Preexposure of macrophages to low doses of lipopolysaccharide inhibits the expression of tumor necrosis factor-alpha mRNA but not of IL-1 beta mRNA

LPS is known to be a potent activator of macrophages and induces the production of TNF-alpha and IL-1. However, the signaling events and regulatory mechanisms required for the activation of macrophages by LPS have not been resolved precisely. We show that LPS modulates its own response in macrophages. Proteose peptone-induced murine peritoneal macrophages (P-PEM) produce significant amount of TNF-alpha and IL-1 after stimulation with LPS. However, preexposure of macrophages to low doses (less than 1 ng/ml) of LPS renders them refractory to stimulation by a second round of LPS, as evaluated by production of TNF-alpha. The loss of sensitivity to a second round of LPS was selective for TNF-alpha production as the LPS-primed macrophages retained the ability to produce IL-1. Northern blot analysis was performed with total RNA obtained from control and LPS- (1 ng/ml) primed P-PEM after 3-h stimulation with a second round of LPS. The expression of TNF-alpha mRNA was inhibited in LPS-primed P-PEM, whereas the expression of IL-1 beta mRNA was the same in control and LPS-primed P-PEM, consistent with the data of biologic activities of these two cytokines. Zymosan-induced TNF-alpha production was the same in control and LPS-primed macrophages, indicating that not all of the pathways required for TNF-alpha production were affected by LPS priming. Monokines such as human (h) rIL-1 alpha, hrTNF-alpha, hrIL-6, and murine rIFN-beta could not substitute for the action of low doses of LPS, and addition of indomethacin could not restore TNF-alpha production. These results suggest that exposure of macrophages to low doses of LPS suppresses the production of TNF-alpha, but not of IL-1, by inhibiting the expression of mRNA through a noncyclooxygenase-dependent mechanism. Thus, LPS-induced production of TNF-alpha and IL-1 in macrophages are differently regulated.     

趋化因子CXCL10在心肌细胞及巨噬细胞中的表达机制

目的探讨心肌缺血-再灌注损伤中趋化因子CXCL10的产生机制。方法分别用LPS、H2O2、Ca2+载体A23187刺激原代培养的心肌细胞、骨髓来源的巨噬细胞或二者混合培养的共培养系统后,ELISA检测培养基上清中的趋化因子CXCL10和促炎性细胞因子IL-1β、IL-6、TNF-α的含量,观察其表达动力学。结果①大剂量(10μg/mL)的LPS刺激心肌细胞主要产生趋化因子CXCL10;刺激骨髓来源巨噬细胞主要产生促炎性细胞因子IL-1β、IL-6、TNF-α。②H2 O2、Ca2+通道激活剂并不能使产生趋化因子CXCL10或IL-1β、IL-6、TNF-α这些促炎性细胞因子。③骨髓来源的巨噬细胞促进心肌细胞表达趋化因子CXCL10;心肌细胞促进骨髓来源的巨噬细胞表达IL-6、TNF-α,但抑制IL-1β的表达。结论心肌细胞是心肌缺血-再灌注损伤中CXCL10潜在的细胞来源;CXCL10的表达,主要依赖于TLR4的激活。     

Quantitative aspects of lipopolysaccharide and cytokine requirements to generate nitric oxide in macrophages from LPS-hyporesponsive (<Emphasis Type="Italic">Lps</Emphasis><Superscript>d</Superscript>) C3H/HeJ mice

