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Biomedical subjects

H Wolf

Publications and source records attributed to H Wolf.

At least 289 records · Page 16Linked to original sources
PubMed1994

Regression of deep vein thrombosis by i.v.-administration of a low molecular weight heparin--results of a pilot study.

Twenty-five patients with phlebographically confirmed deep vein thrombosis of the lower limb were treated with intravenous infusions of low molecular weight heparin for 7 to 29 days. The mean dosage was 15.2 +/- 3.0 Uanti-Xa (equivalent 7.6 +/- 1.5 U-aPTT). Phlebograms were taken before, during and after the treatment with low molecular weight heparin and evaluated using the score system of Marder. Nearly complete recanalization of the occluded veins was found in six (24%) patients, improvement of the Marder score of 60 to 90% was found in four patients and of 30 to 60% in seven patients, while eight patients remained unchanged. With an average dose of 15.2 I.U./kg/h the heptest was prolonged to 70 to 120 seconds while the aPTT-level did not significantly increase. tPA-antigen-levels increased significantly in most of the patients after the third day of treatment, while PAI-activity remained unchanged. A positive conclusion between the decrease of the Marder score and the duration of treatment was found. Thus the low molecular weight heparin used in this investigation proved to be effective and safe in treating deep vein thrombosis.

Adult↗
PubMed1994

A novel and sensitive method for the detection of secreted cell products using time-resolved fluorescence.

A new test has been developed for the quantitative detection of products secreted from isolated cells, based on the use of lanthanide- rather than enzyme-linked streptavidin. Used as a label, europium (Eu3+) can be measured with high sensitivity by time-resolved fluorescence. The main advantages of this assay are both an increased sensitivity and measuring range of cell released substances, when compared to the standard "wet" ELISA. Thus, the immunoglobulin secretion rate of 10(5) splenocytes could be easily measured by time-resolved fluoroimmunoassay (TR-FIA), while it remained below the detection limit of the 'wet' ELISA. In contrast to the classical ELISPOT test, this assay does not detect single antibody secreting cells (ASC), but would be useful for precise quantification of secreted cell products, such as immunoglobulins, cytokines, growth factors.

Animals↗
PubMed1994

Identification of wild-type and mutant p53 peptides binding to HLA-A2 assessed by a peptide loading-deficient cell line assay and a novel major histocompatibility complex class I peptide binding assay.

Mutations of the p53 gene are the most frequently observed genetic changes in human cancers; often leading to an overexpression of the wild-type (wt) p53 protein. Demonstrable T cell reactivity against tumor cells overexpressing wt or mutant p53-derived peptides could support the application of such epitopes in cancer immunotherapies. As the binding of peptide to MHC class I molecules is a prerequisite for antigen-specific T cell recognition, we evaluated the ability of wt and mutant p53 peptides to bind to HLA-A2.1 using two independent flow cytometry-based assay systems, the T2 major histocompatibility complex (MHC) class I peptide stabilization assay (stabilization assay) and the peptide-induced MHC class I reconstitution assay (reconstitution assay). The twenty selected wt sequences each conformed to the previously reported HLA-A2.1 peptide binding motif. Seven of the wt p53 and 2/13 mutant p53 peptides derived from the previously chosen wt peptides bound to HLA-A2.1 in both the stabilization and the reconstitution assays. An additional six wt and six mutant p53 peptides, presumably exhibiting lower affinity for HLA-A2.1, were identified only in the reconstitution assay. Those p53 peptides binding HLA-A2.1 may provide useful immunogens for the generation of HLA-A2.1-restricted cytolytic T lymphocytes in vitro and in vivo.

Amino Acid Sequence↗
PubMed1994

Induction of anergy in resting human T lymphocytes by immobilized anti-CD3 antibodies.

