Showing papers by "Ludwig Maximilian University of Munich published in 1993"

H2O2 and antioxidants have opposite effects on activation of NF-kappa B and AP-1 in intact cells: AP-1 as secondary antioxidant-responsive factor.

Abstract: We show that AP-1 is an antioxidant-responsive transcription factor. DNA binding and transactivation by AP-1 were induced in HeLa cells upon treatment with the antioxidants pyrrolidine dithiocarbamate (PDTC) and N-acetyl-L-cysteine (NAC), and upon transient expression of the antioxidative enzyme thioredoxin. While PDTC and NAC enhanced DNA binding and transactivation of AP-1 in response to phorbol ester, the oxidant H2O2 suppressed phorbol ester activation of the factor. H2O2 on its own was only a weak inducer of AP-1. Activation of AP-1 by PDTC was dependent on protein synthesis and involved transcriptional induction of c-jun and c-fos genes. Transcriptional activation of c-fos by PDTC was conferred by the serum response element, suggesting that serum response factor and associated proteins function as primary antioxidant-responsive transcription factors. In the same cell line, the oxidative stress-responsive transcription factor NF-kappa B behaved in a manner strikingly opposite to AP-1. DNA binding and transactivation by NF-kappa B were strongly activated by H2O2, while the antioxidants alone were ineffective. H2O2 potentiated the activation of NF-kappa B by phorbol ester, while PDTC and NAC suppressed PMA activation of the factor. PDTC did not influence protein kinase C (PKC) activity and PKC activation by PMA, indicating that the antioxidant acted downstream of and independently from PKC.

Alterations of K+ currents in isolated human ventricular myocytes from patients with terminal heart failure.

Abstract: Prolongation of the action potential has been postulated to be a major reason for the altered diastolic relaxation of the heart in patients with severe heart failure. To investigate the electrophysiological basis for this action potential prolongation in terminal heart failure, K+ currents were recorded in single ventricular myocytes isolated from 16 explanted hearts of patients undergoing transplantation. Results from diseased hearts were compared with ventricular cells isolated from six undiseased donor hearts. Action potential duration was significantly prolonged in cells from patients with heart failure. A delayed rectifier K+ current was hardly detectable in most cells, and if it could be recorded, it was very small in both diseased and undiseased cells. When currents were normalized for cell surface area, the average current density of the inward rectifier K+ current was significantly reduced in diseased cells when compared with normal control cells (hyperpolarization at -100 mV, -15.9 +/- 2.2 vs -9.0 +/- 1.2 microA/cm2; P < .01). In addition, a large transient outward K+ current could be recorded in human myocytes. The average current density of the time-dependent component of this transient outward K+ current was significantly reduced in heart failure (depolarization at +40 mV, 9.1 +/- 1.0 vs 5.8 +/- 0.64 microA/cm2; P < .01). Action potential prolongation in severe heart failure may partially be explained by a reduction in current densities of the inward rectifier K+ current and of the transient outward K+ current. These alterations may thereby have a significant effect on cardiac relaxation.

A Comparison of Directional Atherectomy with Coronary Angioplasty in Patients with Coronary Artery Disease

Abstract: Background Directional coronary atherectomy is a new technique of coronary revascularization by which atherosclerotic plaque is excised and retrieved from target lesions. With respect to the rate of restenosis and clinical outcomes, it is not known how this procedure compares with balloon angioplasty, which relies on dilation of the plaque and vessel wall. We compared the rate of restenosis after angioplasty with that after atherectomy. Methods At 35 sites in the United States and Europe, 1012 patients were randomly assigned to either atherectomy (512 patients) or angioplasty (500 patients). The patients underwent coronary angiography at base line and again after six months; the paired angiograms were quantitatively assessed at one laboratory by investigators unaware of the treatment assignments. Results Stenosis was reduced to 50 percent or less more often with atherectomy than with angioplasty (89 percent vs. 80 percent, P<0.001), and there was a greater immediate increase in vessel caliber (1.05 vs. 0....

