PubMed Health⌕ Search

Biomedical subjects

W Falk

Publications and source records attributed to W Falk.

At least 73 records · Page 4Linked to original sources
PubMed1997

Neutralization of tumour necrosis factor (TNF) but not of IL-1 reduces inflammation in chronic dextran sulphate sodium-induced colitis in mice.

The cytokines TNF and IL-1 have been implicated as mediators of the inflammatory processes in patients with inflammatory bowel disease (IBD). To investigate the role of these cytokines in mucosal inflammation we used anti-cytokine strategies in a mouse model of acute and chronic colitis. Mice which received 5% dextran sulphate sodium (DSS) in their drinking water showed signs of acute colitis on day 4, with severe weight loss and bloody diarrhoea. Chronic colitis was established after four cycles of feeding 5% DSS for 7 days and water for 10 days, with the mice showing diarrhoea but no weight loss. In acute colitis, treatment with anti-IL-1 reagents, anti-TNF MoAb, or dexamethasone (DEX) led to aggravation. By contrast, in chronic colitis, treatment of mice with several IL-1 activity-inhibiting reagents failed to show significant effects, whereas anti-TNF MoAb or DEX significantly reduced the colitis. We conclude that in acute colitis IL-1 and TNF are beneficial, whereas in chronic colitis, TNF but not IL-1 seems to play a major role in perpetuation of chronic inflammation.

Acute Disease↗
PubMed1997

The interleukin-1 receptor complex and interleukin-1 signal transduction.

The aim of this mini-review is to give an update on the current developments concerning the IL-1 plasma membrane receptor complex and the modes of signal transduction through the plasma membrane. The mechanisms of signal generation at the cytosolic side of the plasma membrane will be summarized. Possible coupling mechanisms to down-stream signalling elements will be discussed, which result in the complex activation of transcription factors responsible for the cellular responses to IL-1. It is not the aim of this review to extensively summarize IL-1 induced cytosolic kinase cascades and activation of transcription factors.

Animals↗
PubMed1996

Molecular cloning and expression of mouse procalcitonin.

The nucleotide sequence for mouse calcitonin was determined from a cDNA obtained using a polymerase chain reaction (PCR) based method, the rapid amplification of cDNA ends (RACE)-PCR. Primers designed from highly conserved regions in the coding sequences of known rat and human calcitonin cDNAs were used to amplify calcitonin cDNA as 5'-end and 3'-end fragments from mouse thyroid RNA. The obtained cDNA is 850 bp in length most probably representing the entire mouse calcitonin mRNA. It contains an open reading frame coding for a 136 amino acid protein with a calculated M(r) of 15,143. Comparison of the deduced amino acid sequences of preprocalcitonin of mice with other species revealed highest homologies to the rat (93%) and human (77%) sequences. A recombinant form of mouse precalcitonin (rmPCT) of approximately 17 kDa was expressed as a fusion peptide in E.coli transformed with a PCR-cloned expression construct.

Amino Acid Sequence↗
PubMed1996

Neuroimmune control of interleukin-6 secretion in the murine spleen. Differential beta-adrenergic effects of electrically released endogenous norepinephrine under various endotoxin conditions.

In a previous study we demonstrated a superfusion technique which allows for investigation of nerve-immune cell interaction in murine spleen. We demonstrated that under septic-like conditions in the presence of bacteria and lipopolysaccharide (LPS), electrically induced inhibition of interleukin 6 (IL-6) secretion was attenuated by the beta-adrenergic antagonist propranolol. This effect was now investigated more closely under various endotoxin conditions in order to dissect effects of bacteria and endotoxin: (A) bacteria-rich conditions (without penicillin/streptomycin [P/S] and without LPS), (B) LPS-enriched conditions (with P/S and with LPS), and (C) bacteria-free conditions (with P/S and without LPS). Under bacteria-rich conditions, norepinephrine (Emax = 10(-6) M, p = 0.012) and isoproterenol (Emax = 10(-6) M, p = 0.048) concentration-dependently inhibited IL-6 secretion from murine spleen slices in contrast to bacteria-free conditions. In a bacteria-free environment the beta-adrenergic antagonist propranolol did not attenuate the electrically induced inhibition of splenic IL-6 secretion. The insertion of bacterial filters in front of the superfusion chambers to avoid direct contact between bacteria and cells increased the electrically-induced inhibition of IL-6 secretion (p = 0.0036). Added LPS did not change the electrically-induced release of norepinephrine from presynaptic nerve terminals in murine spleen. The study demonstrates two different beta-adrenergic effects on IL-6 secretion of murine spleen slices under bacteria-rich or bacteria-free conditions.

