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Biomedical subjects

A Schmid

Publications and source records attributed to A Schmid.

At least 163 records · Page 9Linked to original sources
PubMed1994

A nucleosome precludes binding of the transcription factor Pho4 in vivo to a critical target site in the PHO5 promoter.

Activation of the Saccharomyces cerevisiae PHO5 gene by phosphate starvation is accompanied by the disappearance of two pairs of positioned nucleosomes that flank a short hypersensitive region in the promoter. The transcription factor Pho4 is the key regulator of this transition. By in vitro footprinting it was previously shown that there is a low affinity site (UASp1) which is contained in the short hypersensitive region in the inactive promoter, and a high affinity site (UASp2) which is located in the adjacent nucleosome. To investigate the interplay between nucleosomes and Pho4, we have performed in vivo footprinting experiments with dimethylsulfate. Pho4 was found to bind to both sites in the active promoter. In contrast, it binds to neither site in the repressed promoter. Lack of binding under repressing conditions is largely due to the low affinity of Pho4 for its binding sites under these conditions. Despite the increased affinity of Pho4 for its target sites under activating conditions, binding to UASp2 is prevented by the presence of the nucleosome and can only occur after prior disruption of this nucleosome in a process that requires UASp1. Protection of the PHO5 UASp2 by the nucleosome is not absolute, however, since overexpression of Pho4 can disrupt this nucleosome even when UASp1 is deleted. Also under these conditions, with only UASp2 present, all four nucleosomes at the PHO5 promoter are disrupted, whereas no chromatin change at all is observed when both UAS elements are destroyed.

Allosteric Regulation↗
PubMed1994

Interaction of Clostridium botulinum C2 toxin with lipid bilayer membranes. Formation of cation-selective channels and inhibition of channel function by chloroquine.

Lipid bilayer experiments were performed with the C2-II binding component of the ADP-ribosylating C2 toxin from Clostridium botulinum. The trypsin-activated but not the nonactivated form of the protein was able to increase the specific conductance of artificial lipid bilayer membranes by the formation of ion-permeable channels. The channels had on average a single-channel conductance of 55 pS in 0.1 M KCl and were found to be cation-selective and voltage-dependent. The single-channel conductance was only moderately dependent on the bulk aqueous KCl concentration, which indicated point charge effects on the channel properties. Incubation of the activated C2-II binding component with antibodies against C2-II or with C2-I toxin inhibited channel formation to a large extent. Addition of chloroquine, a known inhibitor of endocytosis in cells, led to a dose-dependent decrease of the C2-II-induced membrane conductance. This result suggested that the activated C2-II component contains a binding site for chloroquine inside the channel. It is discussed that the channels formed by C2-II component are involved in the translocation of C2-I toxin across the target cell membrane.

Alkanes↗
PubMed1994

Generation and properties of measles virus mutations typically associated with subacute sclerosing panencephalitis.

Subacute sclerosing panencephalitis (SSPE), a very rare but lethal disease caused by measles viruses (MV) persisting in the human central nervous system (CNS) is characterized by lack of viral budding, reduced expression of the viral envelope proteins and spread of MV genomes through the CNS despite massive immune responses. The five major MV genes from several SSPE cases were cloned and sequenced, the two transmembrane envelope glycoproteins hemagglutinin (H) and fusion protein (F) were expressed and their maturation, cellular localization and functionality analyzed. We conclude that 1) mutations in the MV genes arise not only individually, by errors of the MV polymerase, but also in clusters as hypermutations, presumably due to RNA unwinding/modifying activity altering accidentally formed double-stranded RNA regions, 2) MVs spread in SSPE brains after clonal selection, 3) the MV matrix (M) gene is most heavily mutated and dispensable, 4) the two genes encoding envelope transmembrane proteins give rise to functional but altered proteins (typically F is heavily altered in its cytoplasmic domain), 5) H protein is transported poorly to the cell surface, 6) F and H proteins maintain tightly interdepending fusion functions, presumably to allow local cell fusion and MV ribonucleoprotein (RNP) spread through the CNS.