Due to a gene defect (Lps(d)), C3H/HeJ mice are known to be hyporesponsive to the immunobiological potential of lipopolysaccharide (LPS). We studied dose requirements for LPS, IFN-gamma, and cytokines TNF-alpha and IL-10 to produce nitric oxide (NO) in peritoneal macrophages (Mphi) from these animals. In contrast to the Lps(n) C3H/HeN mice, high concentrations of LPS (up to 5 microg/mL) or IFN-gamma (up to 5 ng/mL) by themselves were unable to activate NO production in C3H/HeJ Mphi. The failure to produce NO could not be overcome by addition of L-arginine or tetrahydropterin. The high-output NO biosynthesis was dose-dependently stimulated by combined administration of varying concentrations of IFN-gamma (50-5000 pg/mL) and LPS (approximately 1 ng/mL) or to a lesser extent by IFN-gamma plus TNF-alpha or TNF-alpha/IL-10. Formation of NO in C3H/HeJ MCO triggered by high concentration of LPS (approximately 1 microg/mL) given together with IFN-gamma (0.2-5 ng/mL) reached the values typical for Lps(n) C3H/HeN mice. While Mphi from C3H/HeN mice secreted TNF-alpha, IL-10, and IL-10 upon contact with a low dose of LPS (1 ng/mL), C3H/HeJ Mphi required high concentration of LPS (5 microg/mL) to enhance the secretion of the cytokines. Yet, this dose remained ineffective to stimulate IFN-gamma in Mphi from C3H/HeJ mice. It can be presumed that one of the important factors influencing their deficient ability to form NO is a failure of Mphi to produce IFN-gamma upon LPS contact.     

Inflammatory cytokine production in interferon-gamma-primed mice, challenged with lipopolysaccharide. Inhibition by SK&F 86002 and interleukin-1 beta-converting enzyme inhibitor

Mice challenged with lipopolysaccharide (LPS) produce variable serum levels of pro-inflammatory cytokines, and particularly low levels of interleukin-1 beta (IL-1 beta). Interferon-gamma (IFN-gamma) has been shown to be an important mediator of bacteria-induced hypersensitivity to LPS in mice. In the present study, we show that mice pretreated with IFN-gamma exhibit an enhanced capacity to produce serum IL-1 beta, IL-1 alpha, tumour necrosis factor (TNF-alpha) as well as IL-6 in response to LPS. Priming with intraperitoneal (i.p.) injection of 15 mg rat recombinant IFN-gamma, 18 hours prior to the i.p. LPS (300 mg) challenge resulted in a 4-fold increase in the LPS-stimulated release of IL-1 beta and a 2- to 7-fold increase in the release of IL-1 alpha, TNF-alpha, as well as IL-6 into the serum. LPS induced a concentration-dependent increase in the release of IL-1 beta in isolated peritoneal macrophages from IFN-gamma-primed mice whereas macrophages from unprimed mice released minute amounts of IL-1 beta. In addition, nigericin markedly enhanced the release of IL-1 beta in unprimed mice but not in macrophages from IFN-gamma primed mice. The cytokine synthesis inhibitor SK&F 86002, administered per os (100 mg/kg), 1 hour prior to LPS challenge, strongly inhibited the rise in serum levels of the four cytokines. Furthermore, treatment with the IL-1 beta converting enzyme (ICE) specific reversible inhibitor YVAD-CHO resulted in a sharp dose- and time-dependent inhibition of IL-1 beta secretion in the serum, whereas the other cytokines were not affected. In conclusion, IFN-gamma priming strongly potentiates the release of proinflammatory cytokines in the serum of mice as compared to LPS stimulation alone, and provides therefore a useful way to test the in vivo potency and selectivity of cytokine synthesis inhibitors.     

Induction of interleukin-1alpha production in murine Sertoli cells by interleukin-1

