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Biomedical subjects

M Müller

Publications and source records attributed to M Müller.

At least 739 records · Page 41Linked to original sources
PubMed1996

Redistribution of ribosomal DNA after blocking of transcription induced by actinomycin D.

We report on the effect of different doses and times of incubation of the cytostatic drug actinomycin D (AMD) on nucleolar morphology, rRNA gene transcription and rDNA gene localization using in situ hybridization and the immunocytochemical detection of the human upstream binding factor (UBF) at the electron microscopic level in HeLa cells. Low doses of AMD (0.001 micrograms/ml, 30 min) selectively block rRNA gene transcription but alter neither nucleolar morphology nor the localization of rDNA with respect to the nucleolar components. Treatment with high doses of AMD (0.05 micrograms/ml, 1 h) resulted in a retraction of the rDNA out of the nucleolus in addition to the well-known blocking of rDNA transcription, total nuclear transcription and nucleolar segregation. Under these conditions accumulations of rDNA were found in patches of chromatin at the nucleolar periphery. We conclude that the blocking of rRNA gene transcription and the changes in nucleolar morphology, both induced by AMD at different doses, are independent phenomena.

Cell Nucleolus↗
PubMed1996

Detection of poly(ADP-ribose) polymerase and its reaction product poly(ADP-ribose) by immunocytochemistry.

Poly(ADP-ribose) polymerase catalyses the formation of ADP-ribose polymers covalently attached to various nuclear proteins, using NAD+ as substrate. The activity of this enzyme is strongly stimulated upon binding to DNA single or double strand breaks. Poly(ADP-ribosyl)ation is an immediate cellular response to DNA damage and is thought to be involved in DNA repair, genetic recombination, apoptosis and other processes during which DNA strand breaks are formed. In recent years we and others have established cell culture systems with altered poly(ADP-ribose) polymerase activity. Here we describe immunocytochemistry protocols based on the use of antibodies against the DNA-binding domain of human poly(ADP-ribose) polymerase and against its reaction product poly(ADP-ribose). These protocols allow for the convenient mass screening of cell transfectants with overexpression of poly(ADP-ribose) polymerase or of a dominant-negative mutant for this enzyme, i.e. the DNA-binding domain. In addition, the immunocytochemical detection of poly(ADP-ribose) allows screening for cells with altered enzyme activity.

Animals↗
PubMed1996

What can 18S rDNA do for bivalve phylogeny?

Molecular characteristics, especially 18S rDNA sequences, may be of great value for the study of bivalve evolution and its numerous morphological convergencies once the reliability of these data can be evaluated. The analysis of 11 published complete molluscan sequences and two new ones, Arca noae and Atrina pectinata, reveals considerable differences in relative substitution rates. The gastropod and eulamellibranch species have the fastest and Atrina species have the slowest rates. Two methods are used to assess the information contents of the dataset in addition to bootstrap analysis, spectral analysis, and the "pattern of resolved nodes" technique. Tree reconstructions by parsimony, neighbor-joining, and maximum-likelihood differ in regard to the position of the eulamellibranch family Mactridae and of Crassostrea. Although there is a signal for the monophyly of Bivalvia, Mactridae cluster with Gastropoda in most runs, rendering Bivalvia diphyletic. The position of Crassostrea was extremely variable, probably due to the high substitution rate of this species. Atrina roots deeper than Arca in all trees, although a corresponding signal in spectral analysis is absent. Phylogenetic signals among the three pectinid species are low but sufficient to resolve the branching pattern. The tree inferred from the 18S rDNA and from morphological data has Bivalvia monophyletic with a basal polytomy of Mactridae, Crassostrea, and the remaining Pteriomorphia, where Arca branches off before Atrina and the Pectinidae. Argopecten is sister group to the other two pectinids; 18S sequence data will have great impact on our understanding of bivalve phylogeny, but only when more sequences of similar substitution rates are available.

Animals↗
PubMed1996

Upregulation of gap junction protein connexin43 in alveolar epithelial cells of rats with radiation-induced pulmonary fibrosis.

