Biomedical subjects

W Fiers

Publications and source records attributed to W Fiers.

533 records · Page 30Linked to original sources

PubMedDate not supplied

Effects of tumor necrosis factor on prostacyclin production and the barrier function of human endothelial cell monolayers.

The endothelium controls the influx of macromolecules into the tissues, a process that may be disturbed at sites of inflammation and in atherosclerotic plaques. In this article, we report our evaluations of the effects of the inflammatory mediator, tumor necrosis factor-alpha (TNF-alpha), on the production of prostacyclin and the barrier function of human endothelial cell monolayers in an in vitro model. TNF-alpha (500 units/ml) had no direct effect on the passage of sucrose, peroxidase, and low density lipoprotein through monolayers of human aortic endothelial cells. On the other hand, during the first hours after addition 500 units/ml TNF-alpha induced a reduction of the permeability of umbilical artery endothelial cell monolayers. Within 10 minutes TNF-alpha induced an increase in prostacyclin production by primary cultures of umbilical artery endothelial cells. However, the reduction in permeability was not caused by a change in prostacyclin production or by a change in cyclic AMP concentration because 1) the effect of TNF-alpha on permeability was not prevented by aspirin, 2) no change in the cellular cyclic AMP concentration could be observed after addition of TNF-alpha, and 3) TNF-alpha was still able to reduce the passage rate in the presence of 25 microM forskolin. The reduction in permeability was accompanied by a decrease of F-actin in stress fibers. With prolonged incubation with TNF-alpha, the permeability of umbilical artery endothelial cell monolayers increased, and F-actin was found again in stress fibers. However, these effects of TNF-alpha were only significant at high concentrations of TNF-alpha.(ABSTRACT TRUNCATED AT 250 WORDS)

Actins↗

PubMedDate not supplied

Inhibition of tumor necrosis factor-induced necrotic cell death by the zinc finger protein A20.

Tumor Necrosis Factor (TNF) is a cytokine that induces necrotic and apoptotic forms of cell death. The TNF-induced signalling mechanisms leading to necrosis or apoptosis are partially distinct, and are therefore likely to be regulated in a different way. The zinc finger protein A20 is a TNF-induced primary response gene that has been shown to inhibit TNF-induced apoptosis. However, its ability to inhibit the necrotic route of cell death as well as the underlying mechanism remains unknown. Here we show that stable expression of A20 or a fusion protein consisting of Green Fluorescent Protein (GFP) and A20 protects the TNF-sensitive fibroblast cell line L929 partially from TNF-induced necrotic cell death. TNF-induced necrosis has been shown to involve the activation of several phospholipases, as well as an increased production of reactive oxygen radicals. The reduced TNF-sensitivity of A20-expressing L929 cells was correlated with a decrease of TNF-induced phospholipase A2 (PLA2), phospholipase C (PLC) and phospholipase D (PLD) activation. Furthermore, production of mitochondrial reactive oxygen intermediates was retarded by overexpression of A20. These results demonstrate that A20 not only inhibits TNF-induced apoptosis but also TNF-induced necrosis, suggesting that it interferes with an early step in TNF signalling which is required for both types of cell death.

Animals↗

PubMedDate not supplied

Organization and chromosomal localization of the human interleukin 5 receptor alpha-chain gene.

The gene for the hIL-5R alpha subunit, which is present in a single copy in the human genome, has been analysed in detail. It is located on chromosome 3 in the region 3p26. The gene organization reflects its relationship to the cytokine/haematopoietin receptor superfamily. Three introns are located in the 5' untranslated region. The subsequent exons determine the functional domains of the hIL-5R alpha protein: the signal peptide, three fibronectin type III-like (FN-like) modules, each built up by two exons, the membrane anchor and two exons forming the cytoplasmic tail, the first of which contains the proline cluster region. In addition, a specific exon generating a soluble isoform is located before the membrane anchor exon. This specific exon contains an in frame TAA stop codon, followed by a polyadenylation signal. Hence, a normal splicing event leads to a soluble IL-5R alpha variant, whereas alternative splicing is required for cell membrane anchoring. A second area of alternative splicing is found in the 5' leader sequence, and possibly relates to the presence of short open reading frames preceding the main ATG. All intron-exon junctions meet the GT-AG rule. The gene structures of all cytokine/haematopoietin receptors documented so far have also been compared with respect to intron phasing. This shows that all introns between the FN-III-like modules are of the +1 type, but in addition, splice sites within the Cys-module and WS-WS-module are invariably of the +2 and 0 type, respectively.

