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Biomedical subjects

M Müller

Publications and source records attributed to M Müller.

At least 451 records · Page 25Linked to original sources
PubMed1999

Single daily doses of saquinavir achieve HIV-inhibitory concentrations when combined with baby-dose ritonavir.

Dual protease inhibitor (PI) therapy has been established either in order to increase plasma concentrations of one PI or to combine synergistic effects of two PI's on viral load. Studies with saquinavir (SQV) and small doses of ritonavir (RTV) as well as experiences from our therapeutic drug monitoring suggest that single daily doses may result in sufficient SQV serum levels throughout an interval of 24 h. A controlled, randomized trial with 20 healthy men was conducted for the comparison of serum levels with 1600 mg SQV (group 1) or 1600 mg SQV/200 mg RTV (group 2). The dosages were selected in order to use RTV as an inhibitor of cytochrome P450 3A4 and SQV as protease inhibitor. The volunteers received single daily doses following a standardized breakfast on 3 consecutive days. Serum samples were analyzed for SQV and RTV employing LC-tandem mass spectrometry. The minimum concentration of saquinavir after 24 hours, the AUC, the maximum concentration and the serum half-lives on day 3 served as target parameters. The minimum SQV concentration amounted to 469.4 ng/ml, when combined with RTV and proved to be significantly higher (p <0.05) than the corresponding concentration with SQV alone (127.3 ng/ml). The SQV maximum concentration was raised approximately 6fold and the AUC 9fold when RTV was coadministered. In combination with 200 mg of RTV the predominant elimination half-life of SQV increased from 2.6 to 6.45 hours. These data prove that under single daily doses of 1600 mg SQV/200 mg RTV HIV-inhibitory concentrations of SQV can be achieved for 24 hours. Due to the high variability of the concentrations, which can be seen with all PI s, we recommend continuous therapeutic drug monitoring of serum trough levels.

Adult↗
PubMed1999

Rapid in vitro biocompatibility assay of endovascular stents by flow cytometry using platelet activation and platelet-leukocyte aggregation.

Clinical studies suggest that stent design and surface texture are responsible for differences in biocompatibility of metallic endovascular stents. A simple in vitro experimental setup was established to test stent-induced degree of platelet and leukocyte activation and platelet-leukocyte aggregation by flow cytometry. Heparin-coated tantalum stents and gold-coated and uncoated stainless steel stents were tested. Stents were implanted into silicone tubes and exposed to blood from healthy volunteers. Platelet and leukocyte activation and percentage of leukocyte-platelet aggregates were determined in a whole-blood assay by subsequent staining for activation-associated antigens (CD41a, CD42b, CD62p, and fibrinogen binding) and leukocyte antigens (CD14 and CD45) and flow cytometric analysis. Blood taken directly after venous puncture or exposed to the silicone tube alone was used as negative controls. Positive control was in vitro stimulation with thrombin receptor activating peptide (TRAP-6). Low degree of platelet activation and significant increase in monocyte- and neutrophil-platelet aggregation were observed in blood exposed to stents (P < 0.05). In addition, leukocyte activation was induced as measured by increased CD45 and CD14 expression. Heparin coated stents continuously induced less platelet activation and leukocyte-platelet aggregation than uncoated stainless steel stents of the same length and shorter stents of the same structure. Stent surface coating and texture plays a role in platelet and leukocyte activation and leukocyte-platelet aggregation. Using this simple in vitro assay and whole blood and flow cytometry, it seems possible to differentiate stents by their potency to activate platelets and/or leukocytes. This assay could be applied for improving the biocompatibility of coronary stents.

Adult↗
PubMed1999

Human nerve growth factor beta (hNGF-beta): mammary gland specific expression and production in transgenic rabbits.