How the T cell receptor (TcR)/CD3 complex mediates not only the induction of T cell activation but also suppressive effects like T cell anergy or apoptosis is not well understood. Here we describe a series of preincubation and restimulation experiments which demonstrate that primary stimulation of resting, unseparated human T cells with mitogenic doses of immobilized anti-CD3 antibodies induces hyporesponsiveness upon restimulation of the cells. Various costimuli can prevent this type of anergy to a variable degree if present during the preincubation period, phorbol 12-myristate 13-acetate (PMA) being the most and anti-CD4 antibody the least effective. If employed together with anti-CD3 antibody during the restimulation phase of the assay, interleukin (IL)-2, IL-4 and anti-CD28 antibody break anergy almost completely. Proliferation induced by a submitogenic dose of anti-CD3 antibody supplemented by costimulatory signals (anti-CD2, anti-CD4, anti-CD28, IL-2, IL-4 or PMA) does not result in hyporesponsiveness. Taken together, these results support a modified view of the two-signal model for T cell activation according to which anergy induction in resting T cells occurs if primary proliferation is induced by high density triggering of the TcR/CD3 complex in the absence of accessory signals. We discuss possible implications of these findings for the induction of peripheral tolerance.

Adult↗
PubMed1994

Origin and clonal relationship of common inhibitory motoneurons CI1 and CI3 in the locust CNS.

In the primordial thoracic ganglia of locust embryos, the bromodeoxyuridine (BrdU) technique for labelling proliferating cells and their progeny was combined with intracellular dye injection to investigate the origin and the clonal relationship of common inhibitory motoneurons. Common inhibitors 1 (CI1) and 3 (CI3) were found to be siblings, that is, they are produced by the division of one ganglion mother cell. This ganglion mother cell results from the first division of neuroblast 5-5, at about 30% of embryonic development. A large portion, at least, of the ganglion mother cells produced by subsequent divisions of neuroblast 5-5 give rise to interneurons with contralaterally ascending or descending axons and GABA-like immunoreactivity. Thus, CI1 and CI3 are more closely related to putative inhibitory interneurons than they are to other, that is, excitatory, motoneurons. Consistent with this, the CI somata are associated with cell bodies of putative inhibitory interneurons rather than with clusters of excitatory motoneuron somata. These results elicit speculations regarding the evolutionary origin of inhibitory motoneurons.

Animals↗
PubMed1994

Identification of active cytomegalovirus infection by analysis of immediate-early, early and late transcripts in peripheral blood cells of immunodeficient patients.

Owing to the persistence of viral DNA in leukocytes after primary CMV infection, detection of CMV DNA in these cells does not necessarily represent active infection. To identify CMV replication more precisely we have analysed immediate-early, early and late CMV transcripts by RNA amplification. The assay seems to be specific for active infection since no RNA-derived PCR products were obtained from healthy seropositive persons. The late UL83 transcript was detected in 80% of the patients with active CMV infections. Diagnosis of CMV replication by amplification of early and immediate-early transcripts was considerably less sensitive. In the case of continual CMV DNA detection in blood leukocytes by PCR without pp65 antigenemia the analysis of defined CMV transcripts would allow differentiation of active and non-active infection. In two cases the RNA assay became negative prior to DNA PCR analysis and pp65 antigen detection upon antiviral treatment, indicating that RNA amplification could be a suitable assay for early detection of the end of viral replication. No strong correlation was found between RNA detection and appearance of clinical symptoms. The development of CMV disease probably depends more on the extent of the functional impairment of the immune system.

AIDS-Related Opportunistic Infections↗
PubMed1994

Assembly and extracellular release of chimeric HIV-1 Pr55gag retrovirus-like particles.

The HIV-1 Pr55gag precursors were previously shown to assemble and bud from a variety of different cell types as noninfectious virus-like particles (VLPs) resembling immature HIV virions. The use of these VLPs as an immunogenic and autologous carrier component may allow the presentation of defined epitopes deduced from reading frames other than gag to the immune system, thereby avoiding the induction of adverse immune responses. In order to identify domains within Pr55gag that can be replaced by immunologically relevant epitopes without affecting its capacity to assemble into VLPs, we deleted three domains of a predicted high surface probability. Deletion of amino acids 211-241 within p24CA and amino acids 436-471 within the p6LI portion of Pr55gag had no effect on the assembly, ultrastructure, biophysical properties, and yields of mutant VLPs when expressed via recombinant vaccinia viruses in mammalian cells. Deletion of amino acids 99-154 overlapping the p17MA/p24CA cleavage site completely abolished the capacity of the gag polyprotein to form VLPs and led to a reduction of immature Pr55 VLPs released into the cell-culture supernatants when coexpressed with wild-type Pr55gag. In contrast, assembly and budding of chimeric VLPs could be demonstrated after replacing amino acids 211-241 and 436-471 by immunologically relevant epitopes derived from reading frames other than Pr55gag (e.g., V3 loop; CD4-binding-domain; nef-CTL epitope) or after fusion of these sequences to the carboxy terminus of Pr55gag. The importance of these data for the development of novel HIV candidate vaccines is discussed.