Amino Acid Sequence↗
PubMed1994

Isolation and characterisation of porin from the outer membrane of Synechococcus PCC 6301.

Pore-forming protein (porin) was isolated from N,N-dimethyl-dodecylaminoxid (LDAO)-extracted outer membranes of Synechococcus PCC 6301 and purified by ion exchange chromatography on DEAE-Sephacel column. The apparent molecular mass on SDS-PAGE was determined to be about 52,000. The native porin was reconstituted into black lipid bilayer membranes and showed a single-channel conductance of 5.5 nS in 1 M KCl. The porin was found to be N-terminally blocked. The C-terminal amino acid sequence was identified as Phe-Thr-Phe. Amino acid analysis suggested that the porin protein consists of about 420 amino acid residues, yielding a polarity of 43.6% and a molecular mass of 45,000 in contrast to the mobility on SDS-PAGE.

Amino Acid Sequence↗
PubMed1994

Dual-frequency method for synchronous measurement of cell capacitance, membrane conductance and access resistance on single cells.

A dual-frequency method was developed to monitor changes of membrane capacitance, membrane conductance and serial resistance between patch pipette and cytoplasm of the cell in the whole-cell configuration. Measurement of real and imaginary components of cell admittance during excitation with two superimposed sinusoidal voltage of different frequencies provides mathematical solutions for all three variables. The validity of the method was verified with experiments on mast cells and exocrine pancreatic acinar cells. During degranulation of mast cells, induced by GTP gamma S in the pipette solution, a stepwise increase in membrane capacitance could be observed, indicating that the resolution of the method is below 10 fF. Precalibration of the setup allows all calculated data to be expressed as absolute values. The capacitance measurement proved to be rather independent of changes in the access resistance and in the cell membrane resistance over a wide range. The huge changes in membrane conductance of mouse pancreatic acinar cells during hormonal stimulation with acetylcholine produced a relative error of less than 0.3% in the capacitance trace. This allows a clear distinction between changes of membrane conductance and cell capacitance. The method therefore offers great advantages in the study of exocytosis as well as endocytosis in cell types, such as exocrine gland cells, with major changes in membrane conductance during hormonal stimulation.

Acetylcholine↗
PubMed1994

Megakaryocyte growth in vitro predicts outcome in idiopathic thrombocytopenic purpura.

PURPOSE: The impact of megakaryocyte growth in vitro on clinical data, especially outcome, was studied in 25 consecutive children with idiopathic thrombocytopenic purpura (ITP). PATIENTS AND METHODS: Twenty children with untreated de novo ITP and five children with pretreated ITP were evaluated. The number of megakaryocyte colonies (cloning efficiency), the mean cell number per colony (mitotic amplification) and the percentages of polyploid megakaryocytes after 7 and 12 days in culture (relative size of the endomitotic compartment) were determined in two separate clonal assays. The culture data were related to clinical findings and outcome of the thrombocytopenia. RESULTS: The mean cell number per megakaryocyte colony was significantly correlated with the observed increase in the platelet count 5 days after starting therapy (n = 23; r = 0.642), and a significant negative correlation was found between the relative size of the endomitotic compartment and the duration of thrombocytopenia after bone marrow culture analysis (n = 25; r = -0.503). If all 25 children with ITP (untreated de novo and pretreated ITP) were considered, a normal frequency of polyploid megakaryocytes was associated with a duration of ITP for < 6 months in 14 of 16 cases, whereas an impaired polyploidization predicted a persistence of ITP for > 6 months in 9 of 9 cases (p < 0.0005); if only children with untreated de novo ITP (n = 20) were considered, 13 of 15 children with a normal polyploidization had an acute course of their ITP and 5 of 5 children with an impaired polyploidization developed chronic ITP (p < 0.003). CONCLUSIONS: The results in this small group of patients suggest that the assessment of the relative size of the endomitotic compartment after 7 and 12 days in plasma clot culture actually appears to be the best method for predicting a chronic course in children with ITP.