Transgenic rabbits carrying gene constructs encoding human nerve growth factor beta (hNGF-beta) cDNA were generated. Expression of hNGF-beta mRNA was restricted to the mammary gland of lactating rabbits. Western Blot analysis revealed a polypeptide of 13.2 kDa in the milk of transgenic animals. hNGF-beta was purified from the milk by a two-step chromatographic procedure. Electrospray mass spectroscopy analysis of purified hNGF-beta depicted a molecular weight of 13,261 Da per subunit. The biological activity of the hNGF-beta was tested using PC12W2 cells and cultures of dorsal root ganglion neurons from chicken embryos. Crude defatted milk from transgenic animals and purified hNGF-beta demonstrated full biological activity when compared to commercial recombinant hNGF-beta.

Animals↗
PubMed1999

Lipid analysis by matrix-assisted laser desorption and ionization mass spectrometry: A methodological approach.

Whereas matrix-assisted laser desorption and ionization time-of-flight mass spectrometry (MALDI-TOF-MS) has gained high importance in the field of protein analysis, surprisingly few studies were published about the use of MALDI for lipid analysis. Lipids, however, are well-suited for MALDI since all experiments can be performed in a sole organic phase and, thus, extremely homogeneous matrix/analyte mixtures are formed. We report here for the first time the application of MALDI-TOF-MS for the analysis of diacylglycerols, phosphatidylcholines, and (poly)phosphoinositides. It is shown that in a matrix of 2,5-dihydroxybenzoic acid the molecular ions (M + 1) of phosphatidylcholines as well as the corresponding adducts of different phosphoinositides are easily detected even in complex mixtures, and thus, detailed data on the fatty acid composition are provided. In contrast, diacylglycerols are mainly detected as the corresponding sodium or potassium adducts, but not as the protonated forms. Fragmentation reactions of fatty acids on the double bonds and on the polar lipid head group are observed to a minor extent in the spectra of all investigated lipids. Generally, choline derivatives are most sensitive toward further fragmentation reactions. Due to its very high sensitivity (up to picomolar concentrations) MALDI-TOF-MS can be used for the direct investigation of biologically relevant lipid mixtures occurring, e.g. , in cell membranes. The analysis of the lipid composition of neutrophilic granulocytes is given as a representative example for future applications.

Diglycerides↗
PubMed1999

Sonographic diagnosis of abdominal wall relaxation.

PURPOSE: We report our experience using sonography to diagnose abdominal wall relaxation. METHODS: All patients with abdominal wall abnormalities from 1996 through 1997 underwent clinical and sonographic examination. Sonographic criteria for abdominal wall relaxation were continuity of the fascia (no gap), identification of all muscle layers, and decreased muscle thickness on the affected side. Fifty controls with no abdominal wall abnormalities were also examined for comparison. RESULTS: We sonographically diagnosed 5 cases of abdominal wall relaxation in 625 patients examined. The diagnosis in these 5 cases was confirmed by MRI. Muscle thickness on the affected side decreased by a mean of 38%+/-6% compared with muscle thickness on the unaffected side and was significantly different (p< 0.0001) from the mean muscle thickness seen in the controls. CONCLUSIONS: Sonography enabled the diagnosis of abdominal wall relaxation with the advantages of saving time and money and not requiring use of a contrast medium. Based on our results, we recommend sonography as the first-line imaging modality in the diagnosis of abdominal wall relaxation.

Abdominal Muscles↗
PubMed1999

Differential immunolocalization of VEGF in rat and human adult lung, and in experimental rat lung fibrosis: light, fluorescence, and electron microscopy.