Efficient processing of spatial joins using R-trees

Abstract: Spatial joins are one of the most important operations for combining spatial objects of several relations. The efficient processing of a spatial join is extremely important since its execution time is superlinear in the number of spatial objects of the participating relations, and this number of objects may be very high. In this paper, we present a first detailed study of spatial join processing using R-trees, particularly R*-trees. R-trees are very suitable for supporting spatial queries and the R*-tree is one of the most efficient members of the R-tree family. Starting from a straightforward approach, we present several techniques for improving its execution time with respect to both, CPU- and I/O-time. Eventually, we end up with an algorithm whose total execution time is improved over the first approach by an order of magnitude. Using a buffer of reasonable size, I/O-time is almost optimal, i.e. it almost corresponds to the time for reading each required page of the relations exactly once. The performance of the various approaches is investigated in an experimental performance comparison where several large data sets from real applications are used.

Water-Soluble Ligands, Metal Complexes, and Catalysts: Synergism of Homogeneous and Heterogeneous Catalysis

Abstract: Rapid developments in the field of catalysis are leading to an increased demand for tailor-made catalysts. Water-soluble complex catalysts, which are being intensively investigated at the present time, combine the advantages of homogeneous and heterogeneous catalysis: simple and complete separation of the product from the catalyst, high activity, and high selectivity. From the large number of available water-soluble ligands, the appropriate catalysts can be developed for many reactions. The industrial applications in the fields of hydrogenation and hydroformylation have already indicated the wide scope of this type of catalyst. In addition, the annual production of 300 000 tons of butyraldehyde through application of water-soluble rhodium complexes at Hoechst AG in Oberhausen, Germany, has demonstrated the industrial importance of the concept of complex-catalyzed reactions in aqueous two-phase systems. The efficient operation of catalytic processes increasingly requires the loss-free recycling of the noble metal catalyst, and this can be simply and economically realized in two-phase systems. Special applications in biochemical problems open up developments in the field of water-soluble transition metal complexes that far transcend the familiar kinds of homogeneous catalysis. In the near future, the investigation and application of metal complex catalysts that are compatible with the physiological, cheap, and environmentally friendly solvent, water, is likely to become a high priority in catalysis research.

Peripheral mechanisms of opioid analgesia.

Abstract: Traditionally, opioids exert analgesic effects through actions within the central nervous system (CNS) exclusively (1) Recently, however, evidence has begun to accumulate that opioid antinociception can be initiated by activation of opioid receptors located outside the CNS One of the earliest repo

Pathogenesis of Graves' ophthalmopathy.

Abstract: Ophthalmopathy, the most frequent extrathyroidal manifestation of Graves' disease, results from an increased volume of the orbital tissues (connective and adipose tissue, interstitial enlargement of extraocular muscles) within the enclosed space of the bony orbits. While the primary antigen is still uncertain, the thyrotropin receptor has recently become a prime candidate because it is expressed both by thyroid epithelial cells and by orbital preadipocyte fibroblasts. Tolerance towards the thyroidal and orbital TSH receptor may be disrupted by bacterial or viral antigens through a \"molecular mimicry\"-type mechanism. In the context of appropriate costimulatory signals, this would prompt antigen presentation by local dendritic cells and macrophages followed by the recruitment of TSH receptor-specific T cells to the orbital space. As a result, orbital T cells, fibroblasts, adipocytes and perhaps other residential cells would release numerous cytokines, growth factors and inflammatory mediators, many of which act as potent stimulators of glycosaminoglycan accumulation and edema formation. Once initiated, the orbital immune process frequently takes on a momentum of its own, leading to non-specific but nonetheless harmful consequences such as tissue hypoxia, oxygen free radical damage and fibrogenic tissue remodeling. The clinical signs and symptoms of GO reflect the mechanical consequences of increased orbital tissue volume and pressure within the confines of the bony orbits.

Developing and evaluating cross-cultural instruments from minimum requirements to optimal models

Abstract: In the age of increased international collaboration in medical research, the necessity of having at hand cross-culturally applicable instruments for the assessment of health-related quality of life (HRQL) in clinical trials has been voiced. Several important theoretical bases leading to cultural bias in HRQL measurement include differences in definitions of HRQL across national and cultural contexts, levels of observation relied upon to indicate HRQL states, and the significance or weight placed upon the various HRQL states or dimensions measured. Despite a growing literature on the development and evaluation of existing HRQL measures in other cultures, comprehensive sets of procedures or requirements for the international part of development and evaluation are lacking. This paper reviews major approaches to developing international HRQL measures, and discusses various methods and criteria that have been recommended for evaluating measurement equivalence in comparisons of research across national and cultural contexts. A summary of recent trends and advances in international HRQL assessment is presented.