The degree of immunoreactive connexin43 (C x 43) in rat lung was evaluated during the development of radiation-induced pulmonary fibrosis in rat by a double immunofluorescence technique using polyclonal antisera to Cx43 and monoclonal antibodies to cytokeratins on cryostat sections. In normal rat lungs, Cx43 was detected in pneumocytes type II and I, in large blood vessel endothelia, in peribronchial smooth muscle cells, and in some peribronchial and perivascular interstitial cells. As early as 1 week after irradiation, enhanced immunoreactivity for Cx43 in the epithelial cells was detected. In severely injured lungs (about 3 months after irradiation), Cx43 was found also in the cytoplasm of type II pneumocytes. These findings were confirmed by western blot data. Western blot analysis also revealed increased phosphorylation of Cx43. It remains to be investigated whether the increased content of Cx43 in irradiated rat lung may be due to an enhanced number of gap junctions between type I and II alveolar epithelial cells.

Actins↗
PubMed1996

[Cost aspects in anesthesia. Propofol versus isoflurane anesthesia].

UNLABELLED: Cost control is no longer an option, but a necessity. Propofol anaesthesia is expensive, however, the true differences in comparison to volatile anaesthetics (isoflurane) are not known. METHODS: Sixty patients undergoing either thyroidectomy (n = 30) or laparoscopic cholecystectomy (n = 30) were randomly divided into 3 groups of 20 patients. In group I propofol and fentanyl were used for anaesthesia, in group II isoflurane ('standard' isoflurane anaesthesia), and in group III isoflurane using a low-flow system (fresh gas flow 2 l/min) was given. All patients were ventilated using 70% N2O in oxygen. Vecuronium was used in all cases for muscle relaxation. Isoflurane consumption was measured by weighing the isoflurane vaporizer. RESULTS: Biometric data and time of administration of the anaesthetic were similar in the three groups. Propofol patients stayed significantly shorter than isoflurane patients in the postanaesthesia care unit (PACU). Costs of additional drugs (antiemetics, analgesics) in the PACU were least in the propofol patients. Costs were without differences between the propofol (78.30 DM/patient) and 'standard' isoflurane groups (81.69 DM/patient). Patients in group III showed the lowest overall costs (57.46 DM/patient) (P < 0.05). CONCLUSION: A climate of cost-consciousness and cost-containment prevails at the present time. The costs of propofol and 'standard' isoflurane anaesthesia were without differences; however, isoflurane used in a low-flow system had the lowest cost in this study. Doubts are justified, however, as to whether the choice of anaesthetic agents may considerably lower the costs of an anaesthesia department.

Adult↗
PubMed1996

[Is the elimination of osteosarcoma cells with intraoperative "mesh autotransfusion" and leukocyte depletion filters possible?].

UNLABELLED: Intraoperative autotransfusion is contraindicated in cancer surgery because of the possible risk of systemic tumor spread. The aim of the present study was to investigate whether a cell saver in combination with a white blood cell depletion filter can remove osteosarcoma cells. METHODS: A defined number of osteosarcoma cells from an established cell line were added to red cell concentrates and Ringer solution. The tumor cell concentration was 1000/ ml in the first five experiments, 7111/ml in test no. 6, 1667/ml in test no. 7 and 167/ml in test no. 8. Following thorough mixing, each unit was processed separately by a cell saver (DIDECO BT 795/P) in its normal operation mode to produce a red cell concentrate. This red cell concentrate was filtered using a leukocyte depletion filter (PALL BPF 4). Samples were taken before and after processing with the autotransfuser and after filtration with the white cell depletion filter. Cytospin specimens from all samples were examined for osteosarcoma cells by three different methods (Papanicolaou stain, Vimentin antibodies, DNA analysis). RESULTS: After processing with the autotransfuser, tumor cells were identified in the red cell concentrate. No osteosarcoma cells were evident after the combined use of cell saver and leukocyte depletion filter. CONCLUSION: The sole use of the autotransfuser DIDECO BT 795/P during osteosarcoma surgery is not recommended because of the potential danger of retransfusion of malignant cells. In combination with the leukocyte depletion filter PALL BPF 4, no osteosarcoma cells were identified in the red cell concentrate. Since the adhesiveness of tumor cells from established cell lines may be different from that of tumor cells in the intraoperative salvaged blood, further studies with blood from the surgical field are necessary to determine the efficacy of white cell depletion filters to eliminate osteosarcoma cells.

Blood Transfusion, Autologous↗
PubMed1996

State-dependent modulation of mSlo, a cloned calcium-dependent potassium channel.