Vascular endothelial growth factor (VEGF) is a cytokine with main angiogenetic functions in embryonic development and tumor-formation. In the adult lung, reports of the localization of VEGF were controversial. A precise cell typing of VEGF-positive pulmonary cells is still lacking. Nothing is known about a potential role in pulmonary fibrosis. Immunohistochemistry (IH), double immunofluorescence microscopy (DIF), and immunoelectron microscopy (IEM) were used to study the differential distribution of VEGF in paraffin-embedded (IH, DIF) and in cryo-substituted, Lowicryl-embedded (IEM) specimens of normal rat and human lungs and fibrotic rat lungs. Fibrosis was induced by intratracheal bleomycin treatment. IH and DIF showed that VEGF was present in surfactant protein (SP) D-positive alveolar type II pneumocytes, bronchiolar Clara cells, smooth muscle (SM) cells, and alpha-SM actin-positive myofibroblasts of normal rat and human lungs. Fibrotic lesions in bleomycin-treated rat lungs were rich in VEGF-positive cells presenting with a heterogeneous phenotype (mainly SP-D-positive type II pneumocytes, alpha-SM actin-positive myofibroblasts). There were no signs of angiogenesis. Post-embedding immunogold labeling using protein A-gold and IgG-gold technique revealed a specific localization of VEGF to mitochondria, Clara cell secretory granules, and capillary interendothelial cell junctions. The predominant localization of VEGF to bronchiolar and alveolar epithelial and alpha-SM actin-positive cells, and the marked increase of VEGF-positive type II pneumocytes and myofibroblasts in fibrotic lung lesions, indicate that in adult lungs VEGF is involved in processes other than angiogenesis.

Adult↗
PubMed1999

The genome of Rickettsia prowazekii and some thoughts on the origin of mitochondria and hydrogenosomes.

The sequence of an alpha-proteobacterial genome, that of Rickettsia prowazekii, is a substantial advance in microbial and evolutionary biology. The genome of this obligately aerobic intracellular parasite is small and is apparently still undergoing reduction, reflecting gene losses attributable to its intracellular parasitic lifestyle. Evolutionary analyses of proteins encoded in the genome contain the strongest phylogenetic evidence to date for the view that mitochondria descend from alpha-proteobacteria. Although both Rickettsia and mitochondrial genomes are highly reduced, it appears that genome reduction in these lineages has occurred independently. Rickettsia's genome encodes an ATP-generating machinery that is strikingly similar to that of aerobic mitochondria. But it does not encode homologues for the ATP-producing pathways of anaerobic mitochondria or hydrogenosomes, leaving an important issue regarding the origin and nature of the ancestral mitochondrial symbiont unresolved.

Animals↗
PubMed1999

Reduced antilisterial activity of TNF-deficient bone marrow-derived macrophages is due to impaired superoxide production.

Mice deficient for TNF ligand and receptor type 1 have demonstrated the importance of TNF in the host defense against Listeria monocytogenes. To investigate the particular deficiency of macrophages derived from TNF/lymphotoxin (LT)-alpha(-/-) mice in antilisterial growth control, bone marrow-derived macrophages (BMDM) were used for in vitro infection experiments. After the combined treatment with IFN-gamma and lipopolysaccharide (LPS), production of NO by wild-type (wt) and TNF/LT-alpha(-/-) BMDM was induced to comparable levels, but only wt BMDM controlled L. monocytogenes growth efficiently. Nevertheless, inhibition of NO production led to a remarkable loss of antilisterial activity. This suggests that presence of NO is necessary but not sufficient for L. monocytogenes killing and that elimination of L. monocytogenes requires additional effector molecules. The LPS-inducible superoxide production of TNF/LT-alpha(-/-) BMDM was impaired. Accordingly both scavenging of superoxide and peroxynitrite led to reduced L. monocytogenes killing by wt BMDM. In addition, peroxynitrite was able to kill L. monocytogenes in vitro. Together these findings suggest that the defective host defense of TNF/LT-alpha-deficient mice against L. monocytogenes partially stems from reduced superoxide production of macrophages due to the absence of TNF and imply a function for peroxynitrite, the reaction product of NO and superoxide, in the intracellular killing of L. monocytogenes.

Animals↗
PubMed1999

Proviral load determination of different feline immunodeficiency virus isolates using real-time polymerase chain reaction: influence of mismatches on quantification.