Amino Acid Sequence↗
PubMed1994

Control by pulse parameters of electric field-mediated gene transfer in mammalian cells.

Electric field-mediated gene transfer in mammalian cells (electrotransformation) depends on the pulsing conditions (field intensity, pulse duration, number of pulses). The effect of these parameters was systematically investigated using the transient expression of the chloramphenicol acetyltransferase and the beta-galactosidase activities in Chinese hamster ovary cells. Pulsing conditions inducing reversible permeabilization of the cell plasma membrane are not sufficient to induce gene transfer. The plasmid must be present during the electric pulse if it is to be transferred across the membrane into the cytoplasm. Only the localized part of the cell membrane brought to the permeabilized state by the external field is competent. Pulse duration plays a key role in the magnitude of the transfer. The field induces a complex reaction between the membrane and the plasmid that is accumulated at the cell interface by electrophoretic forces. This leads to an insertion of the plasmid, which can then cross the membrane.

Animals↗
PubMed1994

Low-molecular-weight heparin.

Low molecular weight heparins (LMWHs) are mixtures of heparin molecules in the range of 3000 to 10,000 daltons. As LMWHs of various manufacturers are all produced differently, they are not comparable to each other and are therefore considered to be individual products with different pharmacologic and clinical properties. All these agents have some common characteristics, however, such as a higher availability after subcutaneous administration and a longer biologic half-life. Numerous clinical trials have demonstrated that LMWHs are highly effective and safe for postsurgical prophylaxis of deep vein thrombosis (DVT); therefore, the LMWHs that are commercially available so far, are mainly approved in this indication. LMWHs are also effective in the prophylaxis of DVT in medical indications, as well as for the treatment of established DVT.

Drug Combinations↗
PubMed1994

Inhibition of infectious human immunodeficiency virus type 1 particle formation by Gag protein-derived peptides.

Sequential overlapping Gag protein-derived oligopeptides of human immunodeficiency virus type 1 (HIV-1) 22 to 24 amino acids long, were synthesized and tested in vitro for antiviral activity. Two synthetic peptides, one derived from the matrix protein p17 (NPGLLETSEGCRQ, amino acids 47 to 59) and one located in the capsid protein p24 (PAATLEEMMTA, amino acids 339 to 349) inhibited the production of infectious virus when added to HIV-1-infected cultures when used in the range of 20 to 200 micrograms/ml. As shown by thin section electron microscopy, peptide treatment resulted in the release of immature, deformed virus particles suggesting that the two peptides interfered with assembly and maturation. Other Gag protein-derived oligopeptides had little or no influence on virus production. To characterize further the functionally active regions we synthesized peptide derivatives with three consecutive amino acids substituted by alanine; they did not cause inhibition. Therefore the regions responsible for inhibition were located between amino acids 50 to 61 in p17, and 342 to 350 in p24. These observations might lead to the development of a new antiviral strategy affecting the late stage of virus replication.

Amino Acid Sequence↗
PubMed1994

Negatively cis-acting elements in the distal part of the promoter of Epstein-Barr virus trans-activator gene BZLF1.

Epstein-Barr virus (EBV) replicates in a latent or a lytic way in the infected organism, depending on the type and level of differentiation of the host cell. The switch between latency and lytic replication was previously shown, for Burkitt's lymphoma cell lines, to depend on the viral BZLF1 gene product. Protein-DNA assays were used to identify the cis-acting elements that represent the link between regulating signal transduction pathways and the viral cascade of gene expression. Specific binding of proteins to several sites of the BZLF1 promoter during latency was shown. Induction of the lytic cycle by stimulation with 12-O-tetradecanoyl-phorbol 13-acetate abolished the binding of these proteins to the distal promoter (positions -227 to -551), suggesting a functional role for the down-regulation of promoter activity during latency. Computer analysis identified a multiply repeated sequence motif, HI, in this region and exonuclease III footprints confirmed that these sites act as specific protein recognition sites. Using a set of reporter plasmids we were able to demonstrate a negative regulatory effect of the HI motif in some B lymphoid cell lines, in contrast to epithelial HeLa cells. The HI silencer elements are different from other silencer elements described so far in respect of their sequence and protein-binding pattern during the activation of BZLF1.

