Cytokine gene expression in atopics: effect of IL-4 on IL-1 beta and IL-6 mRNA levels

Mononuclear cells from atopic blood donors showed increased IL-3 steady state mRNA levels. This finding complemented our earlier observations that cells from atopics also showed increased IL-4 but decreased IFN-gamma, IL-1 beta and IL-6 mRNA levels. Therefore, we investigated the effect of human recombinant IL-4 on cytokines mRNA levels in mononuclear cells from normals and atopics. In the presence of IL-4 steady state levels of IL-1 beta and IL-6 mRNA were decreased even if cells were co-stimulated with polyclonal activators such as PMA, PWM or PHA. No influence of IL-4 on granulocyte-macrophage colony-stimulating factor (GM-CSF), IL-3 or IFN-gamma mRNA levels was observed with the exception of a decreased IFN-gamma mRNA level in PWM stimulated cells.     

Effect of cytokines on Japanese encephalitis virus production by human monocytes

Japanese encephalitis (JE) virus was shown to grow in in vitro cultures of human monocytes. Interferon (IFN)-alpha and IFN-gamma inhibited JE virus production by the infected monocytes in the absence of anti-JE virus antibody, but interleukin (IL)-1 alpha, IL-2, IL-3, granulocyte-macrophage colony-stimulating factor (GM-CSF), granulocyte-CSF (G-CSF), and tumor necrosis factor (TNF)-alpha did not show a significant inhibition. Antibody against JE virus increased the JE virus production by the infected monocytes probably by enhanced uptake of virus-antibody complexes via Fc receptors. IFN-gamma and GM-CSF increased JE virus production by monocytes in the presence of anti-JE virus antibody, whereas IFN-alpha inhibited JE virus production even in the presence of the antibody. The other 5 cytokines (IL-1 alpha, IL-2, IL-3, G-CSF, and TNF-alpha) did not show a significant effect on JE virus production by monocytes in the presence or absence of the antibody.     

Expression of colony-stimulating factors and inflammatory cytokines in the uterus of CD1 mice during days 1 to 3 of pregnancy.

Morphological and immunohistochemical analyses have documented the development of an acute inflammatory response, marked by the early appearance of granulocytes and later infiltration of mononuclear cells, in the uterus immediately after mating in mice. The response peaked on Day 1 and subsided by Day 3. In the present study, RNAs for macrophage colony-stimulating factor (CSF-1) and granulocyte-macrophage colony-stimulating factor (GM-CSF) and for interleukin 1 alpha (IL-1 alpha), IL-1 beta, interleukin 6 (IL-6) and tumour necrosis factor alpha (TNF-alpha) were detected in uterine tissue on Day 1. With the exception of IL-6, which was higher on Day 3 than on Day 1, and IL-1 alpha, which was not reduced on Day 2, concentrations of cytokine mRNA decreased to Day 3. No bioactivity was detected for GM-CSF, granulocyte colony-stimulating factor or IL-3, but CSF-1, IL-1, IL-6 and TNF-alpha were detected on Day 1 using bioassays. Changes in concentrations approximately paralleled those for mRNA. The concentrations of mRNA for CSF-1, IL-1, IL-6 and TNF-alpha were higher on Day 1 of pregnancy than in the uteri of cycling mice 24 h earlier. The data are consistent with previous morphological observations demonstrating the expression of an acute inflammatory response in the mouse uterus after mating. Further, the data demonstrate the expression of genes for CSF-1, GM-CSF, IL-1 alpha, IL-1 beta, IL-6 and TNF-alpha is induced in the uterus during mating.     

Modulation of c-kit mRNA and protein by hemopoietic growth factors.

We examined the effects of various hemopoietins on c-kit mRNA and protein expression. Interleukin-3 (IL-3), granulocyte-macrophage colony-stimulating factor, and erythropoietin, but not IL-4, down-regulated levels of c-kit mRNA expressed by mast cells and stem cell progenitors. The effect of IL-3 was dominant and independent of cell growth or viability and was paralleled by reduced expression in c-kit protein. These observations indicate that regulation of c-kit expression is closely interlinked with the molecular mechanisms triggered by erythropoietin, IL-3, and granulocyte-macrophage colony-stimulating factor.     