Adrenergic beta-Antagonists↗
PubMed1996

Increased interleukin 8 expression in the colon mucosa of patients with inflammatory bowel disease.

To test whether there is a difference in the expression of interleukin 8 (IL8) between Crohn's disease and ulcerative colitis and to determine the main site of its synthesis this study analysed IL8 in mucosal biopsy specimens of patients with Crohn's disease and ulcerative colitis by enzyme linked immunosorbent assay (ELISA) and by in situ hybridisation. IL8 was measured by ELISA in 38 normal control patients, eight inflammatory control patients, 55 Crohn's disease biopsy specimens (26 patients), and 67 ulcerative colitis biopsy specimens (35 patients). IL8 mRNA was determined in samples by in situ hybridisation using a specific IL8 RNA probe. IL8 protein was significantly increased in macroscopically inflamed specimens of Crohn's disease (median 118 pg/specimen, p < 0.0001), ulcerative colitis (median 140 pg/specimen, p < 0.001), and inflammatory controls (median 30 pg/specimen, p = 0.010) compared with normal controls (median 4 pg/specimen). IL8 was also increased in uninflamed specimens of Crohn's disease (median 46 pg/specimen, p < 0.001) but not of ulcerative colitis patients (median 9 pg/specimen, p = 0.3). IL8 protein in the mucosa correlated significantly with macroscopic inflammation in Crohn's disease (r = 0.47, p < 0.001) and in ulcerative colitis (r = 0.60, p < 0.001). IL8 mRNA was detected by in situ hybridisation in 31 of 55 biopsy specimens (56%) of Crohn's disease patients, in 38 of 67 specimens of ulcerative colitis patients (57%), in five of eight inflammatory controls (63%) and in five of 38 normal controls (13%). Mucosal IL8 mRNA expression correlated with mucosal IL8 protein (r = 0.46, p < 0.001). IL8 mRNA was only detected in inflammatory cells of the interstitium but not in mucosal epithelial cells. IL8 is produced mainly in the lamina propria of the colon in inflammatory bowel disease and correlates with mucosal inflammation.

Adolescent↗
PubMed1995

Regulation of interleukin-8 production in a human colon epithelial cell line (HT-29).

BACKGROUND/AIMS: Interleukin (IL) 8 is a major neutrophil-activating cytokine synthesized by intestinal epithelial cell lines. The aim of this study was to improve the understanding of the regulation of IL-8 synthesis by investigating the roles of protein kinase C (PKC), protein kinase A (PKA), and protein tyrosine kinase (PTK) in the induction of IL-8. METHODS: HT-29 cells were stimulated with IL-1 beta or tumor necrosis factor alpha (TNF-alpha) together with activators or inhibitors of PKC and PKA or with inhibitors of PTK. The presence of IL-8 protein was detected by enzyme-linked immunosorbent assay and that of IL-8 messenger RNA by Northern blotting and in situ hybridization. RESULTS: TNF-alpha and IL-1 beta dose-dependently induced IL-8 production in HT-29 cells. Activation of PKC by phorbol myristate acetate also stimulated IL-8 production; however, the effects of IL-1 beta or TNF-alpha did not require PKC, as shown by the PKC inhibitor staurosporin or PKC depletion. Stimulation of PKA by forskolin or inhibition by H89 or H7 had no influence on the synthesis of IL-8. However, induction of IL-8 by IL-1 beta or TNF-alpha was reduced by the PTK inhibitors herbimycin (by 79% or 89%, respectively) and genistein (by > 95%). CONCLUSIONS: The synthesis of IL-8 is stimulated in HT-29 cells by IL-1 beta or TNF-alpha. This stimulation is independent from PKC or PKA but depends on protein tyrosine phosphorylation.

Cell Line↗
PubMed1995

In vitro superfusion method for the investigation of nerve-immune cell interaction in murine spleen.

