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Biomedical subjects

D Kirsch

Publications and source records attributed to D Kirsch.

At least 37 records · Page 2Linked to original sources
PubMed1997

[Evaluation and internal quality assurance in general and abdominal surgery].

The measuring of perioperative risk, the finding of intraoperative influences and the objective rating of quality of treatment were achieved by our program of quality assurance. With our system it is possible to measure differences between several surgeons and also differences in comparison to national and international results. Through optimised therapy the program can lead to an improvement of quality.

Data Collection↗
PubMed1996

Analysis of carotenoids and carotenol fatty acid esters by matrix-assisted laser desorption ionization (MALDI) and MALDI-post-source-decay mass spectrometry.

Matrix-assisted laser desorption ionization (MALDI) time-of-flight mass spectrometry and MALDI post-source-decay (PSD) fragment ion mass analysis were employed to examine various model carotenoids and some of their fatty acid esters. It was demonstrated that the use of MALDI helps overcome problems resulting from the inherent thermal instability and lack of solubility which render this group of compounds rather difficult for conventional ionization techniques. Detection limits were in the subpicomolar range. Rather abundant metastable fragmentation occurred under conditions of prompt extraction, but was significantly restricted under conditions of delayed extraction (DE). In DE the quasimolecular ion signals (mainly as odd electron radical ions) became the base peak even in spectra of the most delicate fatty acid esters. PSD fragment ion patterns were similar to those recorded under linked B/E scanning in conventional fast atom bombardment-MS and contained information on structural end group substituents such as hydroxyl, epoxide, or carbonyl functions and on the extent of double bond conjugations. The mono- and bis-fatty acid esters fragmented mainly by simple fatty acid cleavage but also furnished some of the end group-specific fragments.

Carotenoids↗
PubMed1996

Post-source decay and delayed extraction in matrix-assisted laser desorption/ionization-reflectron time-of-flight mass spectrometry. Are there trade-offs?

By the incorporation of delayed extraction (DE) into matrix-assisted laser desorption/ionization time-of-flight mass spectrometry a dramatic improvement of performance with respect to sensitivity, mass resolution and mass accuracy of precursor ions up to approximately 10 kDa has been achieved. Since DE reduces collisional in-source activation to a large extent, the rate of subsequent metastable decay is considerably reduced. Results are presented which demonstrate that under DE the loss of total post-source decay (PSD) fragment ion yield can be as large as one order of magnitude but that, in terms of sensitivity, part of this loss is balanced by a better S/N ratio which results from a significantly improved mass resolution of the PSD fragment ions (M/delta M up to 1800 compared with M/delta M = 200-500 under prompt extraction). While this compensatory effect is true for the middle to high mass range of PSD fragment ions, it gradually vanishes towards the low mass end of the PSD mass scale where, in the case of linear peptides some important information (immonium ions) is lost. It appears, however, that in the majority of practical PSD work, DE improves the qualty of the PSD spectra and that high energy collisional post-source activation can compensate for the occasional loss of analytical information.

Amino Acid Sequence↗
PubMed1995

Photolabeling reveals the proximity of the alpha-neurotoxin binding site to the M2 helix of the ion channel in the nicotinic acetylcholine receptor.

A photoactivatable derivative of neurotoxin II from Naja naja oxiana containing a 125I-labeled p-azidosalicylamidoethyl-1,3'-dithiopropyl label at Lys-25 forms a photo-induced cross-link with the delta subunit of the membrane-bound Torpedo californica nicotinic acetylcholine receptor (AChR). The cross-linked radioactive receptor peptide was isolated by reverse-phase HPLC after tryptic digestion of the labeled delta subunit. The sequence of this peptide, delta-(260-277), and the position of the label at Ala-268 were established by matrix-assisted laser-desorption-ionization mass spectrometry based on the molecular mass and on post-source decay fragment analysis. With the known dimensions of the AChR molecule, of the photolabel, and of alpha-neurotoxin, finding the cross-link at delta Ala-268 (located in the upper part of the channel-forming transmembrane helix M2) means that the center of the alpha-neurotoxin binding site is situated at least approximately 40 A from the extracellular surface of the AChR, proximal to the channel axis.