Cytokine responses of intestinal epithelial-like Caco-2 cells to non-pathogenic and opportunistic pathogenic yeasts in the presence of butyric acid

Candida albicans, Saccharomyces cerevisiae and their cell wall components, zymosan and glucan, have been shown to stimulate interleukin-8 (IL-8/CXCL-8) production by intestinal epithelial cell-like Caco-2 cells pre-cultured with 10 mM butyric acid. We examined in this study whether these yeasts also altered the production of other cytokines and cyclooxygenases (COXs) by Caco-2 cells. Culturing Caco-2 cells with 10 mM butyric acid and 15% FBS for 4 days enhanced the basal levels of mRNA encoding IL-6, IL-8, IL-18, monocyte chemoattractant protein (MCP)-1, stem cell factor, transforming growth factor (TGF)-beta1, TGF-beta3, tumor necrosis factor (TNF)-alpha, COX-1, and COX-2, but not of granulocyte-macrophage colony-stimulating factor (GM-CSF) and TGF-beta2. The inclusion of live S. cerevisiae or C. albicans further enhanced the production of IL-8, but not of the other cytokines and COXs. The non-pathogenic yeasts, C. kefyr, C. utilis, C. versatilis, Kluyveromyces lactis, K. marxianus, Schizosaccharomyces pombe and Zygosaccharomyces rouxii, used for the production of fermented foods and probiotics, and the opportunistic pathogens, C. glabrata, C. krusei, C. parapsilosis and C. tropicalis, isolated from human tissue samples also enhanced IL-8 secretion by Caco-2 cells.     

Characterization and expression kinetics of an endothelial cell activation antigen present in vivo only in acute inflammatory tissues

Endothelial cell activation by endotoxin (LPS), tumor necrosis factor (TNF), Interleukin-1-alpha, beta (IL-1-alpha, beta) and phorbolesters (TPA) results in increased monocyte adhesion. Examination of kinetics of monocyte adhesion shows that the onset of adherence enhancement (AE) is similar in all five agents (about 300% AE at 6 h), while its decrease is delayed in LPS/TNF versus IL-1-alpha, beta/TPA-induced activation (LPS versus IL-1-beta:260% versus 60% at 18 h). Monoclonal antibody (4D10), raised against 24 h LPS-stimulated endothelial cells detects an endothelial cell-specific activation antigen at Mr 81,000 that is induced by LPS, TNF, IL-1-alpha, beta and TPA (within 6 h about 100% positive cells). Decrease in antigen-positive cells is delayed in LPS/TNF versus IL-1-alpha, beta/TPA-induced antigen expression (LPS vs. IL-1-beta: 60% vs. 5% at 24 h). In situ the antigen is not expressed in normal and chronic inflammatory tissues. Acute inflammatory tissues, including contact and atopic dermatitis, psoriasis and periodontitis, however, show endothelial cells staining strongly positive. In contact eczemas at different times after elicitation (0, 6, 24, 72, 96 h), expression of the antigen is first seen after 24 h and is still strong at 96 h. These data indicate that LPS/TNF conduct an endothelial cell activation program in vitro, showing the same prolonged kinetics that is found for endothelial cell activation in the acute inflammatory process in vivo.     

Lithium chloride potentiates tumor necrosis factor-induced and interleukin 1-induced cytokine and cytokine receptor expression.

The antimalignant cell activity of tumor necrosis factor (TNF) in many cell types can be enhanced by lithium chloride (LiCl). This study shows the in vitro effect of LiCl on the TNF-induced or interleukin 1 (IL-1)-induced expression of IL-6, granulocyte-macrophage colony-stimulating factor (GM-CSF), IL-3, IL-2, and the IL-2 receptor-alpha (IL-2R alpha). The levels of IL-6 and GM-CSF in the medium of TNF-treated L929 fibrosarcoma cells were increased by cotreatment with LiCl. In contrast, enhancement of IL-6 production by dibutyryl cyclic AMP or cycloheximide was not affected by LiCl. The production of IL-6 and GM-CSF was not correlated with sensitivity to TNF-mediated cell killing. IL-1 by itself had no measurable effects on L929 cells. However, LiCl potentiated the IL-1-induced synthesis of IL-6, GM-CSF, IL-3, and IL-2 in PC60 murine T-cell hybridoma cells. TNF alone induced only GM-CSF production in these cells, but in the presence of LiCl, increased amounts of GM-CSF as well as small amounts of IL-2 and IL-6 could be detected. It is also shown that in these PC60 cells the expression of the IL-2R alpha was induced by TNF + LiCl treatment but not by TNF alone. IL-2R alpha expression was likewise considerably enhanced by IL-1 + LiCl treatment, as compared with treatment with IL-1 alone. The effects of LiCl on the TNF-induced and the IL-1-induced gene expression seem to be independent of the protein kinase A and C pathways. These results show that LiCl can modulate both TNF-mediated cytotoxicity and TNF-induced and IL-1-induced cytokine expression, suggesting that Li+ acts early in the TNF-signaling pathway, but at a step shared with the IL-1-signaling pathway.     