DNA extraction from Pleistocene bones by a silica-based purification method

Abstract: The polymerase chain reaction has made it possible to include extinct species and past populations in molecular studies of phylogeny and evolution (1). This emerging field, however, is marred by problems, mainly because archaeological remains often yield no amplifiable DNA, extracts often contain components which inhibit the Tag. polymerase, and contamination of trace amounts of contemporary DNA can yield misleading results (2, 3). We have found that the following method, which is a modification of a protocol published by Boom et a/. (4), is highly useful in alleviating the former two types of problems and in several cases allows the study of late Pleistocene animal remains that often are not amenable to other extraction procedures. A layer of approximately 1 mm is removed from the surface of the bone samples by grinding with a drilling machine in order to reduce contamination from previous handling. The sample is ground to a fine powder under liquid nitrogen in a freezer mill (Spex Industries Inc., Edison, NJ). About 0.5 g of bone powder is added to 1 ml of extraction buffer consisting of 10 M guanidinium thiocyanate (GuSCN), 0.1 M Tris-HCl pH 6.4, 0.02 M EDTA pH 8.0 and 1.3% Triton X-100. This is then incubated at 60°C for one to several hours with sporadic agitation. After centrifugation for 5 min at 5,000 rpm about 500 /tl of the supernatant is recovered and added to a mixture of 500 /tl of extraction buffer and 40 /il silica suspension prepared as in ref. 4. The mixture is incubated for 10 min at room temperature. Subsequently, the silica pellet is washed twice with a buffer consisting of 10 M GuSCN and 0.1 M Tris-HCl, pH 6.4, twice with 70% ethanol and once with acetone. After drying the pellet at 56°C, nucleic acids are eluted at 56°C in two aliquots of 65 /tl water or TE and stored at -20°C.

Molecular dynamics simulation of a bilayer of 200 lipids in the gel and in the liquid crystal phase

Abstract: We have constructed and simulated a membrane-water system which consists of 200 molecules of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphatidylcholine forming a rectangular patch of a bilayer and of 5483 water molecules covering the head groups on each side of the bilayer. The total number of atoms is approximately 27 000. The lateral dimensions of the bilayer are 85 A×100 A, and the distance between the bilayer surfaces as given by the average phosphorus to phosphorus distance is 35 A. The thickness of each water layer is up to 15 A. In all, we simulated 263 ps of the dynamics of the system. To prevent system disintegration, atoms within 5 A from the surface were harmonically restrained and treated by Langevin dynamics, forming a stochastic boundary

E-Cadherin Expression in Primary and Metastatic Gastric Cancer: Down-Regulation Correlates with Cellular Dedifferentiation and Glandular Disintegration

Abstract: Expression of the epithelial cell adhesion molecule E-cadherin in primary and metastatic gastric carcinoma was examined using immunohistochemical analyses. Compared to normal mucosa, 92% of the primary tumors (n = 60) showed reduced E-cadherin expression, suggesting that down-regulation of this cell adhesion molecule is a common early event in gastric tumorigenesis. No significant correlation was found between E-cadherin expression and tumor diameter, lymphatic vessel invasion, Borrmann classification, lymph node status, or manifest metastases. Although advanced tumors (tumor stage 3/4) showed a loss of E-cadherin-positive cells (< or = 50% cells/lesion, P = 0.0168), the most significant correlation was observed between low E-cadherin expression and cellular dedifferentiation (grading 3/4, P = 0.0001) and disintegration of tissue architecture (Lauren and WHO classifications, P = 0.0001). Low E-cadherin expression (< or = 50% cells/lesion) was associated with tumor recurrence (P = 0.0013) and mortality (P = 0.0246). E-cadherin expression in metastatic lesions (n = 58) also correlated with the degree of glandular differentiation (P = 0.0001). Significant correlation (rs = 0.686) was observed between E-cadherin expression in primary and metastatic lesions from individual patients (n = 39). However, while metastases derived from E-cadherin-negative tumors remained negative, those originating from E-cadherin-positive tumors frequently demonstrated increased levels of expression. Evaluation of multiple metastases in 11 patients revealed uniformly strong E-cadherin expression in liver metastases, suggesting a possible regulatory role of the microenvironment.