In the present study we examined the involvement of interleukin (IL)-1alpha, -1beta, FSH, and lipopolysaccharide (LPS) in the regulation of IL-1alpha and -1beta production by Sertoli cells under in vitro conditions. Sertoli cell cultures from immature mice produced constitutively basal levels of intracellular IL-1alpha. Stimulation of Sertoli cell cultures with LPS (5 microgram/ml) resulted in a maximal production of intracellular IL-1alpha 2 h after the stimulation. Thereafter, these levels decreased but remained significantly higher within 24 h after stimulation than those in control cultures. The effect of LPS on IL-1alpha production was dose dependent. FSH did not show any effect on intracellular IL-1alpha production by Sertoli cells. IL-1alpha could not be detected in supernatants of unstimulated or stimulated Sertoli cell cultures. Sertoli cell cultures stimulated with recombinant IL-1alpha induced optimal intracellular levels of IL-1alpha within 2 h of stimulation. These levels remained high 24 h after stimulation. However, stimulation of Sertoli cell cultures with IL-1beta induced a peak of IL-1alpha production 8 h after stimulation. These levels decreased 24 h after the stimulation but were still found to be significantly higher than those in control cultures. The addition of IL-1 receptor antagonist (IL-1ra) to Sertoli cell cultures did not significantly alter their capacity to produce IL-1alpha. However, the stimulatory effects of recombinant IL-1alpha on IL-1alpha production by Sertoli cell cultures were reversed by the concomitant addition of recombinant IL-1ra. No immunoreactive IL-1beta could be detected in lysates or conditioned media of immature murine Sertoli cells under any of the stimulatory conditions outlined. Our results may suggest the involvement of physiological (IL-1) and pathophysiological factors (LPS) in the regulation of spermatogenesis and spermiogenesis processes and male fertility.     

Ontogeny of rat pulmonary alveolar macrophage function: evidence for a selective deficiency in il-10 and nitric oxide production by newborn alveolar macrophages

Alveolar macrophages (AM) play a crucial role in host defence by secretion of a large repertoire of biological response modifiers (BRM) following challenge. Newborns manifest increased susceptibility to lung infections, suggesting a deficiency in AM-mediated host defence. Thus, we investigated the ontogeny of BRM production by resting and stimulated AM. We analysed the capacity of rat AM to produce mRNA specific for a range of cytokines including tumour necrosis factor-alpha (TNF-alpha), interleukin (IL)-1beta, IL-6, IL-10, IL-12, IL-18, and the enzyme inducible nitric oxide synthase, in response to in vitro lipopolysaccharide (LPS) challenge. We report that production of nitric oxide by newborn AM under conditions of maximal stimulation was impaired. In addition, expression of IL-10 was only minimally upregulated in AM from newborns in response to LPS compared to adults. Inability to upregulate expression of IL-10 appeared to be influenced by microenvironmental factors, since peritoneal macrophages from newborns responded to LPS with significant upregulation of IL-10. Furthermore, when newborn AM were precultured in vitro, IL-10 responsiveness to LPS was partially restored. In contrast, cytokines such as TNF-alpha, IL-1, IL-6, IL-12 and IL-18 appeared to be expressed at adult levels by newborn AM. These results demonstrate that there may be functional differences in AM of newborns compared to adults, and these may be specific to the tissue compartment.     

Murine eosinophils and IL-1: alpha IL-1 mRNA detection by in situ hybridization. Production and release of IL-1 from peritoneal eosinophils

The presence of IL-1 mRNA in eosinophils from mice infected with larvae of the parasite Mesocestoides corti was investigated by in situ hybridization technique. S35 labeled cDNA probe for alpha IL-1, hybridized with mRNA in murine eosinophils and macrophages. After 6 h of LPS stimulation eosinophils were able to express mRNA in their cytoplasm. This expression was highly increased by the addition of indomethacine. The IL-1 mRNA expression in murine macrophages was higher than in eosinophils in LPS-stimulated cells. This difference was statistically significant, p less than 0.001. To test if eosinophils may produce and release IL-1 in the culture medium, we isolated these cells in a Percoll gradient. Cell preparations with a purity exceeding 94% were cultured with various stimuli and their supernatants were tested for IL-1 activity. Eosinophils produced 169.65 +/- 73 U/ml when stimulated with LPS (n = 14). A dose-dependent response was obtained when the eosinophils were in the presence of the calcium ionophore A23187. Controls were performed to rule out the contribution of the contaminating population on the thymocyte proliferating activity. They were also used to detect other possible causes of interference in the assay, such as leukotrienes or TNF. IL-1 in supernatants was also detected using a conversion assay such as EL-4 thymoma cells. IL-1 activity was first detected in culture supernatants 18 h later, maximal production being in the first 24 h. In accordance with our hybridization results, an increase in IL-1 activity was obtained when eosinophils were stimulated with LPS and treated with indomethacine. The factor had a molecular mass between 16 to 20 kDa that corresponded to the described for murine IL-1. Inasmuch as IL-1 is an important mediator of inflammatory reactions this IL may enhance the proinflammatory action of eosinophils.     