Affinity Labels↗
PubMed1995

The effect of FK506 on glycemic response as assessed by the hyperglycemic clamp technique.

We studied seven nondiabetic subjects with the autoimmune diseases psoriasis, multiple sclerosis, and primary biliary sclerosis who were to receive FK506 as experimental immunotherapy. All subjects underwent two standard oral glucose tolerance tests and two 180-min hyperglycemic clamps immediately before and 10 weeks after starting FK506. There was no significant difference in weight or HbA1c pre- vs. post-FK506 treatment. FK506 levels were therapeutic and non-toxic (0.1-1 ng/ml) for all subjects studied. Repeated measures analysis of variance for interaction between time and treatment was performed on insulin (after outlier removed) and glucose values from the OGTT. There was neither time-by-treatment interaction, nor a treatment effect (P > 0.1). There were no significant differences in pre- vs. post-FK506 treatment values of plasma glucose during the hyperglycemic clamp mean acute insulin response to glucose (AIRG) 164 +/- 38 pmol/L vs. 148 +/- 46 pmol/L (P > 0.1); mean incremental area under the insulin curve (IAUC) during the first 10 min of the study, 473 +/- 109 pmol/L vs. 443 +/- 146 pmol/L (P > 0.1); total area under the insulin curve (TAUC) during the first 10 min of the study, 786 +/- 152 pmol/L vs. 781 +/- 18 pmol/L (P > 0.1); mean glucose infusion rate (GIR) 37.7 +/- 5.0 mumol/kg/min vs. 33.3 +/- 4.4 mumol/kg/min (P > 0.1); or mean insulin sensitivity index (ISI), 3.05 +/- 0.4 vs. 3.13 +/- 0.5 (P > 0.1). Mean steady-state insulin secretion (SSI) was significantly lower 244 +/- 43 pmol/L vs. 200 +/- 25.2 pmol/L (P = 0.03). Peak first-phase insulin secretion values of 321 +/- 62 pmol/L vs. 263 +/- 57 pmol/L approached significance (P = 0.07). No patient progressed to diabetes during the study. FK506 decreased steady-state insulin secretion during the last 60 min of the clamp, regardless of initial glucose tolerance. Insulin sensitivity and glucose infusion rate did not change in the group as a whole with FK506 treatment.

Blood Glucose↗
PubMed1994

A single amino acid substitution (Glu134-->Ala) in NhaR1 increases the inducibility by Na+ of the product of nhaA, a Na+/H+ antiporter gene in Escherichia coli.

The mutation nhaAup (antup) has now been identified as a Glu134 to Ala substitution in NhaR and designated nhaR1. This was demonstrated by sequence analysis showing that the mutant contains a wild-type nhaA but nhaR1 instead of nhaR and by the finding that nhaR1 cloned in a plasmid confers the NhaAup phenotype. Na+ (107 mM) increases by 5- to 10-fold the level of nhaA transcripts, similar to the effect on the NhaR-mediated expression of a nhaA'-'lacZ fusion. These results are in agreement with the notion that nhaR is a positive regulator which controls Na(+)-dependent transcription of nhaA. The promoter region of nhaR and nhaR1 was found to reside within the BglII-BamHI fragment of the C-terminal sequences of nhaA. The mutation nhaR1, while increasing dramatically the level of transcription, reduces the requirement for Na+ by 3- to 5-fold both for nhaA transcription and for the nhaR1-mediated expression of nhaA'-'lacZ fusion. NhaR1, like NhaR, binds specifically to the promoter region of nhaA. However, at equal protein concentration NhaR1 binds more DNA and the NhaR1-DNA complex shows higher mobility than that of NhaR-DNA, suggesting the existence of two different binding complexes. Yet in this assay the DNA binding pattern of neither NhaR nor NhaR1 was affected by the addition of Na+. The possible relevance of these two DNA-binding complexes to the Na(+)-induced NhaR-mediated expression is discussed.

Amino Acid Sequence↗
PubMed1994

[Aortoduodenal fistula. Incidence--diagnosis--therapy].