The mouse slopoke calcium-dependent potassium channel (mSlo) has been expressed heterologously in COS cells, and incorporated from COS cell membranes into artificial phospholipid bilayers. Under control conditions, the channel is not modulated by ATP. However, when mSlo is treated first with the calcium-dependent potassium channel opener NS004, which itself increases the open probability of the channel, subsequent addition of ATP causes a large further increase in channel open probability. An increase in channel activity is not by itself sufficient to confer sensitivity to ATP, because ATP does not modulate channels whose open probability has been increased by elevated calcium or depolarized voltage. The ATP analog AMP-PNP has only minimal effects on channel activity after treatment with NS004, suggesting that hydrolysis of the ATP is required for its action on mSlo. A peptide inhibitor of the calcium/calmodulin-dependent protein kinase II (CaMKII) blocks the modulation of mSlo by ATP, whereas peptide inhibitors of other serine/threonine protein kinases are without effect. The results are consistent with a state-dependent modulation of mSlo by ATP, possibly via phosphorylation.

Adenosine Triphosphate↗
PubMed1996

The E7 protein of human papillomavirus (HPV) type 16 expressed by recombinant vaccinia virus can be used for detection of antibodies in sera from cervical cancer patients.

Sera from 128 Mexican cervical cancer patients (age 30-80; mean 53.6) and from 47 healthy women (age 25-69; mean 49.2) were investigated using a newly developed assay for the detection of serum antibodies to the human papillomavirus (HPV) type 16 early protein E7. This test (CIPA), based upon immunoprecipitation followed by Western blot analysis, uses the complete E7 protein expressed in HeLa cells infected with recombinant vaccinia virus. To determine the sensitivity and specificity of this assay, these results were compared with previous results of the same sera tested by enzyme-linked immunosorbent assay (ELISA; using synthetic peptides derived from HPV 16 E7) and radio-immunoprecipitation (RIPA) using in vitro translated HPV 16 E7 protein. CIPA (45% positives) demonstrated a significant increase in detection rate compared to the peptide-ELISA (30% positives; P = 0.014, chi2-test) and only a slight increase compared to RIPA (38% positives; P = 0.204, chi2-test). Based on the testing of sera from patients with HPV 16 DNA positive tumors the specificity and sensitivity of the CIPA were 0.98 and 0.59, respectively.

Adult↗
PubMed1996

Sequence of a Giardia lamblia gene coding for the glycolytic enzyme, pyruvate,phosphate dikinase.

The sequence of the gene coding for pyruvate,phosphate dikinase (EC 2.7.9.1) in Giardia lamblia (syn. G. duodenalis) has been established. The deduced amino acid sequence is very similar to all of its known homologs from the protist, Entamoeba histolytica, the eubacterium, Clostridium symbiosum and plant chloroplasts. Phylogenetic reconstruction with neighbor-joining and maximum parsimony methods reveals that the sequences form two clades, one comprising the anaerobic microorganisms, and the other the chloroplast enzymes.

Amino Acid Sequence↗
PubMed1996

Transcranial Doppler sonography at the early stage of acute central nervous system infections in adults.

Transcranial Doppler sonography (TCD) of the middle, anterior and posterior cerebral arteries and of the basilar artery was used to evaluate the mean blood velocity (V mean) and the pulsatility index [PI = (V systolic-V diastolic)/V mean] as a vascular resistance index in 63 patients (male 40, female 23, mean age 43 +/- 19 y) with bacterial meningitis (n = 33, including 2 patients with fungal meningitis) and viral meningitis (n = 30) within 12 h after admission of the patients. The findings were similar for all intracranial arteries. Compared with reference values of 69 healthy volunteers [V mean of middle cerebral artery [MCA] 57 +/- 13 cm/s, MCA-PI 0.83 +/- 0.15], MCA-V mean was increased in patients with Glasgow coma scale (GCS) scores of 14 and 15 (71 +/- 18 cm/s; t-test: p < 0.001), not significantly different in the patients with GCS scores of 10-13 (55 +/- 21 cm/s) and decreased in those with GCS scores of 3-9 (42 +/- 21 cm/s, p < 0.01). The MCA-PI increased from 0.93 +/- 0.22 in the patients with GCS scores of 14-15 to 2.81 +/- 2.06 in those with GCS scores of 3-9 (p < 0.001 vs. controls). By regression analysis, MCA-V mean decreased and MCA-PI increased with decreasing GCS scores (p < 0.001). Only in patients with bacterial meningitis was the Glasgow outcome scale (GOS) score lower the more the MCA-PI was increased (regression analysis p < 0.001). We conclude that in patients with bacterial and viral meningitis, and in a good clinical state, the cerebral blood flow seems increased by hyperemia; with clinical deterioration the cerebral haemodynamics worsen. However, the early assessment of the cerebral blood flow by TCD seems useful for predicting outcome in bacterial meningitis only.

