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Query: UNIPROT:P05231 (interleukin-6)
 23,907 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

cDNAs coding for the human hepatic interleukin-6 receptor (IL-6-R) have been isolated from a library made from poly(A) RNA of dexamethasone-treated human hepatoma cells (HepG2). We found the hepatic IL-6-R to be identical to the one expressed by leucocytes. A polyclonal antiserum was raised in rabbits against the IL-6-R protein expressed in Escherichia coli. Although the entire IL-6-R protein was used for immunization, only antibodies to the cytoplasmic domain of the IL-6-R were obtained. It is demonstrated by affinity cross-linking and subsequent immunoprecipitation with antibodies against the ligand as well as against the receptor that the cloned cDNA codes for the functional IL-6-R on HepG2 cells. When the hepatic IL-6-R cDNA was overexpressed in HepG2 cells, these cells became more sensitive to low concentrations of IL-6 with respect to the induction of gamma-fibrinogen mRNA.
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PMID:Structural and functional studies on the human hepatic interleukin-6 receptor. Molecular cloning and overexpression in HepG2 cells. 187 1

Internal deletions of the human interleukin-6 (IL-6) cDNA have been generated in the region encoding residues 29 to 42. Mutant proteins were produced by in vitro transcription-translation or in Escherichia coli and tested for their biological activity using the hybridoma growth factor (HGF) assay or a transcriptional activation assay on human hepatoma cells. The folding of the mutants was also checked by immunoprecipitation with conformation-specific monoclonal antibodies. The results show that only residues 29 to 34 are crucial for IL-6 activity and that the first two amino acids are probably involved in the definition of the IL-6 active site.
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PMID:Internal deletions of amino acids 29-42 of human interleukin-6 (IL-6) differentially affect bioactivity and folding. 187 51

Conditioned medium from human monocyte-macrophages incubated under various conditions was tested for its ability to stimulate fibrinogen mRNA levels in the hepatoma cell line HepG2. Recombinant human interleukin-6 (IL-6) stimulated fibrinogen mRNA levels 4.4-fold over control levels; this response was blocked by an anti-IL-6 antibody. Conditioned medium from 3-day-cultured monocyte-macrophages produced a slight stimulation of fibrinogen synthesis in HepG2 cells which was enhanced when the monocyte-macrophages had been treated with lipopolysaccharide (LPS). This stimulation was blocked by the anti IL-6 antibody. The cytokines, interleukin-1 (IL-1) and tumour necrosis factor (TNF) were also detected in the conditioned medium from the 3-day-cultured monocyte-macrophages. Monocyte-macrophages were cultured for 17 days and then incubated with acetylated low density lipoprotein (AcLDL) for 48 h. Such cells were 'foamy' in appearance and showed a 4-fold increase in apoE mRNA and a 10 to 50-fold increase in apoE secretion. This increase in apoE production was suppressed by almost a third when cells were coincubated with AcLDL and LPS. Conditioned medium from these 17-day-cultured AcLDL-treated human monocyte-macrophages did not stimulate fibrinogen mRNA synthesis in HepG2 cells, nor did the conditioned medium contain detectable levels of cytokines. These results suggest that cytokine production from foam cells in the atherosclerotic lesion is unlikely to be a major contributing factor in determining the elevated fibrinogen levels seen in the plasma of patients with IHD.
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PMID:Cytokine production by cholesterol-loaded human peripheral monocyte-macrophages: the effect on fibrinogen mRNA levels in a hepatoma cell-line (HepG2). 193 38

The current study reinvestigated whether pure plasminolytic fragments D and E from fibrin directly increase fibrinogen synthesized by the hepatocyte. Fibrinogen protein levels and fibrinogen mRNA levels in an interleukin-6 (IL-6) responsive rat hepatoma cell line (FAZA) were measured quantitatively by [35S]-methionine pulse-chase experiments and Northern hybridizations, respectively. The results demonstrate that neither fragments D nor E, alone or in combination, in the presence or absence of dexamethasone, had any influence on the production of fibrinogen.
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PMID:The putative role of fibrin fragments in the biosynthesis of fibrinogen by hepatoma cells. 201 4

Human interleukin-6 or B-cell stimulatory factor-2 is a cytokine involved in acute phase and immune response. Cloning of cDNA for human interleukin-6 in the pT7.7 expression plasmid under the control of a bacteriophage T7 RNA polymerase promoter system allows rapid production of the cytokine in Escherichia coli. Upon cell induction with isopropyl thiogalactopyranoside, recombinant human interleukin-6 is overexpressed and forms insoluble inclusion bodies. Solubilization of the protein with 6 M guanidine hydrochloride and refolding in the presence of a reduction/oxidation system results in a quantitative recovery of recombinant human interleukin-6. This material is already 70% pure and can be further purified to homogeneity with a single passage over a weak anionic-exchange column. Extended structural characterization of the purified protein by electrospray mass spectrometry, automated Edman degradation and peptide mapping by high-pressure liquid chromatography/fast-atom-bombardment mass spectrometry demonstrates that recombinant human interleukin-6 is identical to the natural protein both in amino acid sequence and S-S bridge content. However, it contains a minor component accounting for about 20% of the entire translated protein which exhibits a Met-Ala dipeptide extension at the N-terminus. Purified recombinant human interleukin-6 is biologically active because it is able to induce at least 70-fold the human C-reactive promoter transfected in human hepatoma Hep 3B cells and is stable for several months in 10% glycerol at 4 degrees C. The expression system described in the present work has the main advantage of producing a high yield of recombinant human interleukin-6 (about 25 mg/l) combined with a very simple purification scheme.
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PMID:Single-step purification and structural characterization of human interleukin-6 produced in Escherichia coli from a T7 RNA polymerase expression vector. 205 Jan 35