Immune defence mechanisms can be modulated by brain function. To study such interactions, an in vitro method was developed to examine the release of cytokines and norepinephrine (NE) after electrical stimulation. Slices of mouse spleen were placed in chambers with a volume of 80 microliters and superfused with culture medium. To characterize electrically evoked NE release and cell viability a suitable stimulation protocol was developed using of [3H]NE. As parameter for immune function, modulation of interleukin-6 (IL-6) release by the spleen cells brought about by electrical stimulation was investigated. Splenic [3H]NE overflow was calcium-dependent, tetrodotoxin-sensitive and elicited by 54 mM potassium. Electrically evoked [3H]NE release at 22 h was about 80% of the release at 5.3 h. Electrical stimulation substantially reduced IL-6 secretion at 6 h (control: 143.4 +/- 14.3 vs. electrical: 71.3 +/- 7.9 pg/ml/10(6) leukocytes, P = 0.0001). This effect was inhibited in a concentration-dependent manner by the beta-adrenergic antagonist propranolol (P = 0.0298, EC50 approx. 10(-7) M). In conclusion, this new technique allows long-term investigation of cell function in slices of murine spleen. In addition, these are the first in vitro data indicating the presence of a functional neuroimmunological link in murine lymphoid tissue.

Adrenergic beta-Antagonists↗
PubMed1994

Mechanisms involved in metastasis enhanced by inflammatory mediators.

The enhancement of tumor metastasis by concurrent inflammatory processes is mainly due to the cytokines TNF and IL-1. In the case of TNF this effect is not restricted to metastasis models as measured by in vivo colony formation but also found in experimental model systems of spontaneous metastasis. Direct effects on the tumor cells or interference with the host NK cell system did not seem to account for the observed TNF effect. Experimental evidence from different test systems rather points to TNF- or IL-1-induced enhanced adhesion of tumor cells to the endothelial cell layer as the underlying mechanism. Blocking of integrin-matrix interactions with monoclonal antibodies or competing peptides inhibited tumor cell adhesion to endothelioma cells in vitro and lung colony formation of tumor cells in vivo.

Animals↗
PubMed1994

Activation of the mouse IL-2 gene by okadaic acid: synergy with interleukin-1.

Interleukin-1 (IL-1) has potent immunoregulatory and inflammatory functions. Its activity is mediated by an 80-kDa receptor on the cell surface and leads to activation of other genes. The underlying molecular events are largely unknown. We investigated the role of phosphatases in activation of the IL-2 gene in EL4 thymoma cells. We found that the protein phosphatase PP1 and PP2A inhibitor okadaic acid (OA) alone was able to significantly stimulate IL-2 production by the IL-1-responsive EL4 subline EL4 5D3 and also by the IL-1-nonresponsive EL4 subline EL4D6/76. In the IL-1-responsive cell line OA strongly synergized with phorbol myristate acetate (PMA) and IL-1. In the IL-1-nonresponsive cell line OA synergized with PMA but not with IL-1. Under suboptimal conditions of PMA/OA synergy an additional synergistic effect of IL-1 was shown. This was true for IL-2 and IL-6 production. Sphingomyelinase or sphingosine had no detectable effect. The kinetics of OA- and PMA-induced expression of IL-2 mRNA and IL-2 protein was different. PMA induced maximal expression between 6 and 12 h and was almost undetectable at 24 h. OA-induced expression was first obvious at 12 h and continued longer than 36 h. In both cases IL-1 caused no shift in kinetics, but potentiated the effects of the different tumor promoters. Utilizing IL-2 promoter-CAT constructs we showed in transfection experiments that the synergistic effect was also evident on the transcriptional level. We conclude from the data that phosphatases play an important role for IL-2 expression and that IL-1 can use additional pathways of activation that are different from events induced by PMA or OA.

Animals↗
PubMed1993

Enhancement of experimental metastasis by tumor necrosis factor.

The influence of endogenous and exogenous tumor necrosis factor (TNF) on metastasis was investigated in an experimental fibrosarcoma metastasis model. A single intraperitoneal injection of recombinant human (rh) TNF or recombinant mouse (rm) TNF into mice 5 h before intravenous inoculation of methylcholanthrene-induced fibrosarcoma cells (CFS1) induced a significant enhancement of the number of metastases in the lung. Dose responses of rmTNF and rhTNF demonstrated a stronger metastasis-augmenting effect by rmTNF compared with rhTNF. This effect was time dependent, as administration of rmTNF 5 h before or 1 h but not 24 h after tumor cell inoculation caused an increase of tumor cell colony formation on the lung surface, suggesting an influence of TNF on the vascular adhesion and diapedesis of tumor cells. Since tumor-bearing mice showed an enhanced ability to produce TNF after endotoxin injection compared to control mice, tumor-bearing mice were treated with anti-mTNF antibodies. Neutralization of endogenous tumor-induced TNF led to a significant decrease of the number of pulmonary metastases. Histological analysis of micrometastases in the lung on day 5 by silver staining of proteins associated with nucleolar organizer regions revealed more metastatic foci and augmented proliferative activity of the tumor cells after rmTNF pretreatment of mice. However, no direct effect of rmTNF on the proliferation rate of tumor cells was seen in vitro. These findings suggest that low doses of endogenous TNF or administered TNF during cytokine therapy might enhance the metastatic potential of circulating tumor cells.

