Regulation of plasminogen gene expression by interleukin-6

Plasmin, the primary fibrinolytic enzyme, has a broad substrate spectrum and participates in other biological processes dependent upon proteolytic activity. Consequently, plasmin activity is tightly regulated by plasminogen activators and protease inhibitors. In this study, we examined whether regulation of plasminogen gene expression also might provide a new mechanism for controlling this system. We examined the effects of recombinant human interleukin-6 (rhIL-6), a pleiotropic cytokine, on plasminogen mRNA expression in primary murine hepatocytes and Hep3B human hepatoma cells. In primary hepatocytes, rhIL-6 and hydrocortisone separately increased plasminogen mRNA expression, but hydrocortisone did not markedly enhance the response to rhIL-6. Hep3B hepatoma cells exhibited more modest responses to rhIL-6. We used the polymerase chain reaction to amplify a 1,067-bp fragment of the human plasminogen promoter/5' flanking region. This fragment was cloned upstream of a luciferase reporter gene. Hep3B cells transiently transfected with this construct provided approximately 100-fold higher luciferase activity compared to cells transfected with control plasmids, and luciferase activity was increased approximately 4.5-fold when these cells were treated with rhIL-6. Furthermore, mice injected with rhIL-6 exhibited increases in hepatic plasminogen mRNA. Circulating plasminogen levels were significantly higher in the mice injected with rhIL-6 compared to mice injected with saline. Mice injected with lipopolysaccharide (an inducer of IL-6 in vivo) also showed increased hepatic plasminogen mRNA. Thus, plasminogen gene expression can be modulated by rhIL-6, suggesting a new mechanism for regulating biological systems that use plasmin.     

Regulation of Na+ channel beta 1 and beta 2 subunit mRNA levels in cultured rat astrocytes

This work describes the molecular mechanism of fatty acid and hormonal modulation of retinoid X receptor (RXR alpha) in rat liver. We examined the effects of different fatty acids (myristic-, stearic-, linolenic-, oleic-, arachidonic- and tetradecylthioacetic acid (TTA)) and the synthetic glucocorticoid dexamethasone on RXR alpha mRNA and protein steady-state levels in hepatoma cells and cultured hepatocytes. Fatty acids induced the RXR alpha gene expression where TTA showed the most inductive effect (three-fold induction). Dexamethasone alone resulted in a stronger induction (up to seven-fold in hepatocytes), and in combination with fatty acids, an additive or synergistic effect was observed. The RXR alpha protein level in cultured hepatocytes showed a similar pattern of regulation, with a slight inductive effect of fatty acids and an additive or synergistic effect was observed in combination with dexamethasone. Our results indicate that the RXR alpha gene expression is under distinct regulation by fatty acids and dexamethasone acid which strongly suggests a coupling with the lipid metabolizing system and the retinoid signaling pathway.     

Increased production of tumour necrosis factor-alpha interleukin-1 beta, and interleukin-6 by morphologically normal intestinal biopsies from patients with Crohn's disease

BACKGROUND: Increasing evidence points to a important role for inflammatory cytokines for the pathogenesis of Crohn's disease. AIM: To compare the secretion rate of tumour necrosis factor-alpha (TNF-alpha), interleukin-1 beta (IL-1 beta) and interleukin-6 (IL-6) by morphologically normal and inflamed intestinal mucosa from patients with Crohn's disease. RESULTS: Organ cultures of intestinal biopsy specimens taken from areas of affected mucosa from patients with Crohn's disease spontaneously produced increased amounts of TNF-alpha, IL-1 beta, and IL-6 compared with controls but also biopsy specimens taken in macroscopically and microscopically unaffected areas in the same patients. Concentrations of IL-1 beta and IL-6 measured in the supernatant fluid of biopsy cultures were positively correlated with the degree of tissue involvement measured by both endoscopic and histological grading. By contrast, TNF-alpha concentrations were not correlated to endoscopic and histological grading. CONCLUSIONS: These consistently raised TNF-alpha, IL-1 beta and IL-6 secretions by normal appearing mucosa from patients with Crohn's disease provide evidence for a sustained immune stimulation in Crohn's disease even in the absence of patent inflammation. The results shed a new light on the role of inflammatory cytokines in the onset of intestinal tissue damage in Crohn's disease and suggest that the range of intestinal lesions in Crohn's disease may be wider than suspected on the basis of regular endoscopic and histological examinations.     