Six patients were operated on for an aortoduodenal fistula at the Department of General and Abdominal Surgery, Johannes-Gutenberg-University Mainz. All patients had received an aortic graft implant between 1 and 10 years ago. The etiology, symptoms, and diagnostic and surgical treatment of these six cases are presented, and the results are discussed in comparison with the results of other studies. The main symptom of all of our patients was gastrointestinal bleeding. In our patients arteriography and computed tomography were the best diagnostic procedures. At surgery, five patients underwent graft excision and axillobifemoral bypass (only four patients, as one died before implantation). In the other patient a local repair, with closure of the graft defect and bowel defect, followed by interposition of an omental pedicle, was performed. Two patients died within 6 days of operation due to multiple organ failure.

Aged↗
PubMed1992

Peptide sequencing by matrix-assisted laser-desorption mass spectrometry.

A novel method of peptide sequencing by mass spectrometry is described. Metastable decay of laser-desorbed ions, taking place in the first field-free drift region of a reflection time-of-flight mass spectrometer, has been monitored to get structural information from larger peptides. Fragment ions from metastable decay are mass analysed by adjusting the potentials of the ion reflectron according to the kinetic energies of the ions. The features of the technique and its significance for future applications are outlined.

Amino Acid Sequence↗
PubMed1987

Catecholamines and tumour promoting phorbolesters inhibit insulin receptor kinase and induce insulin resistance in isolated human adipocytes.

The effect of the catecholamine isoprenaline (10(-5) mol/l) and of the tumour promoting phorbolester tetradecanoyl-beta-phorbol acetate (10(-9) mol/l) on insulin stimulated 3-O-methyl-glucose transport was studied in freshly isolated human adipocytes. Both substances reduced the maximal responsiveness of the glucose transport system to insulin by approximately 50%. To test if this is caused by inhibition of the insulin receptor kinase the receptor from phorbolester and isoprenaline treated cells was solubilized, partially purified and its kinase activity studied in vitro. Insulin stimulated 32P-incorporation into the beta-subunit of the insulin receptor of phorbolester or isoprenaline treated cells was reduced to 20-60% of the values found with receptor from control cells at insulin concentrations between 10(-10) mol/l and 10(-7) mol/l. This inhibition of kinase activity of receptor from phorbolester and isoprenaline treated cells was observed at nonsaturating adenosine triphosphate levels (5 mumol/l), and it could be overcome with higher concentrations of gamma-32P-adenosine triphosphate in the phosphorylation assay. A Lineweaver Burk analysis of the insulin stimulated receptor phosphorylation revealed that the Michaelis constant for adenosine triphosphate of the receptor kinase from phorbolester and isoprenaline treated cells was increased to greater than 100 mumol/l compared with less than 50 mumol/l for receptor from control cells. We conclude from the data that catecholamine and phorbolester treatment of human adipocytes modulates the kinase activity of the insulin receptor by increasing its Michaelis constant for adenosine-triphosphate, and propose that this modulation of receptor kinase is a mechanism that can contribute to the pathogenesis of insulin resistance in human fat cells.

3-O-Methylglucose↗
PubMed1987

Insulin receptor kinase defects as a possible cause of cellular insulin resistance.

The insulin receptor contains in its beta-subunit a tyrosine (-) specific protein kinase. It is believed that transmission of an insulin signal across the plasma membrane of target cells of insulin action occurs through activation of this kinase, autophosphorylation of the insulin receptor beta-subunit and subsequent phosphorylation of other cellular substrates. We studied the insulin receptor kinase in a number of insulin resistant cell systems in order to elucidate if defects of this kinase are a possible cause of cellular insulin resistance. Three different patterns of kinase abnormalities were found, in different insulin resistant cells: 1. In an insulin resistance melanoma cell line a reduced receptor kinase autophosphorylation was found apparently due to a defect of the tyrosine autophosphorylation sites of this receptor; 2. Catecholamine and phorbol ester induced insulin resistance of isolated rat fat cells as well as human fat cells was associated with a decreased activity of the insulin receptor tyrosine kinase which was apparently due to a modulation of the ATP binding site of the insulin receptor tyrosine kinase; 3. The receptor kinase isolated from the skeletal muscle of diabetic Zucker rats (fa/fa) was found to be insulin insensitive with no major alteration of maximal responsiveness. These results suggested that different forms of kinase defects exist which can contribute to the pathogenesis of cellular insulin resistance. Based on these data studies in skeletal muscle from type II diabetic patients were started. Results from five patients so far suggest that, here as well, an abnormality of the insulin receptor kinase exists which might be involved in the pathogenesis of insulin resistance in type II diabetes.