Animals↗
PubMed1993

An NF-kappa B-like element plays an essential role in interleukin-1-mediated costimulation of the mouse interleukin-2 promoter.

Interleukin-1 (IL-1) costimulation is required for efficient IL-2 synthesis and IL-2 receptor (IL-2R) expression of T cells. The molecular events leading to these effects are largely unknown. We utilized an IL-1-responsive and an IL-1-non-responsive subclone of the mouse thymoma cell line EL4 to investigate how IL-1 activates IL-2 gene expression. We correlated IL-2 promoter activity with the activity of the endogenous IL-2 gene, thereby showing the biological significance of our results. Our experiments provide new functional data showing that a major target of IL-1 mediated costimulation is the chi B-like site, T cell element distal TCEd (GGGATTTCAC), of the IL-2 promoter. Thus, deletion or mutation of TCEd within a complete IL-2 promoter abrogated IL-1 costimulation in the IL-1 responsive EL4 subclone. Therefore, the TCEd element is functionally essential for the effect of IL-1. We also identified a nuclear factor (NF), IL-1 NF, that binds to the TCEd site after IL-1 stimulation. This factor was only present in the IL-1-responsive EL4 subclone and not in the IL-1-non-responsive subclone after IL-1 stimulation and did not appear after phytohemagglutinin (PHA)-treatment. Binding of IL-1 NF to the TCEd site was competed by a typical chi B oligonucleotide, suggesting that it is similar to NF-chi B in its DNA-binding properties. However, the TCEd element was only activated by costimulation with PHA and IL-1 whereas a typical chi B element was already activated by IL-1 alone. These data suggest that the biological function of the TCEd element of the IL-2 promoter differs from that of a canonical chi B element. Our data provide new evidence that IL-1 acts on the IL-2 promoter by activating the TCEd element via the transcription factor IL-1 NF. Furthermore, activation of this element requires two signals, delivered by IL-1 and PHA, in this way reflecting the activation requirement for the endogenous IL-2 gene.

Animals↗
PubMed1992

Purification and molecular cloning of the APO-1 cell surface antigen, a member of the tumor necrosis factor/nerve growth factor receptor superfamily. Sequence identity with the Fas antigen.

The APO-1 antigen as defined by the mouse monoclonal antibody anti-APO-1 was previously found to be expressed on the cell surface of activated human T and B lymphocytes and a variety of malignant human lymphoid cell lines. Cross-linking of the APO-1 antigen by anti-APO-1 induced programmed cell death, apoptosis, of APO-1 positive cells. To characterize the APO-1 cell surface molecule and to better understand its role in induction of apoptosis, the APO-1 protein was purified to homogeneity from membranes of SKW6.4 B lymphoblastoid cells by solubilization with sodium deoxycholate, affinity chromatography with anti-APO-1 antibody, and reversed phase high performance liquid chromatography. Each purification step was followed by an APO-1-specific solid phase enzyme-linked immunosorbent assay using the monoclonal antibody anti-APO-1. In sodium dodecyl sulfate-polyacrylamide gel electrophoresis, the APO-1 antigen was found to be a membrane glycoprotein of 48-kDa. Endoproteinase-cleaved peptides of the APO-1 protein were subjected to amino acid sequencing, and corresponding oligonucleotides were used to identify a full-length APO-1 cDNA clone from an SKW6.4 cDNA library. The deduced amino acid sequence of APO-1 showed sequence identity with the Fas antigen, a cysteine-rich transmembrane protein of 335 amino acids with significant similarity to the members of the tumor necrosis factor/nerve growth factor receptor superfamily. The APO-1 antigen was expressed upon transfection of APO-1 cDNA into BL60-P7 Burkitt's lymphoma cells and conferred sensitivity towards anti-APO-1-induced apoptosis to the transfectants.

Amino Acid Sequence↗
PubMed1991

Endogenous interferon specifically regulates Newcastle disease virus-induced cytokine gene expression in mouse macrophages.