Regulation of cell growth-dependent expression of mammalian CDC6 gene by the cell cycle transcription factor E2F

CDC6 of Saccharomyces cerevisiae regulates the DNA replication initiation through the origin recognition complex (ORC). Identification of a human homolog of the CDC6 gene (HsCdc6) suggests a universal role of the gene product in DNA replication. Expression of HsCdc6 is growth-regulated. We investigated the molecular basis of growth-regulated expression of mammalian Cdc6. The promoter activity of isolated HsCdc6 upstream region was activated at late G1 and G1/S boundary in the cell cycle of rat embryonic fibroblast REF52 cells by the addition of serum. The isolated promoter was activated by exogenous expression of E2F without serum stimulation. However a mutant promoter lacking the E2F recognition sites failed to respond to serum stimulation and exogenous expression of E2F. Expression of endogenous Cdc6 was induced by exogenous expression of E2F. Therefore, we concluded that the growth-regulated expression of mammalian Cdc6 was mediated by E2F. Moreover, we demonstrated that exogenous overexpression of either HsCdc6 or HsOrc1 failed to induce DNA synthesis unlike overexpression of E2F1, even though E2F1 induced both Cdc6 and Orc1, suggesting that E2F may regulate the expression of another gene(s), besides Cdc6 and Orc1, required for induction of cellular DNA synthesis in mammalian cells.     

Influence of receptor number on the stimulation by salmeterol of gene transcription in CHO-K1 cells transfected with the human beta2-adrenoceptor

We investigated the behavior of the antioxidant enzymes, superoxide dismutase (SOD), catalase (CAT), and ascorbate peroxidase (APx), in potato tubers during storage at low temperature. SOD activity increased temporarily within 3 weeks and was higher at 1 degree C than at 20 degrees C. APx activity also increased more at low (1 degree C) than at higher temperatures (5 and 20 degrees C). The contents of ascorbic acid (AsA), which is the substrate of APx, decreased immediately within 3 weeks and then gradually decreased until 15 weeks. The activity of CAT, the other enzyme which can scavenge hydrogen peroxide, decreased once in the first six weeks and thereafter increased to 15 weeks. Thus, the enhancement of the active oxygen-scavenging system that was induced by low temperature in potato tubers could result not only in a decrease of AsA but also in combined increases in APx and CAT activity whose manners were different.     

Regulation of beta 2-adrenergic receptor mRNA and gene transcription in rat C6 glioma cells: effects of agonist, forskolin, and protein synthesis inhibition

The human CDX1 gene has been isolated from a small intestine cDNA library using a murine Cdx1 cDNA probe. The nucleotide sequence of CDX1 is 81% identical to murine Cdx1 and predicts a 265-amino-acid protein with 85% identity to the mouse protein (98% identity, including conservative amino acid changes). The CDX1 locus has been mapped to a cosmid contig from chromosome 5q31-q33, placing CDX1 approximately 100 kb distal to CSFIR. Expression of CDX1 in adults appears to be limited to the intestine and colon by Northern analysis, suggesting a possible role in the terminal differentiation of the intestine. Further analysis of CDX1 should elucidate the function of caudal-type homeobox genes in human development.     

In vivo expression of interleukin-1 beta (IL-1 beta), IL-2, IL-4, IL-6, tumour necrosis factor-alpha and interferon-gamma in the fetal murine thymus

Cytokines are known to play a role in T-cell lymphopoiesis as potent growth or differentiation factors, but many experiments focusing on their role in the thymus have been conducted only in vitro. We have thus used frozen sections obtained from fetal thymuses of normal C57BL 6 mice to investigate by immunohistochemistry the presence of interleukin-1 beta (I4-1 beta), IL-2. IL-4. IL-6. interferon-7 (IFN-7) and tumour necrosis facor-alpha (TNF-alpha). The results reveal that apart from IL-2, which was not detected, all these cytokines display a time-dependent expression pattern in the normal fetal thymus. First, production of IL-4, IL-6 and TNF-alpha is detected around days 13 14; this is followed by a second wave on days 16 17, with a production of IL-1 beta, IL-4 and IL-6, and finally, just before birth (day 19), by a third wave of IL-1 beta, IL-4, IL-6, IFN-7 and TNF-alpha production. This supports the hypothesis that cytokines play a rote in T-cell lymphopoiesis.     