Tumor necrosis factor-alpha production by astrocytes. Induction by lipopolysaccharide, IFN-gamma, and IL-1 beta

Astrocytes have the capacity to secrete or respond to a variety of cytokines including IL-1, IL-6, IL-3, and TNF-alpha. In this study, we have examined the capacity of astrocytes to secrete TNF-alpha in response to a variety of biologic stimuli, particularly cytokines such as IL-1 and IFN-gamma, which are known to be present in the central nervous system during neurologic diseases associated with inflammation. Rat astrocytes do not constitutively produce TNF-alpha, but have the ability to secrete TNF-alpha in response to LPS, and can be primed by IFN-gamma to respond to a suboptimal dose of LPS. IFN-gamma and IL-1 beta alone do not induce TNF-alpha production, however, the combined treatment of IFN-gamma and IL-1 beta results in a striking synergistic effect on astrocyte TNF-alpha production. Astrocyte TNF-alpha protein production induced by a combined treatment of either IFN-gamma/LPS or IFN-gamma/IL-1 beta occurs in a dose- and time-dependent manner, and appears to require a "priming signal" initiated by IFN-gamma, which then renders the astrocyte responsive to either a suboptimal dose of LPS or IL-1 beta. Astrocyte TNF-alpha production by IFN-gamma/LPS stimulation can be inhibited by the addition of anti-rat IFN-gamma antibody, whereas IFN-gamma/IL-1-induced TNF-alpha production is inhibited by antibody to either IFN-gamma or IL-1 beta. Polyclonal antisera reactive with mouse macrophage-derived TNF-alpha neutralized the cytotoxicity of IFN-gamma/LPS and IFN-gamma/IL-1 beta-induced astrocyte TNF-alpha, demonstrating similarities between these two sources of TNF-alpha. We propose that astrocyte-produced TNF-alpha may have a pivotal role in augmenting intracerebral immune responses and inflammatory demyelination due to its diverse functional effects on glial cells such as oligodendrocytes and astrocytes themselves.     

Interferon gamma priming is not critical for IL-12 production of murine spleen cells

Interferon gamma (IFN-gamma) priming is considered to be critical for interleukin 12 (IL-12) production of murine macrophages and human monocytes by lipopolysaccharide (LPS) stimulation. In our present experiments, freshly prepared spleen cells (f-spleen cells) were confirmed not to produce detectable level of IL-12 by LPS stimulation, although they produced significant amount of IL-12 by the stimulation with LPS plus IFN-gamma. However, the stimulation only with LPS induced IL-12 production of spleen cells preincubated in the absence of IFN-gamma. Findings on IL-12 p40 mRNA accumulation were consistent with their IL-12 production. Essentially the same results were obtained using spleen cells from IFN-gamma deficient mice. In the presence of anti-IL-10, f-spleen cells produced IL-12 upon LPS stimulation, indicating that the failure of f-spleen cells in IL-12 production is caused by IL-10 produced by themselves upon LPS stimulation. In addition, f-spleen cells produced IL-12 upon CD40 ligand stimulation, and the production was hardly affected by the presence of IFN-gamma or preincubation. These results indicate that IFN-gamma priming is not critical for IL-12 production of spleen cells stimulated with LPS or CD40 ligand, although IFN-gamma enhances the production, especially, in response to LPS stimulation.     