Lentiviruses are associated not only with immunodeficiency but also with malignancies. The mechanisms involved in tumorigenesis are still not fully understood. Cats infected with feline immunodeficiency virus (FIV) in the wild represent one model in which the role of viral load in the pathogenesis can be studied, since tumors, especially lymphomas, are quite often observed in cats infected with FIV. To be able to compare the viral load data among cats infected with different FIV isolates, the method used to obtain the viral load has to be unaffected by isolate-specific differences. This is especially true for the real-time polymerase chain reaction (PCR), a new method for viral load determination, since nucleotide sequence mismatches have been used for allelic discrimination with this method. To investigate the influence of these mismatches on PCR efficiency, we have used an FIV-specific real-time PCR and determined the influence of nucleotide sequence variation in several characterized FIV isolates as well as unknown isolates from naturally infected cats. We could demonstrate that minor mismatches, such as point mutations in the primer or the probe region, decrease overall PCR efficiency but do not abolish the quantification, in contrast to major mismatches of three or four nucleotides, which lead to complete inhibition of the real-time PCR detection. Based on these results, it will be possible to design real-time PCR systems allowing the quantification of a broad range of isolates, which is a prerequisite for the investigation of the impact of viral load in tumorigenesis.

Animals↗
PubMed1999

Improving protein identification from peptide mass fingerprinting through a parameterized multi-level scoring algorithm and an optimized peak detection.

We have developed a new algorithm to identify proteins by means of peptide mass fingerprinting. Starting from the matrix-assisted laser desorption/ionization-time-of-flight (MALDI-TOF) spectra and environmental data such as species, isoelectric point and molecular weight, as well as chemical modifications or number of missed cleavages of a protein, the program performs a fully automated identification of the protein. The first step is a peak detection algorithm, which allows precise and fast determination of peptide masses, even if the peaks are of low intensity or they overlap. In the second step the masses and environmental data are used by the identification algorithm to search in protein sequence databases (SWISS-PROT and/or TrEMBL) for protein entries that match the input data. Consequently, a list of candidate proteins is selected from the database, and a score calculation provides a ranking according to the quality of the match. To define the most discriminating scoring calculation we analyzed the respective role of each parameter in two directions. The first one is based on filtering and exploratory effects, while the second direction focuses on the levels where the parameters intervene in the identification process. Thus, according to our analysis, all input parameters contribute to the score, however with different weights. Since it is difficult to estimate the weights in advance, they have been computed with a generic algorithm, using a training set of 91 protein spectra with their environmental data. We tested the resulting scoring calculation on a test set of ten proteins and compared the identification results with those of other peptide mass fingerprinting programs.

Algorithms↗
PubMed1999

Detection of hepatitis C virus in paraffin-embedded liver biopsies of patients negative for viral RNA in serum.

The diagnosis of hepatitis C is based on serological testing for antibodies against various epitopes of the hepatitis C virus (HCV) and detection of HCV RNA in serum, because anti-HCV antibodies alone cannot discriminate patients who are infectious from those who have resolved the infection. If HCV RNA is not detected, which is the case in at least 20% of enzyme immunoassay (EIA)-positive patients, diagnosis remains unclear in a state of disease possibly well suited for therapeutic intervention. Therefore, we investigated if detection of HCV antigens or HCV RNA in routinely processed, formalin-fixed and paraffin-embedded (ffpe) liver biopsy specimens of patients positive for anti-HCV, but negative for HCV RNA in serum, could confirm diagnosis in this serological constellation. We detected HCV RNA by reverse-transcription polymerase chain reaction (RT-PCR) in 27 (61%) of 44 ffpe liver biopsies from EIA-positive, but HCV-RNA-seronegative, patients. Testing of 18 of these biopsies by a panel of polyclonal antibodies against structural and nonstructural HCV proteins revealed positive immunostaining in 6 cases (33%), which were also positive by RT-PCR. Most biopsies showed necroinflammation compatible with chronic hepatitis C, and the detection of tissue HCV RNA correlated significantly with a higher grade of inflammatory activity. Detectability of HCV RNA did not correlate with HCV subtype. In conclusion, the search for HCV RNA by RT-PCR within the liver biopsy specimen can establish rapid and unequivocal diagnosis of hepatitis C in at least 60% of anti-HCV antibody-positive patients who are seronegative for HCV RNA, and thus may help to avoid repeated testing and delayed therapy. Tissue RT-PCR may also be more efficient than serological testing for surveillance of interferon therapy response, because ongoing chronic active hepatitis C is clearly demonstrated in the absence of detectable serum HCV RNA.