Differential induction of metallothionein synthesis by interleukin-6 and tumor necrosis factor-alpha in rat tissues

Metallothionein (MT) synthesis induced by the inflammatory cytokines, interleukin-6 (IL-6) and tumor necrosis factor-alpha (TNF), was studied in vivo. Administration of recombinant human IL-6 or TNF to rats caused the acute phase responses including rapid decreases in plasma zinc (Zn), and increases in plasma copper (Cu) and ceruloplasmin. Hepatic concentration of MT-I, one of MT isoforms, began to increase within 3 h after the injection of IL-6 or TNF. In IL-6-treated rats, MT-I concentration in liver reached a maximum level at 12 h and decreased with a transient rebound, whereas, in TNF-treated rats, a high level of MT-I lasted for about 48 h. MT-II, the other MT isoform, was induced more than MT-I in liver by both cytokines. MT-I was also induced in lung and heart by TNF, but little by IL-6. The data suggest that IL-6 may be responsible for MT synthesis in liver, whereas TNF may be responsible not only in liver but also in lung and heart. Furthermore plasma concentration of MT did not always reflect the enhanced concentration of MT by TNF and IL-6 in liver, suggesting involvement of many factors influencing plasma MT levels. The interrelation between IL-6 and TNF for MT synthesis has also been discussed.     

Transcriptional activation of the factor VIII gene in liver cell lines by interleukin-6

PURPOSE: The pathogenesis of thrombocytopenia in patients with thrombocytopenia with absent radii (TAR) syndrome has not been clarified yet. PATIENTS AND METHODS: This is the first report of a Japanese patient with TAR syndrome. We studied his megakaryopoiesis in vitro and serum levels of thrombopoietin (TPO). RESULTS: Serum levels of TPO in the patient with TAR syndrome were comparable with those of an age-matched control. The bone marrow cells from the patient with TAR syndrome actually generated megakaryocyte colonies in the presence of TPO and the numbers were significantly greater than those from the age-matched control marrow. However, megakaryocyte colonies from the marrow cells with TAR syndrome contained a much lower number of cells per colony and the size of the individual megakaryocytes appeared to be smaller. CONCLUSION: These data suggest that megakaryocyte progenitors from patients with TAR syndrome may have decreased proliferative and differentiative capacity to respond to TPO, leading to thrombocytopenia.     

Production of tumor necrosis factor, interleukin-6 and prostaglandin E2 by LPS-stimulated rat bone marrow macrophages after thermal injury: effect of indomethacin

The effect of thermal injury on the in vitro production of TNF, IL-6, and PGE2 by bone marrow-derived, LPS-stimulated rat macrophages was studied. Thermal injury caused a general hyperactivity in the production of the mediators by the cells. Indomethacin, a cyclooxygenase inhibitor of PGE2 synthesis, inhibited the production of IL-6 and PGE2 but had no effect on the production of TNF. These results suggest that the observed low concentration of PGE2 produced by the cells was insufficient to cause inhibition of TNF synthesis; thus, the effect of indomethacin would be undetectable. The results also suggest that indomethacin may act directly in inhibiting the production of IL-6 by the macrophages. The hyperactive effect of thermal injury on the production of inflammatory mediators by newly differentiated bone marrow derived macrophages can be important in the overall systemic response to the insult.     

GM1 ganglioside potentiates trimethyltin-induced expression of interleukin-1 beta and the nerve growth factor in reactive astrocytes in the rat hippocampus: an immunocytochemical study

An approach to the evaluation and comparison of reversed-phase high-performance liquid chromatography stationary phases with particular emphasis on data analysis and presentation is described. Assessment is based on the peak efficiency, asymmetry (USP tailing factor) and relative retention properties shown by 24 basic compounds having a wide range of structural and physico-chemical properties. A novel approach to data normalisation and presentation is described. This overcomes the problems associated with the quality of the column packing process, as well as differences in stationary phase selectivity which in conjunction with extra column band broadening effects can make comparisons meaningless.     

Effects and interactions of tumour necrosis factor alpha and bradykinin on interleukin-1 production in gingival fibroblasts

We examined the association of serum albumin concentration with diabetes mellitus and other cardiovascular risk factors, prevalent cardiovascular disease, and ultrasonographically assessed carotid artery intima-media thickness using data from 45- to 64-year-old adults in the Atherosclerosis Risk in Communities (ARIC) Study. The mean albumin concentration was 0.04 to 0.12 g/L lower in participants with diabetes and 0.02 to 0.06 g/L lower in those with cardiovascular disease, compared to participants without these conditions. However, lower serum albumin level was also correlated with most traditional risk factors and hemostatic variables. On adjustment for these, there was essentially no association between serum albumin and prevalent cardiovascular disease. Likewise, there was no association between albumin and carotid intima-media thickness (a marker of atherosclerosis). While hypoalbuminemia may be a marker for chronic disease and perhaps renal loss of albumin, it seems unlikely that it is an important cause of atherosclerosis.