IL-10 inhibits cytokine production by activated macrophages

IL-10 inhibits the ability of macrophage but not B cell APC to stimulate cytokine synthesis by Th1 T cell clones. In this study we have examined the direct effects of IL-10 on both macrophage cell lines and normal peritoneal macrophages. LPS (or LPS and IFN-gamma)-induced production of IL-1, IL-6, and TNF-alpha proteins was significantly inhibited by IL-10 in two macrophage cell lines. Furthermore, IL-10 appears to be a more potent inhibitor of monokine synthesis than IL-4 when added at similar concentrations. LPS or LPS- and IFN-gamma-induced expression of IL-1 alpha, IL-6, or TNF-alpha mRNA was also inhibited by IL-10 as shown by semiquantitative polymerase chain reaction or Northern blot analysis. Inhibition of LPS-induced IL-6 secretion by IL-10 was less marked in FACS-purified peritoneal macrophages than in the macrophage cell lines. However, IL-6 production by peritoneal macrophages was enhanced by addition of anti-IL-10 antibodies, implying the presence in these cultures of endogenous IL-10, which results in an intrinsic reduction of monokine synthesis after LPS activation. Consistent with this proposal, LPS-stimulated peritoneal macrophages were shown to directly produce IL-10 detectable by ELISA. Furthermore, IFN-gamma was found to enhance IL-6 production by LPS-stimulated peritoneal macrophages, and this could be explained by its suppression of IL-10 production by this same population of cells. In addition to its effects on monokine synthesis, IL-10 also induces a significant change in morphology in IFN-gamma-stimulated peritoneal macrophages. The potent action of IL-10 on the macrophage, particularly at the level of monokine production, supports an important role for this cytokine not only in the regulation of T cell responses but also in acute inflammatory responses.     

Production of pro- and anti-inflammatory cytokines by monocytes from patients with paracoccidioidomycosis

Monocytes and macrophages can produce a large repertoire of cytokines and participate in the pathogenesis of granulomatous diseases. We investigated the production of pro- and anti-inflammatory cytokines by monocytes from patients with active paracoccidioidomycosis. Peripheral blood monocytes from 37 patients and 29 healthy controls were cultivated with or without 10 microg/ml of lipopolysaccharide (LPS) for 18 h at 37 degrees C, and the cytokine levels were determined in the culture supernatants by enzyme immunoassay. The results showed that the endogenous levels of tumor necrosis factor alpha (TNF-alpha), interleukin-1beta (IL-1beta), IL-6, IL-8, IL-10 and transforming growth factor beta detected in the supernatant of patient monocytes cultivated without stimulus were significantly higher than those produced by healthy controls. These data demonstrated that monocytes from patients with active paracoccidioidomycosis produce high levels of cytokines with both inflammatory and anti-inflammatory activities. However, patient monocytes produced significantly lower TNF-alpha and IL-6 levels in response to LPS when compared to normal subjects, suggesting an impairment in their capacity to produce these cytokines after LPS stimulation. Concentrations of IL-1beta, IL-8 and IL-10 in cultures stimulated with LPS were higher in patients than in controls. These results suggest that an imbalance in the production of pro- and anti-inflammatory cytokines might be associated with the pathogenesis of paracoccidioidomycosis.     

Interleukin-1 (IL-1) receptor blockade reduces endotoxin and Borrelia burgdorferi-stimulated IL-8 synthesis in human mononuclear cells.