Granulocyte-macrophage colony stimulating factor induces both HLA-DR expression and cytokine production by human monocytes

The effect of recombinant human granulocyte-macrophage colony stimulating factor (GM-CSF) on the expression of HLA-DR, and the production of the cytokines interleukin-1 (IL-1) and tumor necrosis factor alpha (TNF alpha) by human peripheral blood monocyte-enriched populations was investigated. GM-CSF was shown to induce both the expression of HLA-DR and the cytokines IL-1 and TNF alpha in a dose-dependent manner. In contrast, interferon-gamma (IFN-gamma), which induced major histocompatibility complex (MHC) class II expression, did not induce IL-1 or TNF alpha production. However, IFN-gamma enhanced the cell surface expression of HLA-DR and the production of IL-1 and TNF alpha on monocyte-enriched cells stimulated by GM-CSF. By itself, GM-CSF did not induce surface class II expression on the human monocytic tumour cell line THP-1, whereas it synergized with IFN-gamma to induce surface expression. These cells responded to GM-CSF by producing IL-1 and TNF alpha; Northern blotting showed that mRNA levels of IL-1 and TNF alpha were transiently induced, similar to other cytokines. Our results indicate that GM-CSF is a major macrophage activating factor that is capable of inducing both the expression of HLA-DR and the cytokines involved in T-cell activation by macrophages; therefore, GM-CSF may be of importance in potentiating antigen presenting function.     

Interleukin 1 induces interleukin 1. I. Induction of circulating interleukin 1 in rabbits in vivo and in human mononuclear cells in vitro

Interleukin 1 (IL-1) plays an important role in host defense mechanisms by increasing body temperature, inducing the synthesis of a variety of lymphokines and hepatic acute phase proteins and acting as a chemoattractant for lymphocytes. However, in some microenvironments such as injured tissue or joint spaces, elevated IL-1 levels may contribute to pathologic processes, for example, proliferation and fibrosis of tissue involved in pannus formation as well as degradation of matrix and abnormal tissue architecture. To investigate potential mechanisms that may lead to excessive production of IL-1, we have examined the ability of IL-1 to participate in an amplification event by inducing its own gene expression leading to synthesis of biologically active IL-1. When injected into rabbits, recombinant human IL-1-alpha induced biphasic fevers, and during the second temperature elevation 3 hr later, a circulating pyrogenic material was detected by passive transfer of plasma to other rabbits. Induction of the biphasic fever was not caused by endotoxin contamination of the recombinant IL-1. The 3-hr circulating pyrogen was heat-labile and was not residual injected IL-1-alpha. Chromatographic separation of this plasma and biologic assay suggested that it was new IL-1 of rabbit origin. We next incubated human blood mononuclear cells with recombinant IL-1-alpha and measured the intracellular and extracellular levels of IL-1 by bioassay using the D10.G4.1 murine T cell line. In order to control for the carryover of recombinant IL-1-alpha used to stimulate the mononuclear cells (MNC), we used neutralizing antibodies that were specific for IL-1-alpha or IL-1-beta. The results of these neutralizations showed that recombinant human IL-1-alpha induces the synthesis of IL-1-beta in human MNC in vitro. These results were verified with a radioimmunoassay specific for IL-1-beta. At concentrations of 100 ng/ml, IL-1-alpha induced prostaglandin E2 production in the MNC culture, and this was associated with decreased production of immunoreactive IL-1-beta. Adding indomethacin to the cultures prevented the decreased production of IL-1-beta induced by high concentrations of IL-1-alpha. Using nonadherent MNC, we observed an increase in IL-1-beta as well as IL-1-alpha mRNA after 4 hr of exposure to recombinant IL-1-alpha. These results demonstrate that IL-1-alpha induces biologically active and immunoreactive IL-1-beta from MNC in vitro and that the same concentrations of IL-1-alpha induce gene expression for both forms of IL-1.(ABSTRACT TRUNCATED AT 400 WORDS)     

Abelson virus transformation of an interleukin 2-dependent antigen-specific T-cell line.

Abelson murine leukemia virus (A-MuLV) carries the gene v-abl, one of a group of oncogenes with structural and functional (tyrosine kinase) homology to three growth factor receptors. Work in this and other laboratories has shown that A-MuLV infection can render myeloid and lymphoid cells independent of the growth factors interleukin 3 and granulocyte-macrophage colony-stimulating factor. We have now shown that v-abl can also relieve interleukin 2 (IL-2) dependence in T cells. We infected a cloned IL-2-dependent antigen-specific cell line. Transformed cells were generated which were factor independent and tumorigenic. The transformants each bore unique v-abl DNA inserts and expressed v-abl mRNA. No elevation of expression of either IL-2 or its receptor could be detected in these cells. Thus, A-MuLV can short-circuit the dependence of hematopoietic cells on IL-2, IL-3, and possibly granulocyte-macrophage colony-stimulating factor, none of whose receptors are known to be of the tyrosine kinase type.     

