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Biomedical subjects

F Herrmann

Publications and source records attributed to F Herrmann.

At least 91 records · Page 5Linked to original sources
PubMed1995

Tumor necrosis factor (TNF)-alpha activates c-raf-1 kinase via the p55 TNF receptor engaging neutral sphingomyelinase.

TNF-alpha mediates proliferation, functional activation and apoptotic death of cells depending upon its concentration and target cell type. The signaling pathways used by TNF-alpha to mount these responses are, at present, not completely understood. We report here that TNF-alpha promotes dose- and time-dependent phosphorylation and activation of the c-raf-1 kinase engaging the type I p55 TNF receptor (TNF-R). c-raf-kinase activation was duplicated by an agonistic monoclonal antibody directed against the p55 TNF-R. Moreover, ectopic expression of the human p55 TNF-R in murine pre-B 70Z/3 cells was sufficient to confer c-raf-1-kinase activation by human TNF-alpha. By inhibiting intracellular activation of acidic sphingomyelinase (SMase) and by using deleted forms of the type I TNF-R it was shown that the neutral, but not the acidic SMase, participated in TNF-alpha-mediated phosphorylation and activation of the c-raf kinase. TNF-alpha-induced transcriptional activation of a heterologous promoter construct harboring the AP-1 binding site was also mediated by the type I p55 TNF-R. In this case the initiation of transcription required the same cytoplasmic domain as that responsible for activation of c-raf-1 kinase and was liberated in the presence of a dominant negative mutant of c-raf-1.

Animals↗
PubMed1995

Functional NF-IL6/CCAAT enhancer-binding protein is required for tumor necrosis factor alpha-inducible expression of the granulocyte colony-stimulating factor (CSF), but not the granulocyte/macrophage CSF or interleukin 6 gene in human fibroblasts.

Tumor necrosis factor (TNF) alpha participates in the regulation of the acute-phase, immune, and inflammatory responses. Target genes known to be transcriptionally activated by TNF-alpha include the granulocyte (G)-colony-stimulating factor (CSF) gene, the granulocyte/macrophage (GM)-CSF gene, as well as the interleukin (IL) 6 gene. Functional nuclear factor (NF)-IL6 recognition sites have been identified in regulatory regions of these genes by transient transfection studies using deleted promoter constructs. In addition, NF-IL6 is known to form heterodimeric complexes with the NF-kappa B transcription factor, which is also engaged in the transcriptional regulation of these genes. The indispensable importance of NF-IL6 for regulating gene expression of proinflammatory cytokine genes in response to inflammatory stimuli in vivo remains, however, unclear. We here report, by using both antisense (AS) oligodesoxyribonucleotide (ODN) and ribozyme (RZ)-mediated specific elimination of NF-IL6 transcripts in human fibroblasts, that TNF-alpha-induced synthesis of G-CSF, but not of GM-CSF or IL-6, is abolished in the absence of functional NF-IL6 in vivo. Both AS ODN and RZ targeting of the NF-IL6 transcript eliminate NF-IL6 protein, as shown in Western blot analysis and electrophoretic mobility shift assays. Similarly, fibroblasts exposed to either the AS NF-IL6 ODN or the NF-IL6 RZ, but not to the sense or nonsense ODN or a mutated ribozyme, also failed to respond with functional activation of NF-IL6 as assayed in transient transfection studies using heterologous promoter constructs harboring the NF-IL6 recognition site. In contrast, protein synthesis, DNA-binding activity, and transcriptional activation capacity of the NF-kappa B transcription factor is not impaired upon exposure to either ODN or RZ. Fibroblasts that had been cultured in the presence of the AS NF-IL6 ODN or the NF-IL6RZ failed to synthesize G-CSF protein in response to TNF-alpha, while TNF-alpha-inducible transcription and release of GM-CSF and IL-6 was preserved.

Base Sequence↗
PubMed1995

MAPKAP kinase 2 is activated by heat shock and TNF-alpha: in vivo phosphorylation of small heat shock protein results from stimulation of the MAP kinase cascade.