Interleukin-1 (IL-1) is a potent stimulator of IL-8 production by fibroblasts and monocytes. In the present study, we asked how much of endotoxin (LPS)-induced IL-8 production by human peripheral blood mononuclear cells was due to IL-1 induced by LPS. Cells were stimulated with either IL-1 beta, LPS, or Borrelia burgdorferi, and total IL-8 was determined by a specific radioimmunoassay. The addition of saturating concentrations of IL-1 receptor antagonist protein (IRAP) reduced the IL-1 beta-, LPS-, and B. burgdorferi-induced IL-8 synthesis by 85, 50, and 40%, respectively. Increasing the concentration of LPS did not affect the reduction in IL-8 synthesis observed in the presence of IRAP. Significant inhibition of the IL-1 beta-induced IL-8 synthesis was observed when IRAP was added 60 or 90 min after IL-1 beta; similarly, IL-8 synthesis after LPS was also reduced by delayed addition of IRAP. These data suggest that the ameliorative effects of IL-1 receptor blockade in models of inflammation and infection may be due, in part, to suppression of IL-1-induced IL-8.     

IL-2 induction of IL-1 beta mRNA expression in monocytes. Regulation by agents that block second messenger pathways

We have previously shown that in mixed cultures of PBL incubation with human rIL-2 induces the rapid expression of IL-1 alpha and IL-1 beta mRNA. Because studies have demonstrated that IL-2R can be expressed on the surface of human peripheral blood monocytes, we chose to investigate whether IL-1 beta mRNA could be directly induced in purified human monocytes by treatment with Il-2 and, if so, to analyze the second messenger pathways by which it may be controlled. Human monocytes do not spontaneously express IL-1 beta mRNA, but can express the gene as soon as 1 h after treatment with IL-2. The level of IL-1 beta mRNA induced by IL-2 at 5 h in human monocytes was about one-fourth that induced by LPS. LPS induction of IL-1 beta mRNA in human monocytes can be blocked by either an inhibitor of protein kinase C (PKc) 1-(5-isoquinolinesulfonyl)-2-methylpiperazine or an inhibitor of calcium/calmodulin (CaM) kinase N-(6-aminohexyl) 5-chloro-1-naphthalenesulfonamide, suggesting that both PKc and CaM kinase are involved in transducing signals initiated by LPS. In contrast, IL-2 induction of IL-1 beta mRNA expression is blocked only by 1-(5-isoquinolinesulfonyl)-2-methylpiperazine, suggesting that PKc, and not CaM kinase, is activated by IL-2. These data suggest that overlapping but distinct second messenger pathways are involved in the transduction of signals initiated by IL-2 and LPS.     

Cytokine production by mature and immature thymocytes.

We have studied the ability of subpopulations of activated thymocytes to produce four cytokines (IL-2, IL-4, IFN-gamma and TNF-alpha) which are believed to play roles in T cell development. Supernatants from various thymocyte subsets activated with calcium ionophore and PMA were tested for these cytokines. All CD3hi thymocyte subsets (CD4+8-, CD4-8- and CD4-8+) produced high titers of these four cytokines except CD3+4-8+ thymocytes, which did not produce IL-4. In contrast, CD4+8+ thymocytes did not produce any detectable cytokines. CD3-4-8- thymocytes produced IL-2, IFN-gamma, and TNF-alpha (but not IL-4) when activated by calcium ionophore + PMA and IL-1. We then separated CD3-4-8- thymocytes into IL-2R+ and IL-2R-. CD3-4-8-IL-2R+ thymocytes only produced small amounts of IL-2 when activated with calcium ionophore + PMA + IL-1, whereas CD3-4-8-IL-2R- thymocytes did not require IL-1 to produce IL-2, IFN-gamma, and TNF-alpha. Finally, CD4-8+3- thymocytes (an immature population believed to be an intermediate between CD3-4-8- and CD4+8+ thymocytes) only produced marginally detectable levels of IL-2 upon stimulation with calcium ionophore, PMA, and the addition of IL-1 did not result in increased levels of cytokine production. These observations indicate discrete patterns of cytokine production by the subsets studied and suggest specific controls of cytokine gene expression during T cell development.     