The novel subset of CD14+/CD16+ blood monocytes exhibits features of tissue macrophages

Abstract: The CD14+/CD16+ cells account for about 10% of all blood monocytes. They are characterized by a low level expression of the CD14 molecule and a high level expression of the CD16 (Fc gamma R III) molecule. Polymerase chain reaction analysis of mRNA prevalence in CD14+/CD16+ cells (compared to the regular CD14++ blood monocytes) demonstrates low levels of CD14 transcripts and high levels of CD16 transcripts, suggestive of a transcriptional control for both of these proteins. Analysis of additional cell surface molecules in three-color immunofluorescence reveals that CD14+/CD16+ cells express the Fc gamma R II in all, and Fc gamma R I and ICAM-1 in some donors. Furthermore, class II antigens are expressed at fourfold higher levels, while both, CD11b and CD33 cell surface proteins, are decreased by a factor of two. Transcript levels were reduced in CD14+/CD16+ cells for all three cell surface molecules. Since these phenotypic markers of the CD14+/CD16+ blood monocytes are reminiscent of tissue macrophages, we performed a comparative analysis with alveolar macrophages (AM). These cells are similar to the CD14+/CD16+ monocytes in that they show low levels of CD14 and strong expression of CD16. Furthermore, similar to the CD14+/CD16+ cells, the AM also exhibit higher levels of class II and lower levels of CD11b and CD33 when compared to the regular CD14++ blood monocytes. In vitro induction of maturation of blood monocytes by 5 day culture of peripheral blood mononuclear cells in 10% human serum will result in decreased CD14 and increased CD16 cell surface expression on the monocyte derived macrophages. At the same time, these cells acquire increased levels of class II and decreased levels of CD11b and CD33. Taken together, these data show that CD14+/CD16+ monocytes, while still in circulation, have acquired features in common with mature tissue macrophages.

Neural Networks in Chemistry

Abstract: The capabilities of the human brain have always fascinated scientists and led them to investigate its inner workings. Over the past 50 years a number of models have been developed which have attempted to replicate the brain’s various functions. At the same time the development of computers was taking a totally different direction. As a result, today’s computer architectures, operating systems, and programming have very little in common with information processing as performed by the brain. Currently we are experiencing a reevaluation of the brain’s abilities, and models of information processing in the brain have been translated into algorithms and made widely available. The basic building-block of these brain models (neural networks) is an information processing unit that is a model of a neuron. An artificial neuron of this kind performs only rather simple mathematical operations; its effectiveness is derived solely from the way in which large numbers of neurons may be connected to form a network. Just as the various neural models replicate different abilities of the brain, they can be used to solve different types of problem: the classification of objects, the modeling of functional relationships, the storage and retrieval of information, and the representation of large amounts of data. This potential suggests many possibilities for the processing of chemical data, and already applications cover a wide area: spectroscopic analysis, prediction of reactions, chemical process control, and the analysis of electrostatic potentials. All these are just a small sample of the great many possibilities.

Social structure of pilot whales revealed by analytical DNA profiling

Abstract: Long-finned pilot whales swim in large, extremely cohesive social groups known as pods. Molecular typing revealed that pod members form a single extended family. Mature males neither disperse from nor mate within their natal pods, a situation unusual for mammals. Such behavior could be explained in terms of inclusive fitness benefits gained by adult males helping the large number of female relatives with which they swim.