Adolescent↗
PubMed1994

Prolactin: a possible mediator of graft-versus-host disease following allogeneic bone marrow transplantation in humans.

We have evaluated post-transplant serum levels of prolactin with respect to source of donated bone marrow (BM) and to the occurrence of either acute and/or chronic graft-versus-host disease (GVHD). Forty adult patients underwent allogeneic (n = 35), autologous (n = 4) or syngeneic (n = 1) bone marrow transplantation for haematologic malignancy (n = 32) or aplastic anaemia (n = 8), respectively. Serum prolactin levels measured within 100 days post-transplant were related to patients' sex but otherwise proved unrelated to the occurrence or severity of GVHD and to the source of the BM graft (allogeneic, autologous, syngeneic). Beyond day 100 post-graft, however, serum prolactin levels proved significantly elevated in allogeneic recipients exhibiting chronic GVHD (p = 0.0004) and were unrelated to the patients' sex. In this group of patients, serum prolactin levels were not related to serum cyclosporin levels. In allogeneic recipients exhibiting no GVHD, serum prolactin levels were positively correlated with serum cyclosporin levels (p < 0.05). These data show that serum prolactin levels are significantly elevated beyond day 100 post-graft in recipients exhibiting chronic GVHD. Prolactin, a hormone recently shown also to be released by mononuclear leucocytes and to be involved in lymphocyte activation plays a hitherto unrecognized role in the pathogenesis of GVHD in humans.

Adolescent↗
PubMed1994

[The metabolism of foreign substances in the horse with reference to other animal species].

In this paper, the metabolism of xenobiotics in the horse, including differences to other species, is discussed. The most important metabolic reactions of phase I (oxidation, reduction and hydrolysis of substrates), as well as of phase II (conjugation of substrates with glucuronic acid, sulphuric acid, acetic acid, alkyl groups, amino acids, amino acid derivatives, glutathione etc.), are discussed and enzymes involved in the metabolic reactions are considered.

Animals↗
PubMed1993

Metabolism of 2,2'-dihydroxybiphenyl by Pseudomonas sp. strain HBP1: production and consumption of 2,2',3-trihydroxybiphenyl.

Cells of Pseudomonas sp. strain HBP1 grown on 2-hydroxy- or 2,2'-dihydroxybiphenyl contain NADH-dependent monooxygenase activity that hydroxylates 2,2'-dihydroxybiphenyl. The product of this reaction was identified as 2,2',3-trihydroxybiphenyl by 1H nuclear magnetic resonance and mass spectrometry. Furthermore, the monooxygenase activity also hydroxylates 2,2',3-trihydroxybiphenyl at the C-3' position, yielding 2,2',3,3'-tetrahydroxybiphenyl as a product. An estradiol ring cleavage dioxygenase activity that acts on both 2,2',3-tri- and 2,2',3,3'-tetrahydroxybiphenyl was partially purified. Both substrates yielded yellow meta-cleavage compounds that were identified as 2-hydroxy-6-(2-hydroxyphenyl)-6-oxo-2,4-hexadienoic acid and 2-hydroxy-6-(2,3-dihydroxyphenyl)-6-oxo-2,4-hexadienoic acid, respectively, by gas chromatography-mass spectrometry analysis of their respective trimethylsilyl derivatives. The meta-cleavage products were not stable in aqueous incubation mixtures but gave rise to their cyclization products, 3-(chroman-4-on-2-yl)pyruvate and 3-(8-hydroxychroman-4-on-2-yl)pyruvate, respectively. In contrast to the meta-cleavage compounds, which were turned over to salicylic acid and 2,3-dihydroxybenzoic acid, the cyclization products are not substrates to the meta-cleavage product hydrolase activity. NADH-dependent salicylate monooxygenase activity catalyzed the conversions of salicylic acid and 2,3-dihydroxybenzoic acid to catechol and pyrogallol, respectively. The partially purified estradiol ring cleavage dioxygenase activity that acted on the hydroxybiphenyls also produced 2-hydroxymuconic semialdehyde and 2-hydroxymuconic acid from catechol and pyrogallol, respectively.