The activation of MAPKAP kinase 2 was investigated under heat-shock conditions in mouse Ehrlich ascites tumor cells and after treatment of human MO7 cells with tumor necrosis factor-alpha (TNF-alpha). MAPKAP kinase 2 activity was determined using the small heat-shock proteins (sHsps) Hsp25 and Hsp27 as substrates. In both cell types, about a threefold increase in MAPKAP kinase 2 activity could be detected in a time interval of about 10-15 min after stimulation either by heat shock or TNF-alpha. Phosphorylation of MAPKAP kinase 2, but not the level of MAPKAP kinase 2 mRNA, was increased after heat shock in EAT cells. It is further shown that activation of MAPKAP kinase 2 in MO7 cells is accompanied by increased MAP kinase activity. These data strongly suggest that increased phosphorylation of the sHsps after heat shock or TNF-alpha treatment results from phosphorylation by MAPKAP kinase 2, which itself is activated by phosphorylation through MAP kinases. Hence, we demonstrate that MAPKAP kinase 2 is responsible not only for phosphorylation of sHsps in vitro but also in vivo. The findings link sHsp phosphorylation to the MAP kinase cascade, explaining the early phosphorylation of sHsp that is stimulated by a variety of inducers such as mitogens, phorbol esters, thrombin, calcium ionophores, and heat shock.

Animals↗
PubMed1995

Cancer gene therapy: principles, problems, and perspectives.

Despite enormous efforts focused on the development of new drugs and the use of novel drug combinations, including high-dose regimens supported by bone marrow and blood stem cell transplantation procedures, progress in the treatment of disseminated human cancer has been marginal. Remarkable advances in our understanding of the molecular biology of cancer has provided the possibility to employ new, selective tools of genetic intervention for more successful tumor treatment. We are now witnessing the inception of gene therapy. However, gene therapists face many drawbacks, including selectivity, specificity, sensitivity, and safety of gene transfer. Despite this there are already over 70 clinical protocols accepted for genetic approaches to cancer worldwide. Strategies currently under clinical investigation and discussed here include: (a) the enhancement of tumor immunogenicity by insertion of cytokine genes, genes coding for products of the major histocompatibility complex, and those for lymphocyte costimulatory ligands, (b) the vectoring of tumoricidal cytokines into cells that can potentially home on tumors to release their toxic products locally, (c) the use of tumor-specific pro-drug activators, i.e., the insertion of enzymatically pro-drug-activating genes fused to promoter systems which rely on differential (ideally tumorspecific) transcription control, (d) gene-marking strategies which may provide new indicators for minimal, residual, and relapsed tumor disease, (e) artificial repression of gene functions by insertion of genes encoding for complementary (antisense) mRNA to the gene of interest (e.g., oncogenes, drug resistance genes).

Forecasting↗
PubMed1995

Dose-intensified treatment of breast cancer: current results.

Chemotherapy has an established role in the treatment of carcinoma of the mammary gland (breast cancer), but when administered at conventional doses the net benefit in terms of relapse prevention and overall survival duration remains limited largely to operable patients at first diagnosis with limited tumor burden (stage II disease with less than four involved axillary lymph nodes). Pilot studies evaluating significant dose escalation in patients with advanced disease have yielded high remission rates, but the impact on long-term survival remains controversial. Recent trials involving dose-intense treatment in stage IV breast cancer do not support its use except in patients who have achieved complete remission or show no evidence of disease prior to high-dose consolidation. More optimism may be warranted for the use of dose-intense chemotherapy in the high-risk adjuvant setting (stage II/IIIA) and as part of the initial treatment for patients with inflammatory breast cancer or locally advanced primary inoperable carcinoma (stage IIIB). Prospective randomized trials in all settings are ongoing, although definitive results are not expected before 1998. Apart from single-course myeloablative high-dose chemotherapy, the availability of hematopoietic growth factors through recombinant DNA technology and the easy procurement of hematopoietic cell support through mobilization of peripheral blood progenitors has spurred the development of new strategies employing dose-intense treatment within the past 10 years. Repetitive application of chemotherapy at submyeloablative doses and sequential accelerated dose-intense application of single agents are now increasingly being integrated into more complex dose-escalated protocols. This review will focus on the results of mature trials and newer approaches to dose escalation.

Breast Neoplasms↗
PubMed1995

Ribozymes: biology, biochemistry, and implications for clinical medicine.

Ribozymes are a class of ribonucleic acid (RNA) molecules that possess enzymatic properties. Upon binding to complementary nucleic acid strands, catalytic degradation takes place via a cleavage reaction. In effect, inactivation of susceptible substrate RNA molecules takes place at a catalytic rate and with a high degree of substrate specificity. This article reviews the biology and biochemistry of this class of molecules and its potential applications in clinical medicine.

Genetic Therapy↗
PubMed1995

Interleukin 5 expressing allergen-specific T-lymphocytes in patients with house dust mite sensitization: analysis at a clonal level.