Urokinase (uPA) and its inhibitor PAI-1 are strong and independent prognostic factors in node-negative breast cancer

Abstract: Evidence has accumulated that invasion and metastasis in solid tumors require the action of tumor-associated proteases, which promote the dissolution of the surrounding tumor matrix and the basement membranes. The serine protease urokinase-type plasminogen activator (uPA), which is elevated in solid tumors, appears to play a key role in these processes. We used enzyme-linked immunoassays (ELISA) to test for uPA antigen and its inhibitor PAI-1 in tumor tissue extracts of 247 breast cancer patients who were enrolled in a prospective study. The relation of these data to known prognostic factors and to other variables such as DNA analysis and cathepsin D was studied. Disease-free and overall survival were analyzed according to Cox's proportional hazard model. The major new finding is that breast cancer patients with either high uPA (> 2.97 ng/mg protein) or high content of the uPA inhibitor PAI-1 (> 2.18 ng/mg protein) in their primary tumors have an increased risk of relapse and death. Multivariate analyses revealed uPA to be an independent and strong prognostic factor. The impact of uPA is as high as that of the lymph node status. In node-negative patients the impact of uPA is closely followed by that of PAI-1. Since uPA and PAI-1 are independent prognostic factors, the node-negative patients could be subdivided further by combining these two variables. In this refined analysis, patients whose primary tumors have lower levels of both antigens evidently have a very low risk of relapse (93% disease-free survival at three years) in contrast to patients with high uPA and high PAI-1 (55% disease-free survival at three years). The combination of uPA and PAI-1 in our group of patients with axillary node-negative breast cancer allows us to identify the 45 percent of patients having an increased risk of relapse. Consequently, more than half of the patients had less than a 10% probability of relapse and thus would possibly be candidates for being spared the necessity of adjuvant therapy.

An OspA serotyping system for Borrelia burgdorferi based on reactivity with monoclonal antibodies and OspA sequence analysis.

Abstract: A total of 136 Borrelia burgdorferi sensu latu strains from various biological sources (ticks, human skin, and cerebrospinal fluid) and geographical sources (Europe and North America) were investigated by Western blot (immunoblot) with eight monoclonal antibodies against different epitopes of the outer surface protein A (OspA). On the basis of the differential reactivities of these monoclonal antibodies, seven OspA serotypes were defined. As determined by 16S rRNA sequence analysis, these serotypes correlated well with recently delineated genospecies: serotype 1 corresponds to B. burgdorferi sensu strictu, serotype 2 corresponds to group VS461, and serotypes 3 to 7 correspond to Borrelia garinii sp. nov. (G. Baranton, D. Postic, I. Saint Girons, P. Boerlin, J.-C. Piffaretti, M. Assous, and P. A. D. Grimont, Int. J. Syst. Bacteriol. 42:378-383, 1992). Antigenic differences were confirmed by partial sequence analysis of OspA of representatives of each serotype. Comparative sequence analysis suggested that serotype 5 OspA resulted from genetic recombination of serotype 4 and 6 ospA genes. Serotype 2 (group VS461) was most prevalent among European skin isolates (49 of 62 isolates). Among all B. garinii strains included in this study, serotype 6 was most frequently found in ticks and only rarely in human skin and cerebrospinal fluid, whereas serotypes 4 and 5 were isolated from patients but never from ticks. Our data suggest different pathogenic potentials and organotropisms of distinct OspA serotypes and raise the question of true antigenic variation among B. garinii strains.

Signaling in the elicitation process is mediated through the octadecanoid pathway leading to jasmonic acid.

Abstract: Fungal cell walls and fragments thereof (elicitors) induce the formation of low and high molecular weight defense compounds in plant cell suspension cultures. This induced synthesis requires a signal molecule transmitting the message between the elicitor plant cell wall receptor and gene activation. We demonstrate in this study that cis-jasmonic acid is rapidly synthesized in plant cell cultures of diverse taxonomic origin (gymnosperms and mono- and dicotyledonous plants) after challenge with a fungal elicitor preparation. The rapid decline of cis-jasmonic acid in some of these tissues is attributed to rapid metabolism of this pentacyclic acid. The induction of alkaloids by several different molecules provoking the elicitation process is strictly correlated with the synthesis of jasmonates. Elicitation leads to a rapid release of alpha-linolenic acid from the lipid pool of the plant cell. alpha-Linolenic acid and 12-oxophytodienoic acid, the formation of which is also induced, are known to be distant precursors of jasmonic acid. We assume cis-jasmonic acid and its precursors to be the signaling molecules in the elicitation process.

