Biosynthesis Of Interleukin-6 Research Articles

Acid β-Glucosidase 1 Counteracts p38δ-dependent Induction of Interleukin-6: POSSIBLE ROLE FOR CERAMIDE AS AN ANTI-INFLAMMATORY LIPID

Activation of protein kinase C (PKC) by the phorbol ester (phorbol 12-myristate 13-acetate) induces ceramide formation through the salvage pathway involving, in part, acid beta-glucosidase 1 (GBA1), which cleaves glucosylceramide to ceramide. Here, we examine the role of the GBA1-ceramide pathway, in regulating a pro-inflammatory pathway initiated by PKC and leading to activation of p38 and induction of interleukin 6 (IL-6). Inhibition of ceramide formation by fumonisin B1 or down-regulation of PKCdelta potentiated PMA-induced activation of p38 in human breast cancer MCF-7 cells. Similarly, knockdown of GBA1 by small interfering RNAs or pharmacological inhibition of GBA1 promoted further activation of p38 after PMA treatment, implicating the GBA1-ceramide pathway in the termination of p38 activation. Knockdown of GBA1 also evoked the hyperproduction of IL-6 in response to 4beta phorbol 12-myristate 13-acetate. On the other hand, increasing cellular ceramide with cell-permeable ceramide treatment resulted in attenuation of the IL-6 response. Importantly, silencing the delta isoform of the p38 family significantly attenuated the hyperproduction of IL-6. Reciprocally, p38delta overexpression induced IL-6 biosynthesis. Thus, the GBA1-ceramide pathway is suggested to play an important role in terminating p38delta activation responsible for IL-6 biosynthesis. Furthermore, the p38delta isoform was identified as a novel and predominant target of ceramide signaling as well as a regulator of IL-6 biosynthesis.

P38α Stabilizes Interleukin-6 mRNA via Multiple AU-richElements

AU-rich elements (AREs) in the 3'-untranslated region (UTR) of unstable mRNA dictate their degradation or mediate translational repression. Cell signaling through p38alpha MAPK is necessary for post-transcriptional regulation of many pro-inflammatory cytokines. Here, the cis-acting elements of interleukin-6 (IL-6) 3'-UTR mRNA that required p38alpha signaling for mRNA stability and translation were identified. Using mouse embryonic fibroblasts (MEFs) derived from p38alpha(+/+) and p38alpha(-/-) mice, we observed that p38alpha is obligatory for the IL-1-induced IL-6 biosynthesis. IL-6 mRNA stability is promoted by p38alpha via 3'-UTR. To understand the mechanism of cis-elements regulated by p38alpha at post-transcriptional level, truncation of 3'-UTR and the full-length 3'-UTR with individual AUUUA motif mutation placed in gene reporter system was employed. Mutation-based screen performed in p38alpha(+/+) and p38alpha(-/-) mouse embryonic fibroblast cells revealed that ARE1, ARE2, and ARE5 in IL-6 3'-UTR were targeted by p38alpha, and truncation-based screen showed that IL-6 3'-UTR-(56-173) was targeted by p38alpha to stable mRNA. RNA secondary structure analysis indicated that modulated reporter gene expression was consistent with predicted secondary structure changes.

Thromboxane A2Promotes Interleukin-6 Biosynthesis Mediated by an Activation of Cyclic AMP-Response Element-Binding Protein in 1321N1 Human Astrocytoma Cells

1321N1 human astrocytoma cells express thromboxane A2 (TXA2) receptors (TP). However, physiological consequences of TXA2 signaling in glial cells remain unclear. Herein, we show that TXA2 promotes interleukin-6 (IL-6) biosynthesis in glial cells. A TP agonist, 9,11-dideoxy-9alpha,11alpha-methanoepoxy-prosta-5Z,13E-dien-1-oic acid (U46619), enhanced IL-6 production in both 1321N1 cells and cultured mouse astrocytes. It has been shown that IL-6 gene expression is regulated by various transcription factors. Among them, we found a significant increase in cyclic AMP-response element-binding protein (CREB) activity with its phosphorylation at Ser133 by U46619 in 1321N1 cells. Although U46619 increased IL-6 promoter activity, a mutation at cyclic AMP-response element (CRE) on the promoter clearly suppressed the effect, suggesting that CRE is involved in U46619-induced IL-6 expression. Furthermore, both CREB and IL-6 promoter activities were suppressed by SB203580 [4-(4-fluorophenyl)-2-(4-methylsulfinylphenyl)-5-(4-pyridyl)-1H-imidazole], a p38 mitogen-activated protein kinase (MAPK) inhibitor, and H89 [N-[2-(4-bromocinnamylamino)-ethyl]-5-isoquinoline], a protein kinase A (PKA) inhibitor, indicating involvements of p38 MAPK and PKA in CREB activation and IL-6 expression. To determine which G-proteins are implicated in the U46619-induced IL-6 synthesis, the interfering mutants of Galpha(q), Galpha12, or Galpha13 by were overexpressed in 1321N1 cells adenoviral approach. It is noteworthy that the Galpha(q) or Galpha13 mutant blocked the IL-6 production by U46619. The constitutively active mutant of Galpha(q), Galpha12, or Galpha13 enhanced IL-6 production, indicating that Galpha(q) and Galpha13 were involved in U46619-induced IL-6 production. In conclusion, TXA2 enhances the IL-6 biosynthesis via the PKA p38 MAPK/CREB pathway in 1321N1 cells. IL-6 induction depends on Galpha(q) and Galpha13 as well. This is the first report showing TP-mediated IL-6 production in glial cells.