Amino Acid Sequence↗

PubMedDate not supplied

Tumor necrosis factor induces acute phase proteins in rats.

Inoculation of WAG rats with recombinant mouse tumor necrosis factor results in a rapid and marked increase in several acute phase proteins in the serum (haptoglobin, alpha 1 acid glycoprotein, alpha 2 macroglobulin) and in the plasma (fibrinogen). We conclude that TNF may play an important role in the inflammatory response in vivo and possibly in the pathogenesis of inflammatory disorders.

Acute-Phase Proteins↗

PubMedDate not supplied

TNF in combination with interferon-gamma is cytotoxic to normal, untransformed mouse and rat embryo fibroblast-like cells.

The cytostatic and cytolytic effects of the combination of Tumour Necrosis Factor (TNF) and Interferon-gamma (IFN-gamma) on freshly isolated, untransformed and non-tumorigenic mouse embryo fibroblast-like (MEF) as well as rat embryo fibroblast-like (REF) cells are reported. Both TNF and IFN-gamma decreased the proliferation rate of REF cells, and prolonged treatment with the combination of TNF and IFN-gamma was cytolytic to the cells. IFN-gamma alone also inhibited the growth of MEF cells, an activity not shared by TNF, and was even cytolytic after treatment for more than one week. Growth inhibition as well as cytolysis of MEF cells by IFN-gamma was strongly enhanced by TNF. This is one of the few reports regarding the cytotoxic action of the combination of recombinant TNF and IFN-gamma on untransformed cells.

Animals↗

PubMedDate not supplied

Effect of oncogene transfection or passage in vivo on malignant phenotypes of rat2 cells.

Rat2 cells are thymidine kinase-deficient derivatives from the immortalized rat embryo cell line Rat1. They show no phenotypic correlates of malignancy in vitro and produce tumors in syngeneic Fischer rats after long latency periods. We have investigated how transfection with oncogenes would alter the in vitro and in vivo behavior of Rat2 cells. Thus we have manipulated Rat2 cultures in various ways. The cell lines obtained were categorized as parental, in vitro subclones, untransfected in vivo derivatives, non-oncogene (neor and tk) transfectants, oncogene (mutated c-Ha-ras, polyoma middle-T, FBR v-gag-fos-fox) transfectants, and in vivo derivatives of transfectants. They were tested in vitro for morphotype, colony formation in soft agar, growth in organ culture, invasion in organ culture, and in vivo for latency period of tumor formation, tumor growth rate, invasiveness, and metastasis. Differences between the consequences of various manipulations were found in the number of malignancy-related phenotypic alterations. The following trend could be deduced from our data: induction of invasiveness in organ culture by all manipulations; morphotypic transformation and shortening of tumor-latency period by all oncogene transfections and by passage with tumor formation in vivo; growth in organ culture and increased tumor growth rate in vivo by transfection with ras-, or fos-oncogenes and by passage in vivo. Metastatic capability (present in parental Rat2 cell tumors) and colony formation in soft agar (absent in Rat2 cells) were not affected by the present manipulations. We concluded that differences between the oncogene-transfectants and the untransfected in vivo derivatives do not lie in the expression of malignancy-related phenotypes but in the time needed to acquire them.

Animals↗

PubMedDate not supplied

Production of interferon gamma by peripheral blood mononuclear cells from normal subjects and from patients with rheumatoid arthritis.

A radioimmunoassay for human interferon gamma (IFN-gamma) has been carried out using a recombinant glycosylated interferon (Hu IFN-gamma) as tracer, the N.I.H. reference preparation (Gg 23-901-530) and a polyclonal rabbit antiserum. The assay is highly specific for IFN-gamma: there is no cross-reaction either with interferons alpha and beta, Interleukins 1 and 2, tumor necrosis factor alpha and beta or with various brain peptides. The sequential saturation procedure allowed a sensitivity of 0.4 U/ml with intra and between assay coefficients of variation less than 8 and 12%, respectively. The in-vitro production of IFN-gamma by peripheral blood mononuclear cells (P.B.M.C.) was also measured. In unstimulated cultures, IFN-gamma production remained undetectable, i.e. below the 0.4 U/ml sensitivity level. After stimulation of P.B.M.C. from normal subjects with increasing amounts of PHA, both the 3H-thymidine incorporation and IFN-gamma release followed bell-shaped curves. There was no significant difference of 3H-thymidine incorporation between PHA stimulated cultures (0.2 and 2.5 ug/ml) from normal subjects (36 cases) and those with active (16 cases) or non-active (14 cases) rheumatoid arthritis. At two PHA concentrations of 0.2 and 2.5 ug/ml, mononuclear cells from patients with active disease produced significantly less IFN-gamma than those from either controls or cases with non-active disease.