Interleukin 5 (IL-5) is a T-cell lymphokine known to stimulate development, functional activity, and in vitro survival of eosinophils. Tissue and blood eosinophilia occurring during allergic responses of the immune system are potentially mediated by IL-5 secreting T-cells. To test this hypothesis a series of allergen-specific T-cell clones were established from peripheral blood and skin lymphocytes of patients with atopic dermatitis and house dust mite sensitization. In addition, alloreactive T-cell clones were also prepared from peripheral blood lymphocytes of healthy donors. Cloned T-cells were analyzed for IL-5 mRNA expression and IL-5 secretion by means of in vitro gene amplification using the reverse transcriptase polymerase chain reaction and IL-5 specific oligonucleotide hybridization, as well as IL-5-specific ELISA. A majority of allergen-specific long-term cultured T-cell clones (84%) of different donors and of either phenotype (CD8+ or CD4+) disclosed IL-5 transcripts on stimulation with lectins. Almost all clones exhibiting IL-5 transcripts also released immunoreactive IL-5 protein into their culture supernatants. In contrast, only 2% of alloreactive T-cell clones obtained from healthy donors and none of alloreactive T-cell clones of one atopic patient investigated expressed detectable amounts of IL-5 mRNA in response to lectin stimulation, all of whom were CD4+. These results suggest that eosinophilia observed in allergic responses in the peripheral blood and in tissues at the site of induced late-phase cutaneous reaction may be associated with IL-5 release by allergen-specific T-cells.

Animals↗
PubMed1995

Is neuroblastoma screening evaluation needed and feasible?

Despite the five million children who have been screened for neuroblastoma in Japan through detection of catecholamine metabolites, it is still uncertain whether screening for this disease is beneficial. The Japanese study has clearly indicated that screening at 6 months or earlier leads to heavy overdiagnosis. It is shown in this paper that screening at a later age may give the same reduction in mortality with possibly less overdiagnosis. However, it is estimated that, even with two screens at 12 and 18 months, the reduction in mortality would not greatly exceed 25%, under realistic hypotheses on the length of the preclinical phase of the disease. The evaluation of the efficacy of this screening strategy would need the recruitment of half a million children per year over 5-7 years and the follow-up of an equal number of controls. Such a trial would improve our knowledge of the natural history of the disease and might help to answer some questions raised recently regarding its biological heterogeneity.

Age Factors↗
PubMed1995

The role of tyrosine kinases and their substrates in signal transmission of hematopoietic growth factors: a short review.

Both structure-function analysis of hematopoietic growth factor receptors and identification of novel signal transduction molecules have provided new insights into the processes involved in signal transmission pathways engaged by hematopoietic growth factors. These investigations have pointed to the importance of post-translational modifications of pre-existing proteins, in particular tyrosine phosphorylation, in transmitting signals and thereby linking extracellular signals to the activation of nuclear effector molecules which govern gene expression. These observations not only contribute to our understanding of the pleiotropism and redundancy ascribed to hematopoietic growth factors, but also help to trace some of the molecules conferring signal specificity. It is to be expected that this rapidly evolving research field will provide us with a significant collection of new findings in the near future and that the precise understanding of the processes involved in ligand-binding and signal transmission will also stimulate the development of novel therapeutic drugs affecting these processes. This article gives a short overview on the role of tyrosine kinases and their substrates in signal transmission processes initiated by hematopoietic growth factors.

Amino Acid Sequence↗
PubMed1995

Interleukin (IL)-6 signaling leads to phosphorylation of the small heat shock protein (Hsp)27 through activation of the MAP kinase and MAPKAP kinase 2 pathway in monocytes and monocytic leukemia cells.

Interleukin-6 is a multifunctional cytokine which regulates various aspects of the host immune response. Here we show that signaling events transferred by IL-6 in monocytes and the U937 human monocytic leukemia cell line lead to the phosphorylation of the small heat shock protein (Hsp)27. Phosphorylation of Hsp27 is both dose- and time-dependent. In the absence of NaF, a serine/threonine phosphatase inhibitor, IL-6 failed to initiate Hsp27 phosphorylation in vitro. IL-6 also failed to phosphorylate Hsp27 when cells had been deactivated with tyrosine kinase inhibitors such as genistein. The capacity of cellular extracts to phosphorylate Hsp27 could be, however, restored when either immunoprecipitated activated MAP kinase or purified MAPKAP kinase 2 was added to cell lysates. These findings suggest that IL-6-mediated phosphorylation of Hsp27 results from activation of MAPKAP kinase 2, a serine/threonine kinase which is activated by MAP kinase. Taking together, our findings indicate that IL-6-induced activation of MAP kinase by IL-6 entails the activation of MAPKAP kinase 2 and subsequent phosphorylation of the Hsp27.

Calcium-Calmodulin-Dependent Protein Kinases↗
PubMed1995

Expression of c-fos correlates with IFN-alpha responsiveness in Philadelphia chromosome positive chronic myelogenous leukemia.