Inhibition of interleukin 1 synthesis by tenidap: a new drug for arthritis

Tenidap is a new antiarthritic drug of novel chemical structure. This study shows the effects of tenidap on the in vitro synthesis of interleukin 1 (IL-1). IL-1 production by murine peritoneal macrophages was induced either by stimulation with lipopolysaccharide (LPS) or by phagocytosis of zymosan. With either stimulus, tenidap inhibited IL-1 production as measured by a quantitative competitive IL-1 receptor binding assay. Approximately 20 ng/mL of IL-1 was produced by 10(6) macrophages in response to LPS and about half that amount was produced in response to zymosan. Fifty percent inhibition of IL-1 production by tenidap was found at 3 microM for both stimuli. Using goat anti-IL-1 alpha and Western blot analysis, the appearance of intracellular 34 kDa pro-IL-1 alpha was inhibited by tenidap down to 3 microM. Tenidap decreased [35S]Met incorporation into cellular protein at 30 microM but not at 10 or 3 microM, indicating selectivity for IL-1 inhibition relative to total protein synthesis. Because tenidap inhibited IL-1 induction by both zymosan and LPS, it must act subsequently to receptor triggering. As the appearance of IL-1 was inhibited both intracellularly and extracellularly, the primary drug effect cannot be on secretion.     

Calcium mediates one of the signals required for interleukin 1 and 2 production by murine cell lines

The murine T lymphoma cell line EL-4 can be induced to produce interleukin 2 (IL-2) by concurrent stimulation with interleukin 1 (IL-1) and a T-cell lectin such as phytohemagglutinin (PHA) or concanavalin A (Con A). The results presented here demonstrate that the requirement for the lectin, but not IL-1, could be completely replaced by the calcium ionophore A23187. The optimal effective concentration of A23187 was found to be 2.5 X 10(-7) M, and the costimulating effect of IL-1 was dose-dependent. The stimulatory effect of A23187 was completely eliminated by incorporation of 5 mM ethylene glycol bis (beta-aminoethyl ether) N,N,N',N'-tetracetic acid (EGTA) in the culture medium, and this inhibition could in turn be reversed by addition of 5 mM CaCl2 to the medium. Release of IL-2 from IL-1/A23187-stimulated EL-4 was detected within 5 hr after initiation of the cultures, and both signals were required at the same time to initiate synthesis or release of IL-2. In addition, the calcium ionophore also augmented release of IL-1 from the P388D1 murine macrophage cell line. These results suggest that a calcium-mediated event may serve as a common mechanism for the induction of secretion of lymphokines and monokines from murine cell lines.     

IFN-gamma stimulates IgG2a secretion by murine B cells stimulated with bacterial lipopolysaccharide

rIFN-gamma strikingly enhances the secretion of IgG2a by murine splenic B cells stimulated with bacterial LPS in vitro and concomitantly suppresses the production of IgG3, IgG1, IgG2b, and IgE while sparing IgM secretion. IFN-gamma stimulates highly purified B cell populations to secrete IgG2a, strongly suggesting that it acts directly on B cells. It increases the frequency of precursors of IgG2a-expressing soft agar colonies and enhances the number of IgG2a+ cells in colonies indicating that it both increases the frequency of precursors of IgG2a+ cells and enhances the number of IgG2a+ daughter cells emerging from each precursor. IFN-gamma completes its action within the first 24 to 48 h of a 6-day culture with LPS and its addition cannot be delayed beyond the first 48 h. Preincubation of resting B cells in the presence of IFN-gamma leads to a time dependent increase, up to 42 h, in IgG2a secretion upon subsequent addition of LPS. IFN-gamma can exert this action on resting B cells that have been selected for absence of membrane IgG expression by cell sorting. The promotion of IgG2a secretion appears to be a specific property of IFN-gamma in that IFN-alpha, IFN-beta, IL-1, IL-2, IL-3, IL-4, IL-5, granulocyte-macrophage-CSF, granulocyte-CSF, and CSF-1 fail to enhance IgG2a secretion by LPS-stimulated B cells.