Base Sequence↗
PubMed1994

Immune response in the lungs following oral immunization with bacterial lysates of respiratory pathogens.

We have investigated the local immune response of the BALB/c mouse respiratory tract after oral immunization with a bacterial lysate of seven common respiratory pathogens. After two immunization on five consecutive days, we examined the immunoglobulin (immunoglobulin G [IgG], IgM, and IgA) secretion rates of cells isolated from the lungs and compared them with those of spleen cells of orally immunized and nonimmunized animals by using a new test system based on time-resolved fluorescence. The procedure followed the principle of the classical ELISPOT test with nitrocellulose-bottomed microtiter plates, but europium (Eu3+)-linked streptavidin rather than enzyme-conjugated streptavidin was used, with the advantage of quantifying secreted immunoglobulins instead of detecting single antibody-secreting cells. Lymphocytes isolated from the lungs of treated animals revealed significant increases in total and antigen-specific IgA synthesis compared with the rates of the controls, whereas IgG and IgM production rates showed no remarkable differences. In addition, the sera of treated mice revealed higher antigen-specific IgA titers but not increased IgM and IgG levels. We conclude that priming the gut-associated lymphoid tissue with bacterial antigens of pneumotropic microorganisms can elicit an enhanced IgA response in a distant mucosal effector site, such as the respiratory tract, according to the concept of a common mucosa-associated immune system.

Adjuvants, Immunologic↗
PubMed1994

Different activation of Epstein-Barr virus immediate-early and early genes in Burkitt lymphoma cells and lymphoblastoid cell lines.

Specific expression of the Epstein-Barr virus (EBV) immediate-early and early gene products Zta, Rta, I'ta, and MSta by a recombinant vaccinia virus system allowed us to analyze the first steps in the induction of the lytic cycle in EBV-infected Burkitt lymphoma (BL) cells and lymphoblastoid cell lines (LCLs). Significant differences in the induction of early genes were found between these cell types: whereas in BL cells the trans activator Zta was found to induce key steps of the early lytic cycle, only minor activities of Zta were noted in LCLs. Contrary to Zta, the trans activator Rta was found to be highly effective in LCLs. These observations suggest that Rta may play an important role in the activation of the early lytic cycle in LCLs, although it cannot be activated by Zta. The latter may be a reason for the lower tendency of LCLs to switch into the lytic cycle compared with BL cells or differentiated epithelial cells.

B-Lymphocytes↗
PubMed1994

Rhythmic modulation of the responsiveness of locust sensory local interneurons by walking pattern generating networks.

1. Spiking local interneurons in the thoracic ganglia of the locust are known to process mechanosensory information from leg receptors and to contribute to postural reflexes. It is not clear, however, whether these interneurons are also involved in the control of leg movement during walking or whether their role in mediating local reflexes is phasically modulated by the walking pattern generating networks. To address these questions, intracellular recordings from spiking local interneurons in a midline population were made both in a "fictive walking" preparation reduced to isolated thoracic ganglia and in semi-intact locusts walking on a treadwheel. 2. In fictive walking (and therefore deafferented) preparations, rhythmic synaptic inputs were observed in 72% of the sampled local interneurons. These inputs resulted in a modulation of the interneurons' firing rate in phase with the centrally generated rhythm. A similar rhythmic drive was seen in spiking local interneurons recorded from semi-intact walking locusts. This drive persisted after amputation of the leg from which the interneurons received their sensory inputs. These results indicate that this population of interneurons receives inputs from central networks that participate in the generation of walking. 3. In fictive walking preparations where a hindleg was left attached to its segmental ganglion, individual local interneurons could be characterized physiologically by their receptive field properties. The central inputs were found to modulate the activity of members of all subclasses of interneurons (i.e., purely exteroceptive, purely proprioceptive, and with mixed receptive fields. The mode of modulation observed could oppose or assist the effect of sensory input expected to occur during normal walking.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