DNA Primers↗
PubMed1999

Regulation of hepatic transport systems involved in bile secretion during liver regeneration in rats.

We investigated the expression of hepatic transport systems involved in bile secretion during liver regeneration after partial hepatectomy (PH) in rats. Initial studies showed maximal BrdU incorporation 24 hours after PH. Therefore, transporter expression and bile secretion were analyzed in detail at this time. The mRNA levels of the multidrug resistance genes mdr1a and mrp1 slightly increased, whereas mdr1b mRNA levels showed an extensive increase after PH. The mRNA levels of the conjugate transporter, mrp2, decreased slightly, whereas mrp2 protein levels did not change. Bilirubin secretion did not change, but the biliary glutathione secretion markedly decreased and the hepatic GSH content increased. The messenger RNA levels of the bile salt uptake transporters ntcp, oatp1, and oatp2 and the bile salt exporter, bsep/spgp, all decreased with ntcp showing the most prominent decrease. Protein levels of ntcp dramatically decreased whereas oatp2 only slightly decreased. Oatp1 protein expression slightly increased and bsep/spgp protein levels did not change. Decreased levels of bile salt uptake systems were associated with a 10-fold increase in the plasma bile salt concentration, yet, bile flow and bile salt secretion were increased when expressed per gram liver and unaffected when expressed on the basis of body weight. In conclusion, during the initial phase of rat liver regeneration ntcp is down-regulated whereas other transporter proteins involved in bile secretion are only slightly affected. Despite increased serum bile salt levels the remnant liver is not cholestatic: bile flow is maintained by uptake of bile salts probably via oatp isoforms and their secretion via bsep/spgp.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1999

Multidrug resistance protein MRP1, glutathione, and related enzymes. Their importance in acute myeloid leukemia.

Multidrug resistance (MDR), which is cross-resistance to structurally and functionally unrelated drugs such as anthracyclines, epipodophyllotoxins and vinca alkaloids, is a major cause of treatment failure in malignant disorders. Known mechanisms of MDR are overexpression of the ATP-dependent membrane proteins P-glycoprotein (P-gp) and multidrug resistance protein (MRP1), or an increased detoxification of compounds mediated by glutathione (GSH) or GSH related enzymes. MRP1 appeared to transport drugs conjugated to GSH and also unmodified cytostatic agents in presence of GSH. The relation between MRP1, GSH and enzymes involved in GSH metabolism or GSH dependent detoxification reactions recently has drawn a lot of attention. Coordinated induction of MRP1 and GSH related enzymes is reported in malignant cells after exposure to cytostatic agents. Besides MRP1, a number of MRP1 homologs are identified, named MRP2, MRP3, MRP4, MRP5 and MRP6. The relation between MDR and expression of these MRP1 homologs is currently under research.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1999

Irene, a case study of a bulimia nervosa patient: the therapeutic process of integrative painting therapy.

The bulimic symptomatology--supposedly an expression of certain developments in our society--increased immensely. The progress of Integrative Painting Therapy applied to a bulimia nervosa patient, is simultaneously visible and documented. This gives insight into the psychodynamics and psychopathology of this disorder, as well as its development during treatment. The painting group provides a quick access to the patient's emotions and it permits the establishment of a strong therapeutic relationship. Paintings generated within the group reflect a patient's inner experiences and provide a starting-point for the therapeutic process. In her pictures, Irene confronts her bulimia disorder and is finally able to come to terms with her social role as a woman, her growing up and her longing for a healthy family background especially meaning contact with her father. In this spirally evolving process, where painting groups, single and family therapy work together, she was finally able to break free from her bulimic symptoms and find a new access to herself and her family, especially her father.

Adult↗
PubMed1999

[Minimized polypropylene mesh for preperitoneal net plasty (PNP) of incisional hernias].