Inverse regulation of interleukin-6 (IL-6) and IL-6 receptor in histamine deficient histidine decarboxylase-knock-out mice

Interleukin-6, a multifunctional cytokine upon binding to its receptor on hepatocytes regulates production of acute phase proteins involved in local and systemic inflammation. Gene expression and biosynthesis of IL-6 and its receptor (IL-6 R/gp130) is under complex regulation. Histamine, in addition to its principal role in immediate type hypersensitivity has been described to modulate IL-6 production and expression of IL-6 receptor. In this study, the IL-6 and IL-6 receptor expression was examined in histamine deficient histidine decarboxylase (HDC) knock-out mouse model. Our data suggest that in histamine deficient mice the inducibility of IL-6 is significantly reduced, whilst more IL-6 receptor/gp130 mRNA expresses in the liver than in wild type (HDC +/+) mice. These in vivo findings confirm earlier in vitro results and emphasize the efficacy of antihistamines in local IL-6 related processes.

Biosynthesis of interleukin-6, an autocrine growth factor for melanoma, is regulated by melanoma-derived histamine

Interleukin-6 is an autocrine growth factor in advanced stage melanoma and biosynthesis of IL-6 is increased by histamine in various cell lines. In our study we analysed the direct relation of histamine and IL-6 synthesis in human melanoma cell lines. All melanoma cells contained IL-6 mRNA, but only metastatic melanoma cells secreted the IL-6 protein. The H1 histamine receptor antagonist triprolidine decreased gene expression and biosynthesis of IL-6, while other histamine receptor antagonists had no effect. These data indicate that endogenous histamine has a definite role in the regulation of local IL-6, suggesting that histamine and IL-6 could be part of autocrine growth regulation of the tumor.

Inhibitory effect of resveratrol on interleukin 6 release by stimulated peritoneal macrophages of mice.

In the present investigation, interleukin 6 (IL-6) activity in the supernatant of cultured mouse peritoneal macrophages was monitored using a sensitive bioassay involving the IL-6-dependent murine hybridoma B9 cell line. The effects of resveratrol on Il-6 release by mouse peritoneal macrophages stimulated with calcium ionophore A23187 and fMLP were explored. Resveratrol, at a concentration range from 5 x 10(-6) to 4 x 10(-5) mol.l-1, was found to dose-dependently inhibit IL-6 release by cultured macrophages induced by A23187 and fMLP, and showed no direct cytotoxic effect, but induced proliferation of cultured mouse thymus cells. Resveratrol, at a concentration range from 10(-8) to 10(-5) mol.l-1, was shown to dose-dependently inhibit calcium ion influx into the cells with the stimulation of fMLP (10(-6) mol.l-1). These results suggest that the blocking of calcium ion influx into cells by reveratrol is one of the possible mechanisms of the IL-6 biosynthesis inhibitory action of resveratrol.

Interleukin-6 biosynthesis in human preovulatory follicles: some of its potential roles at ovulation.

In the present work we explored cellular sites of interleukin-6 (IL-6) biosynthesis in human follicular aspirates from patients undergoing in vitro fertilization therapy and the effects of this cytokine on oocyte fertilization and granulosa cell (GC) steroidogenesis. Biological IL-6 activity from 20-40 IU/mL was present in follicular fluids from 22 patients; it was also detected in 10 of 22 supernatants of cultured oocyte-cumulus complexes and in cumulus cell and GC cultures. Biological IL-6 activity in oocyte-cumulus complex cultures was not related to fertilization rates. Total ribonucleic acid was isolated from follicular aspirates and GC-enriched preparations. After reverse transcriptase and polymerase chain reaction cycles using oligonucleotide primers corresponding to known cDNA sequences for IL-6, a 126-basepair band characterized the amplification product of IL-6 transcripts on gel electrophoresis. To localize IL-6 messenger ribonucleic acid, in situ hybridization analysis was performed using a [35S]IL-6 riboprobe. The distribution of transcripts was more dense (15% vs. 3% stained cells) in GC-enriched preparations, which contained more than 95% GCs, than in original follicular preparations, which contained 20-40% viable GCs; it was not significantly modified by the presence of macrophage contaminants. The expression of IL-6 protein was assessed by positive immunohistological stainings. Biological IL-6 activity was higher, and in situ hybridization signals were more dense and more intense in 24-h GC cultures than in 72-h GC cultures, suggesting that IL-6 biosynthesis was transiently induced. Under experimental conditions of low IL-6 endogenous levels in cultures, adding recombinant human IL-6 from 10-200 IU/mL had no effect on progesterone production or aromatase activity in GC cultures free of macrophages, whereas in GC cultures including macrophage contaminants, stimulatory effects on basal and hCG-stimulated progesterone production and on basal and FSH-stimulated aromatase activities were observed. The present study provides strong support for the view that IL-6 is produced by GCs in the preovulatory follicle at the time of ovulation. In addition, we showed that IL-6 might be an intraovarian regulatory factor concerned with steroidogenesis.