In macrophages from inbred mice, the magnitude of the interferon (IFN) response to Newcastle disease virus (NDV) infection is under genetic control of the If-1 locus, which carries the allele for either high (h) or low (l) IFN production. Here, we report that the activity of genes within the If-1 locus is influenced by macrophage-derived endogenous IFN. In addition to various other biological effects, we observed that endogenous IFN specifically downregulated NDV-induced IFN and interleukin 6 production. Preculture of bone marrow-derived macrophages (BMM) from BALB/c (If-1l) mice in macrophage colony-stimulating factor plus anti-IFN-beta provoked a 30- to 50-fold increase in NDV-induced cytokine production compared with induced control cultures in macrophage colony-stimulating factor alone, whereas only a 4- to 6-fold increase was observed in anti-IFN-beta-treated BMM from C57BL/6 (If-1h) mice. This resulted in nearly complete abrogation of the genetically determined difference in the response to NDV. The increase was specific for NDV and was marked by strong additional activation of IFN-alpha genes. Studies using BMM from B6.C-H28c If-1l congenic mice gave results identical to those obtained with BALB/c BMM. Addition of 20 IU of recombinant IFN-alpha 4 to anti IFN-beta-treated macrophages from B6.C-H28c mice 20 h prior to NDV infection strongly downregulated the IFN-alpha, IFN-beta, and interleukin 6 responses. The genetic difference between macrophages from If-1h and If-1l mice was thus reestablished, since the same treatment caused only weak reduction of NDV-induced cytokine gene expression in BMM from C57BL/6 mice. These data suggest that the If-1h and If-1l alleles harbor IFN-inducible genes that, following activation, specifically suppress subsequent cytokine gene expression in response to NDV.

2',5'-Oligoadenylate Synthetase↗
PubMed1990

Requirement of endogenous tumor necrosis factor/cachectin for recovery from experimental peritonitis.

By intrasplenic immunization we raised a rat mAb (mAb V1q; IgG2a, kappa) with a potent neutralizing activity against natural mouse TNF (1 microgram/ml mAb V1q/100 U/ml TNF). mAb V1q was used to study the role of endogenous TNF in experimental peritonitis induced by sublethal cecal ligation and puncture. mAb V1q persisted for over 5 days in the serum of mice injected with 100 micrograms of the antibody and, therefore, proved useful for in vivo experiments. As little as 20 micrograms mAb V1q/mouse prevented lethal shock of the animals by 400 micrograms LPS/mouse. In sublethal cecal ligation and puncture i.p. injection of mAb V1q directly and up to 8 h after induction of experimental peritonitis lead to death of the animals within 1 to 3 days. The lethal effect of mAb V1q was compensated by injection of recombinant mouse TNF. Similar mAb V1q effects as in immunocompetent mice were shown in severe combined immune deficiency mice deficient of mature functional B and T cells. Taken together, these data suggest that during the early phase of peritonitis endogenous TNF may stimulate nonlymphoid cells such as granulocytes, macrophages, platelets, and fibroblasts to ingest bacteria and to localize inflammation, respectively. These beneficial effects of TNF may determine survival. Thus, our data may have implications for the therapeutic management of a beginning peritonitis.

Animals↗
PubMed1990

Inhibition of murine cytotoxic T cell responses by progesterone.

In the present study we evaluated the effect of sex hormone on the generation of murine cytotoxic T cell responses. We show that the in vitro CTL response was strongly inhibited by progesterone but not by E1, E2 or testosterone. Our experiments attempting to understand the mechanism of the hormone action on CTL development have revealed that the ability of the cells to generate helper signals was not affected. This was demonstrated by the fact that neither IL-2 production nor IL-2 receptor expression was altered by the hormone. Rather, it appears that the capacity of the cells to respond to the signals and to become cytotoxic was modified. Furthermore, we show that the hormone mediated an inhibition of CTL development in thymocyte cultures externally supplemented with all the required helper factors. These results strongly suggest that progesterone has a direct effect on the differentiation of cytotoxic effector cells.

Animals↗
PubMed1990

Tumor-induced tumor necrosis factor production in macrophages.

Tumor-associated tumor necrosis factor (TNF) production in patients as well as a TNF-inducing membrane constituent of tumor cells have been reported. In a murine fibrosarcoma model we analyzed TNF production during growth of a tumor transplant. In situ hybridization showed that a gradually increasing number of cells within the tumor tissue became positive for TNFmRNA. Also, in spleen cells of tumor-bearing mice TNFmRNA became more abundant in later stages of tumor growth compared to early stages. In plasma of these animals, however, TNF activity was not detected at any time even after stimulation with bacterial endotoxin. Neutralization with monoclonal antibodies of endogenous TNF during tumor growth did not affect the growth rate of the tumor, indicating that either the antibodies did not reach the relevant TNF production and action sites or that endogenously produced TNF did not play a significant role in this tumor model.

Animals↗