Transcription of the human C-reactive protein (CRP) gene is induced by interleukin-6 (IL-6) during acute inflammation. Important information for inducible CRP expression is located within the 90 bases preceding the transcriptional start site. We show that the CRP promoter contains two adjacent binding sites (beta and alpha) that interact with at least two hepatocyte-specific nuclear proteins, H-APF-1 and H-APF-2. Point mutations that abolish or reduce binding drastically affect the level of CRP gene expression. Binding to beta is identical when extracts from uninduced or IL-6-induced Hep3B cells are used. On the contrary, both quantitative and qualitative changes in the alpha binding can be detected with extracts from uninduced cells or from cells treated with IL-6 or IL-6 + cycloheximide. A synthetic promoter based on the multimerization of the beta-binding domain, but not of the alpha-domain, is highly inducible when transfected in hepatoma cells. These results are discussed in relation to the structure of the promoter region of other acute phase inducible genes.
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PMID:Constitutive and IL-6-induced nuclear factors that interact with the human C-reactive protein promoter. 215 72

To study the mechanism of induction of human C-reactive protein (CRP) gene expression, we have utilized an in vitro liver cell system to analyze the cis-acting DNA sequences located within the 5'-flanking region of human CRP gene. Stable transfection of human hepatoma cells, PLC/PRF/5, by a CRP gene construct containing the 1 kilobase pair of upstream sequence of the CRP gene demonstrated that this region contained the inducible element(s) which regulated human CRP gene transcription. Dissection of this region by 5', 3' and internal deletion constructs of upstream region of the CRP gene fused to a reporter gene, chloramphenicol acetyl transferase, indicated the presence of two inducible elements located proximal to the site of initiation of transcription, two constitutive enhancer-like elements located distal to the promoter, and a negative regulatory region located between the two inducible elements. We had previously shown that a protein factor from monocytes or HTLV1-infected T-cells, was responsible for CRP induction in hepatoma cells. We have found this factor to be synonymous with interleukin-6. By stable and transient transfection assays in hepatoma cells, recombinant interleukin-6 alone was sufficient to activate both inducible elements.
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PMID:cis-acting elements responsible for interleukin-6 inducible C-reactive protein gene expression. 215 96

IL6-PE40 is a chimeric toxin composed of human interleukin-6 (IL6) linked by a peptide bond to PE40, a form of Pseudomonas exotoxin (PE) devoid of its cell recognition domain. To identify cancer cell lines with high numbers of IL6 receptors and to assess the usefulness of IL6-PE40 as a possible anticancer agent, we evaluated the toxicity of IL6-PE40 on a variety of tumor cell lines and demonstrated that certain human myeloma and hepatoma cell lines were particularly sensitive. IL6 binding to selected hepatoma and myeloma cell lines were determined by using [125I]IL6. IL6 receptor mRNA levels were measured by polymerase chain reactions. When comparisons were made among different hepatoma cell lines, the sensitivity to IL6-PE40 correlated with the number of IL6 receptors. However, the hepatoma line PLC/PRF/5, which contains 2,300 IL6 receptors, was more sensitive to IL6-PE40 (amount of protein required to inhibit protein synthesis by 50% was 5 ng/ml) than both the myeloma cell lines U266 and H929 (for both cell lines, the 50% inhibitory dose was 8 ng/ml), which contain 15,500 and 16,500 IL6 receptors, respectively. RNA analysis confirmed that the sensitivity of these cells to IL6-PE40 and the amount of IL6 receptor RNA detected did not correlate. These data suggest that factors in addition to the number of IL6-binding sites contribute to the sensitivity of cells to IL6-PE40.
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PMID:Cell-specific toxicity of a chimeric protein composed of interleukin-6 and Pseudomonas exotoxin (IL6-PE40) on tumor cells. 216 May 79

Affinity cross-linking of 125I-labeled recombinant human interleukin-6 (IL-6) to human hepatoma cells (HepG2) allowed the detection of three IL-6-containing complexes with molecular masses of 100 kDa, 120 kDa and 200 kDa. Treatment of HepG2 cells with dexamethasone led to a time- and dose-dependent up-regulation of IL-6-receptor mRNA levels. By the use of cross-linking this effect was also seen at the protein level, where all three IL-6-binding complexes increased upon incubation of HepG2 cells with dexamethasone. Under conditions of IL-6-receptor up-regulation by dexamethasone, gamma-fibrinogen mRNA induction by IL-6 is stronger and occurs earlier than without dexamethasone. We propose therefore that the expression of the IL-6 receptor might be a rate-limiting step in acute-phase-protein induction.
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PMID:Studies on the structure and regulation of the human hepatic interleukin-6 receptor. 216 35

Human C-reactive protein (CRP) is the major acute phase reactant during inflammation. Regulation of CRP gene expression has been studied in two experimental systems: transgenic mice and human hepatoma cells. In the first system the human CRP gene flanked by approximately 10(4) bases of 5' and 3' sequences is expressed in a liver-specific and inducible manner. The chromatin configuration of the CRP transgene is characterized by the presence of constitutive and inducible liver-specific DNase I-hypersensitive sites. Inducible sites map precisely at the level of the CRP promoter region. In hepatoma cells we analysed the expression of the bacterial chloramphenicol acetyltransferase (CAT) gene driven by various segments of the CRP promoter. This latter approach has led to the identification of promoter elements responsive to interleukin-6 and of hepatocyte-specific nuclear proteins that interact with them.
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PMID:Regulation of the human C-reactive protein gene, a major marker of inflammation and cancer. 217 Aug 8


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