Dorsal skinfold chamber technique for intravital microscopy in nude mice.

Abstract: For many years, observation chambers, implanted in various animal species and in man, have been used for intravital microscopy of the microcirculation in granulation tissue, preformed tissue, or of the microvascularization of tissue implants. We describe herein a skinfold chamber model for the intravital microscopic investigation of striated skin muscle in immunoincompetent, nude mice over a minimum period of 2 weeks. Using fluorescent markers for contrast enhancement of plasma (fluorescein isothiocyanate-dextran) and leukocytes (acridine orange), the presented model allows the quantitative analysis of 1) the microhemodynamic parameters microvessel diameter and red blood cell velocity in arterioles (16 to 50 mu diameter), capillaries (4 to 9 mu diameter), and post-capillary venules (19 to 60 mu diameter), 2) leukocyte/endothelium interaction in these vessel segments, 3) functional capillary density and intercapillary distance, and 4) endothelial cell integrity. These parameters can be assessed in the microcirculation of the striated muscle tissue under normal or pathological conditions, as well as in the microcirculation of transplanted xenogeneic (human) neoplastic and non-neoplastic tissue grafts.

Nuclear uptake control of NF-kappa B by MAD-3, an I kappa B protein present in the nucleus.

Abstract: I kappa B proteins specifically inhibit the DNA binding of NF-kappa B/Rel transcription factors. An additional role as inhibitors of nuclear uptake was supposed by subcellular fractionation and enucleation experiments. Using indirect immunofluorescence labeling of cells, we show here that the DNA-binding p50 and p65 subunits of NF-kappa B, as well as the I kappa B protein MAD-3, all occur in the nucleus when overexpressed on their own. Nuclear uptake of p65 and, to a lesser extent of p50, was, however, suppressed when MAD-3 was coexpressed. Likewise, nuclear uptake of MAD-3 was blocked by overexpressed p65 or p50. This directly demonstrates that I kappa B is a nuclear uptake regulatory protein and that the various subunits of NF-kappa B can mutually control their access to the nucleus. In the presence of MAD-3, antibodies specific for peptides overlapping the nuclear location signal (NLS) sequences of p65 and p50 could not recognize their epitopes on NF-kappa B, suggesting that the I kappa B protein rendered the signals inaccessible for NLS receptors.

Skeletal muscle sonography: a correlative study of echogenicity and morphology.

Abstract: In skeletal muscle sonography high echogenicities have proved to be of diagnostic value. The following study examines whether these echointensities are caused mainly by interstitial fat or fibrosis. Consequently, the echogenicities of 86 muscles, their diameters, and the thickness of subcutaneous fat layers superficial to these muscles were measured and compared for content of fat and connective tissue, which were assessed by morphometry and biochemical testing in the corresponding muscle biopsy samples. The results indicate that fat replacement constitutes the main cause of increased muscle echogenicity, whereas intramuscular fibrosis did not significantly affect the muscles' echogenicity.

Motor recovery following capsular stroke. Role of descending pathways from multiple motor areas

Abstract: The functional anatomy of motor recovery was studied by assessing motor function quantitatively in 23 patients following capsular or striatocapsular stroke While selective basal ganglia lesions (caudate and/or putamen exclusively) did not affect voluntary movements of the extremities, lesions of the anterior (plus caudate/putamen) or posterior limb of the internal capsule led to an initially severe motor impairment followed by excellent recovery, hand function included In contrast, lesions of the posterior limb of the internal capsule in combination with damage to lateral thalamus compromised motor outcome In experimental tracing of the topography of the internal capsule in macaque monkeys, we found axons of primary motor cortex passing through the middle third of the posterior limb of the internal capsule Axons of premotor cortex (dorsolateral and post-arcuate area 6) passed through the capsular genu, and those of supplementary motor area (mesial area 6) through the anterior limb Small capsular lesion can therefore disrupt the output of functionally and anatomically distinct motor areas selectively The clinically similar motor deficits with a similar course of functional restitution following disruption of these different descending motor pathways indicate a parallel operation of cortical motor areas They may have the further capability of substituting each other functionally in the process of recovery from hemiparesis