Acute Disease↗
PubMed1996

Spine loss in experimental epilepsy: quantitative light and electron microscopic analysis of intracellularly stained CA3 pyramidal cells in hippocampal slice cultures.

The sequence of neuronal alterations resulting from epileptic activity is poorly understood. In the hippocampus of some epileptic patients, there is a loss of certain neuronal types in the hilar region and in CA3. The neuronal alterations preceding this degeneration probably affect synaptic structures. Here we have estimated the number of dendritic spines, major postsynaptic elements of hippocampal neurons, in defined dendritic segments of identified (intracellularly stained) CA3 pyramidal neurons in "epileptic" slice cultures of hippocampus and in control cultures. Slice cultures were prepared from five- or six-day-old rat pups and maintained in vivo for 23 days before epileptic activity was induced by application of the convulsants bicuculline and picrotoxin for three days. Individual CA3 pyramidal neurons were then intracellularly injected with horseradish peroxidase, and the number of dendritic spines was counted in proximodistal dendritic segments by applying the Sholl method. In addition, the total dendritic length was measured and the branching index evaluated. The number of spines on CA3 pyramidal cell dendrites in the "epileptic" cultures was found to be decreased by 40%. This spine loss affected proximal and peripheral dendritic segments of the CA3 pyramidal neurons to a similar extent. No significant differences were observed between control and "epileptic" cultures in dendritic length or in the branching index. Quantitative electron microscopic analysis did not reveal differences between "epileptic" cultures and control cultures in the spine area of the labelled CA3 pyramidal cells, indicating that there was a real spine loss, not just a reduction in the size of the spines. We conclude that epileptic activity causes morphological alterations in defined postsynaptic compartments of hippocampal pyramidal cells surviving under these conditions.

Animals↗
PubMed1996

The cochlear place-frequency map of the adult and developing Mongolian gerbil.

Gerbils are frequently used in auditory research; however, only limited information on the exact frequency representation in the cochlea is available. Therefore the place-frequency map of the gerbil was determined by iontophoretic application of horseradish peroxidase in physiologically characterized single auditory nerve fibers. Locations of the labeled nerve fibers at the inner hair cells were determined from a '3-dimensional' reconstruction of the cochleae. The map was established for frequencies between 0.3 and 31.9 kHz. As in other mammals, a baso-apical frequency gradient from high to low frequencies was found. The slope of the place-frequency map amounted to 1.5 mm/octave for higher frequencies. Below 4 kHz the slope decreased to a value of 1 mm/octave at 0.5 kHz. A shift in the frequency map occurs during cochlear maturation. The place-frequency map in subadult gerbils (18 days after birth) is shifted by a distance corresponding to an octave, at least for frequencies above 6 kHz.

Acoustic Stimulation↗
PubMed1996

JAKs and STATs branch out.

The JAK-STAT signal-transduction pathway is utilized by a wide range of cytokines to regulate gene expression. Cytokines activate members of the JAK family o f protein tyrosine kinases, which in turn activate, by tyrosine phosphorylation, one or more STAT transcription factor family members. Activated STATs form dimers, translocate to the nucleus and bind to response elements to induce transcription. Recent findings are beginning to connect JAKs and STATs with other signalling pathways: JAKs may phosphorylate and activate signalling proteins other than STATs, and STATs can be phosphorylated by non-]AK tyrosine kinases. STAT activity can also be modulated by serine phosphorylation.

Journal Article↗
PubMed1996

Cyclosporine-induced detachment of vascular endothelial cells initiates the intrinsic coagulation system in plasma and whole blood.