Repair of incisional hernias requires the extensive implantation of alloplastic materials. The extent of the scar tissue is markedly regulated by the amount and structure of the incorporated material and is responsible for the increased rate of local wound complications. Correspondingly, minimization of the alloplastic implants should be favorable. In a randomized, prospective clinical study, the early results were compared after implantation of either a minimized, low-weight (26.8 g/m2) mesh with a pore size of 5 mm or a common, heavy-weight (90.2 g/m2 polypropylene) mesh with a pore size of 0.8 mm. Indicators for clinical suitability were the rate and volume of seroma, subjective paraesthesia, physical capability, abdominal wall compliance, and the histologically analyzed tissue reaction of samples removed on the occasion of revision operations. As result, the optimized, low-weight mesh showed a remarkable decrease in the rate of seroma, patient complaints, less restriction of abdominal wall mobility, and improved abdominal wall compliance as verified by 3D stereography. These clinical findings corresponded to a pronounced decrease in inflammation and scar reaction, indicating improved incorporation of the alloplastic material. No other major complications except for one recurrence have been found.

Female↗
PubMed1999

Alterations in the immunohistochemical distribution patterns of vascular endothelial growth factor receptors Flk1 and Flt1 in bleomycin-induced rat lung fibrosis.

To investigate the role of vascular endothelial growth factor (VEGF) in fibrogenesis, the distribution patterns of the VEGF receptors Flt1 and Flk1 were studied by immunohistochemistry, double immunofluorescence, and immunoelectron microscopy in normal (n=2) and bleomycin-treated (n=21) adult rats. Lungs were studied at 5, 24, 28, 35, and 42 days after treatment (p.t.). Flt1, Flk1, and VEGF immunoreactivity localised predominantly to the pulmonary epithelium. In control lungs, Flt1 immunoreactivity was present in ciliated bronchial epithelium and type 2 pneumocytes, Flk1 in Clara cells, and VEGF in Clara cells and type 2 pneumocytes. Flk1 localised to mast cells, present in the peribronchovascular and pleural interstitium only. Flt1- and Flk1-mRNAs were observed in Clara cells and type 2 pneumocytes. Bleomycin-induced fibrogenesis was characterised by a decrease in Flk1 immunoreactivity of Clara cells, and an increase in VEGF-immunoreactive myofibroblasts and type 2 pneumocytes by day 5 p.t., followed by a progressive accumulation of Flk1-immunoreactive mast cells by day 24 p.t. in fibrotic lesions containing VEGF-immunoreactive myofibroblasts. After 42 days, fibrotic regions were densely populated by mast cells. Since mast cells are known to be chemotactically attracted by VEGF, we suggest that VEGF/Flk1 represents the molecular link between proliferation of myofibroblasts, accumulation of mast cells, and the burst of fibrosis at sites of initial lesions in bleomycin-induced fibrosis.

Animals↗
PubMed1999

Usefulness of clinical scores to predict outcome in bacterial meningitis.

The predictive usefulness of clinical scores in patients with acute bacterial meningitis was investigated. Fifty-one consecutive patients with acute bacterial meningitis were scored on days 1, 3, 5, 8, and 14 after admission according to the Sandinavian Stroke Scale (SSS), Glasgow Coma-Scale (GCS) and Hunt & Hess Scale (HH). As an index of their usefulness to predict the outcome, the scales were correlated with short-term outcome on day 21 assessed by the Glasgow Outcome Scale (GOS). The scores of all three scales correlated highly significantly with short-term outcome. Depending on the day of assessment, Spearman correlation coefficients ranged between 0.52 and 0.88 for SSS, between 0.50 and 0.84 for GCS, and between -0.47 and -0.82 for HH. The scales differed in their ability to predict outcome on and after day 1: mortality was best predicted by GCS, and complete recovery was best predicted by SSS. The use of scales in bacterial meningitis provides a rational quantitative basis to predict outcome more graduated than in dead or alive. Because the scales accentuate different aspects of outcome (e.g. mortality, restitution), the selection of a scale to be used in clinical trials should take into consideration the main focus of the study.

Acute Disease↗