Interleukin-6 biosynthesis in human preovulatory follicles: some of its potential roles at ovulation

Interleukin-6 biosynthesis in human preovulatory follicles: some of its potential roles at ovulation

Regulation of interleukin 6 gene expression in rat.

The effects of ip endotoxin administration on interleukin 6 (IL6) transcripts in brain and in peripheral tissues of rats were studied together with the effects of this treatment on IL6 and corticosterone concentrations in blood serum. Northern blot analyses showed a rapid increase of IL6 transcripts in spleen, pituitary gland, and adrenals that was paralleled by pronounced elevations in serum IL6 and corticosterone levels. Adrenalectomy further enhanced the induction of IL6 messenger RNA (mRNA) in spleen and pituitary gland and augmented the increase in serum IL6 bioactivity after lipopolysaccharide (LPS) injection. Corticosterone pretreatment (10 mg/kg) completely blocked the increase of IL6 in serum and IL6 mRNA in spleen, adrenals, and hypophysis. In several brain areas, low amounts of IL6 mRNA were detected under basal, noninflammatory conditions, but in response to LPS there was no change in the IL6 mRNA in hippocampus, hypothalamus, and cerebellum. Neither adrenalectomy nor peripheral injections of sublethal LPS doses of up to 10 mg/kg were capable of increasing IL6 mRNA in the hippocampus. The data do not support the hypothesis that central IL6 biosynthesis via transcription of the gene contributes to the endotoxin-mediated activation of the hypothalamic-pituitary-adrenal system. The results, however, clearly demonstrate that LPS-induced IL6 gene expression is subject to glucocorticoid suppression in peripheral tissues.

Biosynthesis of interleukin-6 by cultured human chorion laeve cells: regulation by cytokines.

Intrauterine infection is an important cause of preterm labor and delivery and is characterized by increased production of inflammatory cytokines by gestational tissues. We evaluated the biosynthesis of the inflammatory cytokine interleukin-6 (IL-6) by human chorion laeve cells and its regulation by other cytokines essential to the inflammatory process. We found that cultured chorion cells secrete IL-6 in the presence of growth medium supplemented only with 10% fetal calf serum. IL-1 beta, tumor necrosis factor, and lipopolysaccharide all induced a significant concentration-dependent stimulation of IL-6 production by chorion cells. The concentration range of each cytokine tested (0.1-10 ng/mL) is within the range of values found in the amniotic fluid of women destined to deliver preterm due to infection of gestational tissues. Additionally, treatment of chorion cells with IL-1 beta in combination with actinomycin-D or cycloheximide attenuated the stimulatory action of IL-1 beta on IL-6 production. Northern blot analysis of total RNA from cultured chorion cells stimulated with IL-1 beta demonstrated that IL-6 mRNA increases over time. Our data suggest that IL-6 is produced by human fetal chorion in response to infection of maternal gestational tissues. In conjunction with other inflammatory mediators, fetally derived IL-6 may play a role in the pathophysiology of preterm labor due to infection.

Biosynthesis of interleukin-6 by cultured human chorion laeve cells: regulation by cytokines

Biosynthesis of interleukin-6 by cultured human chorion laeve cells: regulation by cytokines

Inhibition by cyclo-oxygenase inhibitors of interleukin-6 production by human peripheral blood mononuclear cells

The effects of cyclo-oxygenase inhibitors on interleukin-6 (IL-6) production by human peripheral blood mononuclear cells were examined. Indomethacin and Y-9223, a novel cyclo-oxygenase inhibitor, inhibited the increases in the IL-6 level in the culture medium of both mitogen-stimulated adherent cells and non-adherent cells fractionated from mononuclear cells. Northern blotting showed that the mitogen-induced increase in the expression of IL-6 mRNA was inhibited by indomethacin and Y-9223, indicating that these agents inhibit IL-6 biosynthesis. Aspirin, ibuprofen, and phenylbutazone also inhibited IL-6 production by adherent cells stimulated with lipopolysaccharide (LPS). There was, however, no direct relationship between inhibition of IL-6 and prostaglandin E 2 (PGE 2) production by these agents. The addition of PGE 2 corresponding to the amount produced by adherent cells stimulated with LPS slightly increased IL-6 production by unstimulated adherent cells, but to a lower level than that reached with LPS. An anti-PGE 2 antibody partially blocked IL-6 production by adherent cells stimulated with LPS. These results suggest that, in addition to the inhibition of PGE 2 production, other mediators including cyclo-oxygenase products or other action mechanisms are involved in the inhibition of IL-6 production by these drugs.