Adult↗

PubMedDate not supplied

Studies on the induction of the interleukin-6 promoter in cell lines of human and simian origin.

In a number of eukaryotic cell types, the promoter of the interleukin-6 (IL-6) gene is inducible by several agents. We studied this inducibility in two human (MG63 and HeLa H21) and four simian cell lines (BSC-1, AP8, CV-1 and VERO). Furthermore, we also investigated whether this inducibility was maintained when a heterologous gene was placed under control of the human IL-6 promoter. As a heterologous gene we used the Simian Virus 40 (SV40) large T antigen, because of its stimulatory effects on SV40-based expression vectors. For the human cell lines tested, we observed that this gene was equally well induced and controlled as the endogenous IL-6 gene. In contrast, the aforementioned simian cell lines were only slightly inducible for IL-6 production; moreover, after transfection with the IL-6 promoter--SV40 T antigen gene construct, a continuous, low level of expression was found, even in those lines which only showed a low (CV-1) or no (BSC-1) endogenous IL-6 background. This apparent difference between the human and simian cells analyzed does not reflect a species specificity, but presumably is related to a different differentiation state of these cells. Furthermore, the IL-6 gene induction could be correlated with activation of the required transcription factor NF-kappa B.

Animals↗

PubMedDate not supplied

Platelet-activating factor is a mediator in tumor necrosis factor/galactosamine-induced lethality.

We here report that administration to mice of WEB2170, a potent platelet-activating factor (PAF) receptor antagonist, prevents both PAF-induced and murine tumor necrosis factor (TNF)-induced lethality in galactosamine (GalN)-sensitized mice. Furthermore, we demonstrate that pretreatment with alpha 1-acid glycoprotein (AGP) or interleukin-1 (IL-1) protects against TNF-induced, but not against PAF-induced lethality. We conclude that PAF is a mediator in TNF/GalN-induced lethal shock, but that the protection conferred by AGP or IL-1 pretreatment is not at the level of scavenging PAF.

Animals↗

PubMedDate not supplied

Inhibition of and sensitization to the lethal effects of tumor necrosis factor.

The extension of the positive results obtained with tumor necrosis factor (TNF) in the locoregional treatment of cancer to systemic treatments requires the selective inhibition of its shock-inducing properties. In this paper, recent data regarding the mechanisms by which infections and tumors render mice extremely sensitive to the lethal effects of TNF as well as regarding the inhibition of the dose-limiting toxicities, hypotension and hepatotoxicity, are summarized. An interleukin-12 (IL-12) driven induction of interferon-gamma (IFN-gamma), probably in synergism with endogenous TNF, was found to mediate infection-induced sensitization. The sensitization induced by tumors develops independent of the IL-12/IFN-gamma axis but ultimately leads to a common step, which can be inhibited by alpha-CD11a and is specific for sensitization. Hypotension can be inhibited by methylene blue (MB), an inhibitor of the nitric oxide (NO)-induced activation of the cytosolic guanylate cyclase, without the indispensable protective properties of NO being affected. Finally, two acute phase proteins, alpha 1-acid-glycoprotein (AGP) and alpha 1-antitrypsin (AT), were able to protect against the TNF-induced liver failure. None of these three inhibitors seems to affect the antitumor effects of TNF.

Animals↗

PubMedDate not supplied

TNF-induced intracellular signaling leading to gene induction or to cytotoxicity by necrosis or by apoptosis.

TNF-induced apoptosis, e.g. in murine PC60 cells, requires the TNF receptor p55 (TNF-R55) and the TNF receptor p75 (TNF-R75); the latter even does not have to be triggered. The intracellular domain of TNF-R55 can be activated in the cytosol by linking it to the trimeric CAT protein; induction of this fusion protein leads to a full TNF response. A new MAP kinase, p38, has been shown to be also activated by TNF. This activation is essential for gene induction, but not for cytotoxicity in L929 cells. TNF treatment of L929 leads to reactive oxygen formation in the mitochondria, resulting in cell death by necrosis. TNF treatment of many other cell types results in apoptosis, and this process involves activation of one or more ICE homologs (IHO). In the mouse, seven cysteine proteases of the IHO family have been cloned and partially characterized. One or more of these IHOs is involved in cell killing by proteolysis of critical substrate(s). One substrate, which may be a key effector molecule in the apoptotic process, is PITSLRE kinase.

Animals↗