The network of hemopoietic regulatory proteins in myeloid cell differentiation.

There are clones of myeloid leukemic cells that can be induced to undergo terminal cell differentiation to macrophages by normal hemopoietic regulatory proteins. Induction of differentiation in two different clones of myeloid leukemic cells with interleukin 6 (IL-6) or granulocyte-macrophage colony-stimulating factor (GM-CSF) resulted in induction of mRNA for the hemopoietic regulatory proteins IL-6, GM-CSF, interleukin 1 alpha and interleukin 1 beta, tumor necrosis factor, and transforming growth factor beta 1. In one of these clones, induction of differentiation with GM-CSF was also associated with induction of mRNA for macrophage colony-stimulating factor (M-CSF) but not for the receptor for M-CSF (c-fms), whereas in the other clone, induction of differentiation with IL-6 was associated with induction of mRNA for both c-fms and M-CSF. The clones also differed in their responsiveness to these regulators. There was no induction of mRNA for granulocyte colony-stimulating factor or interleukin 3 during differentiation of either clone. The results indicate that the genes for a nearly normal network of positive and negative hemopoietic regulatory proteins are induced during differentiation of these myeloid leukemic cells and that there are leukemic clones with specific defects in this network.     

不同诱导因子对人外周血单个核细胞P2X7受体表达的作用

ATP激活P2X7受体可产生一系列的白细胞功能反应,因此P2X7受体的表达调控引起我们的兴趣。然而P2X7受体在正常人外周血单个核细胞(peripheral blood mononuclear cells,PBMC)、单核细胞中的表达调控机制尚未阐明。本文用半定量RT-PCR方法检测多种细胞因子、细菌抗原、丝裂原对P2X7受体表达的诱导作用,探索P2X7受体的诱导表达模式。结果表明,单个核细胞和单核细胞可检出P2X7受体的表达;白细胞介素2、4、6(interleukin-2、-4、-6,IL-2、IL-4、IL-6)、肿瘤坏死因子仪(tumour necrosis factor-α,TNF-α)等细胞因子和金黄色葡萄球菌CowanⅠ株(Staphylococcus aureus Cowan strainⅠ,SAC)、脂多糖(lipopolysaccharide,LPS)能上调PBMC的P2X7受体表达,而γ干扰素(interferon-γ,IFN-γ)、粒-巨噬细胞集落刺激因子(granulocyte-macrophage colony-stimulating factor,GM-CSF)、巨噬细胞集落刺激因子(macmphage colony-stimulating factor,M-CSF)和植物血凝素(phytohemagglutinin-M,PHA-M)等则没有作用;LPS和M-CSF可以提高单核细胞的P2X7受体表达,IFN-γ、TNF-α、GM-CSF作用较弱,但是这些因子的预处理并不能增强LPS对P2X7受体表达的诱导。炎症因子促进P2X7受体的表达,提示P2X7受体可能在对抗细菌感染的免疫反应中起一定作用,这有待于进一步研究。     

Soluble interleukin-6 receptor alpha inhibits the cytokine-Induced fractalkine/CX3CL1 expression in human vascular endothelial cells in culture

Soluble form of IL-6 receptor alpha (sIL-6R) is known to serve as an agonist, without exogenous IL-6, on endothelial cells which do not express IL-6R but have only IL-6 receptor beta chain, gp130. We investigated the effect of sIL-6R on fractalkine expression in human umbilical vein endothelial cells (HUVECs) in culture. sIL-6R markedly inhibited HUVEC fractalkine/CX3CL1 expression induced by interleukin (IL)-1alpha, tumor necrosis factor (TNF)-alpha, or interferon (IFN)-gamma. IL-1alpha-induced fractalkine expression was inhibited by sIL-6R in time- and concentration-dependent manners. The experiment using actinomycin D indicated that sIL-6R lowered the stability of fractalkine mRNA. The inhibitory effect of sIL-6R was reversed by anti-gp130 neutralizing antibody. sIL-6R inhibited adhesion of mononuclear cells (MNCs) to HUVEC monolayers stimulated with IFN-gamma, but it did not inhibit the adhesion to monolayers stimulated with IL-1alpha. MNC chemotactic activity of conditioned medium of HUVEC stimulated with IL-1alpha or IFN-gamma was inhibited by co-treatment with sIL-6R. sIL-6R may play a regulatory role in immune responses by modulating the interaction between leukocytes and the vascular endothelium.