Cyclosporine A (CsA) is supposed to alter the metabolism of vascular endothelial cells, leading to a prothrombotic state. We examined by which mechanism human umbilical vein endothelial cells (HUVECs) treated with CsA would promote coagulation in human plasma and in whole blood. Treatment of HUVECs with CsA at concentrations clinically used led to dose-dependent cell detachment, with the subsequent exposure of the highly procoagulant connective tissue. As determined by scanning electron microscopy, cell counting of detached and adherent cells, and antigenic measurement of collagen exposure, HUVECs treated with 0.4 micrograms/ml CsA (or more) for 4 days exhibited significant amounts of subendothelial areas. On CsA-treated HUVEC monolayers, the clotting time of recalcified citrated platelet-rich plasma (PRP), but not platelet-poor plasma (PPP), was dose-dependently shortened. Likewise, the onset of thrombin generation was significantly earlier. Except at a high concentration of 8.0 micrograms/ml CsA, there was no procoagulant effect when PPP was used. To investigate CsA-treated HUVECs in whole blood, cells were cultivated on globular microcarriers and were incubated with nonanticoagulated whole blood. When untreated cells were used, generation of factor Xa, thrombin, and kallikrein was completely suppressed for 30 minutes. HUVEC beads treated with 0.4 and 0.8 micrograms/ml CsA, however, led to a dose-dependent generation of all three coagulation factors, with peak values at 2.5 to 5 minutes. Extrinsic activation was excluded, since CsA treatment did not induce tissue factor activity in HUVECs. Furthermore, the thrombomodulin activity of HUVECs w as not altered by CsA. In conclusion, treatment of HUVECs with CsA for 4 days at concentrations clinically used leads to the exposure of subendothelial areas that induce activation of the intrinsic coagulation in recalcified PRP and nonanticoagulated whole blood.

Blood Coagulation↗
PubMed1996

The multidrug transporters--proteins of an ancient immune system.

The multidrug resistance proteins, discovered as membrane transporters producing chemotherapy-resistance in cancer, are functioning as complex cellular defence systems through recognition and energy-dependent removal of a large variety of toxic agents. The multidrug transporters belong to the ATP-binding cassette (ABC) transporters, present both in prokaryotes and eukaryotes and built from a combination of characteristic membrane-spanning helices and cytoplasmic ATP-binding domains. In mammals the MDR1 (P-glycoprotein) extrudes large hydrophobic compounds and provides the basis of the blood-brain and the blood-testis barrier for such molecules. The multidrug resistance-associated protein (MRP) and its homologues have a major role in the cellular export of large organic anions, including e.g. conjugated bile salts and glutathione-conjugates. The substrate recognition, that is the self and non-self discrimination and the ATP-dependent foreign agent extrusion are directly coupled within the structure of these large plasma membrane proteins. Here we suggest that the multidrug transporters are essential parts of our immune-defence system, working as 'cellular antitoxic' mechanisms.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1996

Multispecific amphipathic substrate transport by an organic anion transporter of human liver.

BACKGROUND: Hepatic uptake of differently charged amphipathic endo- and xenobiotics is thought to occur via distinct carrier-mediated transport systems. Alternatively, a single rat organic anion transporting polypeptide (oatp) has recently been demonstrated to mediate hepatocellular uptake of differently charged amphipathic substrates. AIM: To investigate whether a cloned human liver organic anion transporting polypeptide (OATP) also can mediate charge- and class-independent hepatocellular uptake of amphipathic substrates. METHODS: Xenopus laevis oocytes were injected with OATP-cRNA. Sodium-independent uptake of estrone-3-sulfate, ouabain and the organic cation N-(4,4-azo-n-pentyl)-21-ajmalinium was compared in OATP-expressing and uninjected (or water injected) control oocytes. RESULTS: Our results indicate that OATP, in addition to bromosulfophthalein and bile salts, can also transport anionic estrone-3-sulfate (Km approximately 59 microM), neutral ouabain (K(m) approximately 5.5 mM) and cationic N-(4,4-azo-n-pentyl)-21-ajmalinium. For each of these compounds, OATP-mediated uptake was cis-inhibited by the OATP substrate taurochenodeoxycholate and the transport activities correlated well with the amounts of cRNA injected. CONCLUSION: Similar to the rat liver oatp, the human liver OATP can also mediate multispecific and charge-independent uptake of lipophilic amphipathic organic compounds. Thus, OATP may play an important role in the first pass clearance of drugs and other xenobiotics by the human liver.

Animals↗
PubMed1996

Distribution of von Willebrand factor in capillary endothelial cells of rat lungs with pulmonary fibrosis.

To determine the value of von Willebrand factor (vWF) antigen as a marker of endothelial injury in radiation-induced fibrosis of rat lungs, we studied endothelial immunoreactivity to antibodies against vWF using the indirect immunoperoxidase technique combined with morphometric analysis. Using immunoelectron microscopy of LR White embedded lung samples to detect vWF, immunogold-labelled Weibel Palade bodies were found in endothelial cells of capillary endothelium. The irradiated lungs showed a statistically significant elevation of vWF expression, ie, vWF positive endothelia per unit area, and a significant increase of vWF expression per unit of parenchyma as well. The results suggest that vWF antigen expression and the number of vWF positive structures is modulated under condition of injury in radiation-induced fibrosis.

Animals↗