This study evaluates (i) constitutive levels of oncogene and p53 transcripts in chronic phase CML patients and (ii) their modulations subsequent to in vivo therapy with rIFN-alpha 2c. Peripheral blood mononuclear cells (pbmc) and bone marrow cells of 26 patients were examined for c-fos, c-myc, p53 and the hybrid bcr/abl mRNA levels. Results indicated that (i) constitutive c-fos transcript levels are significantly higher in patients subsequently responding to IFN-alpha therapy (p < 0.01) and positively correlated with the proportion of lymphocytes (r = 0.6895, p < 0.01) and negatively with the proportion of immature cells (r = -0.568, p < 0.01) contained in the pbmc preparations tested, (ii) constitutive mRNA levels of the hybrid bcr/abl, c-myc and p53 are positively correlated with each other, but failed to relate to disease parameters, and (iii) acute and chronic in vivo exposure to IFN-alpha is accompanied by upregulation of c-fos and downregulation of c-myc mRNA levels in responder patients.

Bone Marrow Cells↗
PubMed1995

Modulation of vindesine and doxorubicin resistance in multidrug-resistant pleural mesothelioma cells by tumor necrosis factor-alpha.

Tumor necrosis factor-alpha (TNF-alpha) has been shown to enhance the cytotoxicity of a variety of antineoplastic agents. To examine whether multidrug-resistant cells are targets of TNF-alpha, and whether TNF-alpha is capable of modulating chemoresistance of these cells, a pleural mesothelioma cell line (PXF1118L) and two multidrug-resistant sublines thereof were used as experimental models. Drug resistance of these cells was due to P-glycoprotein expression, as confirmed by (1) staining with a monoclonal antibody (MRK16) specific for human P-glycoprotein, (2) decreased accumulation of [3H]vinblastine that was reversed by verapamil, and (3) enhanced cytotoxicity of vindesine in the presence of verapamil. Parental and multidrug-resistant cells exhibited little but comparable sensitivity to TNF-alpha alone. Combining TNF-alpha with vindesine or, to a lesser extent, with doxorubicin, but not with cisplatin, resulted in greater cytotoxicity towards multidrug-resistant cells than seen for each compound alone, indicating a synergism. In contrast, TNF-alpha failed to modulate vindesine or doxorubicin cytotoxicity in parental cells. [3H]Vinblastine accumulation was unaffected by TNF-alpha, and chemoresistance was reduced by TNF-alpha also in the presence of verapamil (10 microM), indicating that TNF-alpha was acting in a way different from calcium-channel blockers. Though the molecular mechanism by which TNF-alpha was enhancing vindesine and doxorubicin cytotoxicity remained undefined in this study, the numbers of TNF-alpha binding sites on parental and on multidrug-resistant cells were similar, and P-glycoprotein expression was unmodulated during the entire 48 h incubation period. In conclusion, we show that TNF-alpha increases the cytotoxicity of anticancer drugs in multidrug-resistant tumor cells by a mechanism that differs from most chemosensitizing agents, including verapamil. Further studies will be needed to clarify the mechanism by which TNF-alpha synergizes with anticancer drugs.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1995

Submitogenic concentrations of anti-CD3 monoclonal antibody exert antiapoptotic effects in preactivated CD4+ but not CD8+ human T cells.

Immunological memory has been ascribed to the presence of long-lived memory cells. The mechanisms underlying their generation are not completely understood, but dependence on antigen persistence has been discussed in this regard. However, in spite of in vivo evidence favoring this model, studies on TCR/CD3 stimulation of T cell lines or unseparated peripheral blood T cell in vitro have failed to demonstrate prolonged survival of preactivated cells. We have examined the dose-dependent effect of TCR/CD3 engagement mimicked by immobilized anti-CD3 antibody. To this end, well-defined populations of CD4+ and CD8+ lymphoblasts isolated from bulk cultures of preactivated PBMCs by flow sorting were examined. These cells were restimulated with immobilized anti-CD3 in the presence or absence of various costimulatory factors, and were analyzed for their viability state, as well as their apoptotic and proliferative behavior. We have shown that inhibition of apoptosis following CD3 stimulation occurs at submitogenic concentrations, while activation-driven apoptosis requires high-density TCR/CD3 activation. Prevention of apoptosis by submitogenic CD3 stimulation was, however, observed only when CD4+ but not when CD8+ cells were investigated, and was not readily influenced by other costimulatory factors present in cultures. This observation points to the importance of antigen persistence in regulating survival of memory CD4+ but not CD8+ cells.

Antibodies, Monoclonal↗