Biphenyl Compounds↗
PubMed1993

On the function of histamine in the central nervous system of arthropods.

Histamine is localized in the CNS of many invertebrate taxa by various methods. It is described to act as a neurotransmitter, neuromodulator, and neurohormone. Because of its distribution throughout these taxa it is tempting to forward the following hypothesis: Histamine acts as a neuroactive substance in all invertebrates. Its function as a transmitter of photoreceptors is restricted to arthropods. A special system of six large histamine-immunoreactive neurons exists exclusively in arachnids.

Animals↗
PubMed1993

[Pattern of injuries in skateboard accidents].

In this study 101 patients who had sustained a total of 123 skateboard injuries and had been treated in the Department of Surgery at the Elisabeth-Krankenhaus in Essen between December 1989 and July 1991 were reviewed. The ages of the patients ranged between 6 and 22 years; most injuries were sustained between the ages of 12 and 17 years (67.3%). The upper extremity was injured in 50.4%. Lower extremity injuries occurred in 23.5%, head and face injuries in 22.7%, and chest injuries in 3.4%. The most common type of injury was a fracture (37.3%), most frequently to the wrist and forearm (56.5% of the fractures), followed by contusions (18.7%), lacerations (13.8%) and abrasions (10.6%). Of the 101 patients, 20 had to be hospitalized altogether 25 times, for an average duration of 7.3 days per person. The reasons for hospitalization were fractures in 18 patients, which had to be treated by open reduction and internal fixation in 7 cases and closed reduction in 14 cases. Concussion of the brain made hospitalization necessary for 2 cases.

Adolescent↗
PubMed1993

Characterization procedure for ion-selective electrode assays of magnesium activity in aqueous solutions of physiological composition.

A magnesium ion-selective liquid membrane electrode based on a synthetic neutral carrier is presented. The selectivity for Mg2+ over Na+, K+, and H+ is sufficient for assays in the physiological range, but a chemometric correction of approximately 10% is still necessary for the Ca2+ interference. We optimized the membrane composition especially with respect to selectivity and accuracy by performing a new characterization procedure in which we simulated the variety of blood serum samples with 10 aqueous solutions. After a three-point calibration, these solutions were measured alternately with the middlerange calibrator for 3 min. The magnesium activity was evaluated after correcting for the calcium interference according to the Nikolsky equation. The selectivity coefficient, KpotMgCa, was itself fitted to give the minimum deviations from the assigned magnesium activities. Electrodes with an optimized membrane composition show an average deviation from theoretical activity values of about 1.6% with logKpotMgCa = -0.8.

Calcium↗
PubMed1992

Identification of two general diffusion channels in the outer membrane of pea mitochondria.

Reconstitution experiments were performed on lipid bilayer membranes in the presence of detergent solubilized mitochondrial membranes of pea seedlings (Pisum sativum). The addition of the detergent-solubilized material to the membranes resulted in a strong increase of the membrane conductance. To identify the proteins responsible for membrane activity the detergent extracts were applied to a hydroxyapatite (HTP) column and the fractions were tested for channel formation. The eluate of the column contained a protein which migrated as a single band with an apparent molecular mass of 30 kDa on SDS-PAGE. This channel was identified as the porin of pea mitochondria since it formed voltage-dependent channels with single-channel conductances of 1.5 and 3.7 nS in 1 M KCl and an estimated effective diameter of about 1.7 nm. Further elution of the column with KCl containing solutions yielded fractions which resulted in the formation of transient channels in lipid bilayer membranes. These channels had a single-channel conductance of 2.2 nS in 1 M KCl and had also the characteristics of general diffusion pores with an estimated effective diameter of 1.2 nm. Zero-current membrane potential measurements suggested that pea porin was anion-selective in the open state. The selectivity of the second channel was investigated by the measurement of the reversal potential. It was also slightly anion-selective. Its possible role in the metabolism of mitochondria is discussed.

Anions↗