase‐4 (IRAK‐4), the phosphorylation of IKK‐β and IκB‐α, the nuclear translocation of NF‐κB and the activation of mitogen‐activated protein kinases in LPS‐stimulated peritoneal macrophages. Furthermore, lancemaisde A inhibited the interaction between LPS and TLR4, as well as IRAK‐4 expression in peritoneal macrophages. Based on these findings, lancemaside A expressed anti‐inflammatory effects by regulating both the binding of LPS to TLR4 on macrophages. J. Cell. Biochem. 111: 865–871, 2010. © 2010 Wiley‐Liss, Inc.     

Eosinophils and immune mechanisms. III. Production of the lymphokine eosinophil stimulation promoter by mouse T lymphocytes.

The lymphoid cell population responsible for production of eosinophil stimulation promoter (ESP), a lymphokine which increases migration of eosinophils, was investigated in murine Schistosoma mansoni infection. Con A challenge induced ESP production, whereas LPS did not. Prior treatment with anti-thetaC3H alloantiserum plus complement in vitro eliminated ESP production; in vivo treatment with rabbit anti-mouse thymocyte serum consistently reduced ESP production by splenic lymphoid cells, but affected lymph node cell ESP production only after exceptionally large doses. Thymocytes did not produce significant amounts of ESP; nor did lymphoid cells from congenitally athymic mice. Depletion of B lymphocytes and macrophages by nylon fiber adherence eliminated antigen-induced ESP production; this was partially restored by addition of non-immune, 72-hr peritoneal exudate cells. Con A-induced ESP production was not affected by nylon fiber treatment. These results demonstrate that ESP is produced by an ATS-sensitive, peripheralized T lymphocyte population, and suggest a macrophage requirement for antigen-induced production of this lymphokine.     

Potentiation of lymphokine-induced macrophage activation by tumor necrosis factor-alpha

In this study, we examined the possible role of TNF-alpha and lymphotoxin (TNF-beta) as cofactors of macrophage activation. The results demonstrate that both TNF were capable of enhancing the cytostatic and cytolytic activity of murine peritoneal macrophages against Eb lymphoma cells. The potentiation of tumor cytotoxicity became apparent when macrophages from DBA/2 mice were suboptimally activated by either a T cell clone-derived macrophage-activating factor or by IFN-gamma plus LPS. Neither TNF-alpha nor TNF-beta could induce tumor cytotoxicity in IFN-gamma-primed macrophages, indicating that TNF cannot replace LPS as a triggering signal of activation. In LPS-resistant C3H/HeJ macrophages, which were unresponsive to IFN-gamma plus LPS, a supplementation with TNF fully restored activation to tumor cytotoxicity. Furthermore, TNF-alpha potentiated a variety of other functions in low-level activated macrophages such as a lactate production and release of cytotoxic factors. At the same time, TNF-alpha produced a further down-regulation of pinocytosis, tumor cell binding and RNA synthesis observed in activated macrophages. These data demonstrate new activities for both TNF-alpha and TNF-beta as helper factors that facilitate macrophage activation. In particular, the macrophage product TNF-alpha may serve as an autocrine signal to potentiate those macrophage functions that were insufficiently activated by lymphokines.     

Induction of macrophage Ia antigen expression by rIFN-gamma and down-regulation by IFN-alpha/beta and dexamethasone are mediated by changes in steady-state levels of Ia mRNA

In previous studies, the induction of Ia antigens on murine peritoneal exudate macrophages by recombinant IFN-gamma (rIFN-gamma) and the antagonism of rIFN-gamma-induced Ia expression by the inhibitors IFN-alpha/beta and glucocorticoids have been examined. In this report, these findings have been extended to an analysis of total or cytoplasmic mRNA from macrophage cultures treated with rIFN-gamma in the absence or presence of these two inhibitors. Recombinant IFN-gamma induced a 5.7- to 6.5-fold increase in steady-state levels of Ia (A alpha-specific) mRNA. Coordinate increases in steady-state mRNA for A beta, and E alpha were observed in response to rIFN-gamma. Maximum induction occurred 24 hr post-treatment and required the continued presence of rIFN-gamma. Induction of A alpha-specific mRNA was sensitive to the protein synthesis inhibitor cycloheximide. Simultaneous treatment of macrophage cultures with rIFN-gamma and IFN-alpha/beta or the glucocorticoid dexamethasone (DEX) resulted in a significant decrease in steady-state, A alpha-specific mRNA levels compared with treatment with rIFN-gamma alone. This analysis suggests that both the induction of Ia expression by rIFN-gamma, and the antagonism of rIFN-gamma-induced Ia gene expression by IFN-alpha/beta and DEX, are regulated by cognate changes in Ia mRNA.     

Regulation of macrophage activation by IL-3. II. IL-3 and lipopolysaccharide act synergistically in the regulation of IL-1 expression

We have previously reported that IL-3, a cytokine produced by both Th1 and Th2 type CD4+ T cells, displays macrophage-activating potential. IL-3, like IFN-gamma, readily induced functions related to Ag presentation (e.g., Ia and lymphocyte function-associated Ag-1 expression). However, in contrast to the response elicited by IFN-gamma, tumor cytotoxicity was not induced by IL-3. In this paper we have evaluated the capacity of IL-3 to regulate IL-1 expression. Our data demonstrate that although IL-3 alone was unable to induce the production of substantial IL-1 bioactivity in peritoneal exudate cells, it contributed synergistically to the induction of IL-1 bioactivity in the presence of suboptimal doses of LPS. It was of interest that IFN-gamma, which can also interact synergistically with LPS, was unable to complement the partial signals provided by IL-3 for the expression of IL-1 bioactivity, suggesting that IL-3 and IFN-gamma may be providing similar stimulatory signals in this respect. Our studies on the mechanism of synergy between IL-3 and LPS indicated that the effect of LPS did not appear to be mediated by the well-characterized LPS-inducible cytokines of macrophage origin (i.e., IL-1, alpha and beta, TNF-alpha, and IL-6). The best characterized function of IL-3 is its multicolony-stimulating activity as a CSF; in this context we also studied granulocyte-macrophage CSF and noted that it behaves similarly to IL-3 in that it can synergistically contribute to IL-1 induction. A similar, but more dramatic induction of IL-1 synthesis in response to IL-3 was demonstrated by the P388.D1 murine macrophage cell line. The kinetics and the molecular mechanism of the response of P388.D1 to IL-3 indicate several unique features of IL-3-induced IL-1 expression: 1) IL-3 itself induced IL-1 mRNA expression, which was unaccompanied by substantial production of bioactivity, either cell-associated or secreted into the culture supernatant; 2) IL-3 synergized with suboptimal doses of LPS to induce not only heightened IL-1 mRNA levels but bioactivity as well; and 3) IL-3, when combined with LPS, altered the kinetics of IL-1 message and bioactive protein production in response to LPS: IL-3 and LPS induced an early release (3 to 7 h poststimulation) of the IL-1 protein as well as a second peak of mRNA and bioactivity (at 12 to 36 h), which was not observed in response to either IL-3 or LPS alone.(ABSTRACT TRUNCATED AT 400 WORDS)