Adipose Tissue↗
PubMed1986

Tumor-promoting phorbol esters increase the Km of the ATP-binding site of the insulin receptor kinase from rat adipocytes.

Phorbol ester treatment of isolated rat adipocytes inhibits insulin stimulation of the glucose transport system. We studied whether this effect is related to a modification of the insulin receptor kinase. Insulin receptor of tetradecanoyl-beta-phorbol acetate (TPA)-treated adipocytes was solubilized and partially purified, and its kinase activity was studied in vitro. We found that insulin (10(-7) M) increased the tyrosine autophosphorylation of the insulin receptor kinase from TPA-treated cells only 3-fold in contrast to a 12-fold stimulation in control cells. The rate of insulin-stimulated 32P incorporation into the receptor of TPA-treated cells at insulin concentrations between 10(-10) M and 10(-7) M and at nonsaturating [32P]ATP levels (5 microM) was reduced to only 5-8% of the values found with receptor from control cells. 125I-insulin binding to the solubilized receptor of TPA-treated cells was reduced as well, apparently due to a decreased affinity of the binding site. Decreased binding was however, not sufficient to explain the difference of kinase activity. The inhibition of kinase activity of the receptor from TPA-treated cells decreased when the concentration of [gamma-32P]ATP in the phosphorylation assay was increased. A Lineweaver-Burk analysis of receptor phosphorylation revealed that the Km for ATP of the receptor kinase from TPA-treated cells was increased to greater than 100 microM compared to 25 microM for the receptor of control cells. An analogous change of the Km for ATP was found when we studied the phosphorylation of a synthetic polymer of tyrosine and glutamic acid as a substrate of the receptor kinase. We conclude from the data that phorbol treatment of rat adipocytes modulates the kinase activity of the insulin receptor by increasing its Km for ATP and that this is part of a mechanism leading to insulin resistance in these cells.

Adenosine Triphosphate↗
PubMed1986

Decreased tyrosine kinase activity of insulin receptor isolated from rat adipocytes rendered insulin-resistant by catecholamine treatment in vitro.

Catecholamine treatment of isolated rat adipocytes decreases insulin binding and inhibits insulin stimulation of the glucose-transport system. There is increasing evidence that the insulin signal is transmitted after insulin is bound to the receptor via a tyrosine kinase, which is an intrinsic part of the receptor. To find whether the receptor kinase is modified by catecholamines, we solubilized and partially purified the insulin receptor of isoprenaline-treated adipocytes and studied the effect of insulin on its kinase activity. (1) Insulin increased the tyrosine autophosphorylation of the insulin receptor kinase from catecholamine-treated cells only 4-fold, compared with a 12-fold stimulation in control cells. (2) The rate of insulin-stimulated 32P incorporation into the receptor of isoprenaline-treated cells at non-saturating [32P]ATP concentrations (5 muM) was decreased to 5-8% of the values for receptor from control cells. (3) 125I-insulin binding to the partially purified receptor from catecholamine-treated cells was also markedly decreased. The insulin receptor from catecholamine treated cells bound 25-50% of the amount of insulin bound by the receptor from control cells at insulin concentrations of 10 pM-0.1 muM. Part of the impaired insulin-responsiveness of the receptor kinase of catecholamine-treated cells is therefore explained by impaired binding properties; however, an additional inhibition of the kinase activity of the insulin receptor from catecholamine-treated cells is evident. (4) This inhibition of kinase activity decreased when the concentration of [gamma-32P]ATP in the phosphorylation assay was increased. A Lineweaver-Burk analysis revealed that the Km for ATP of the receptor kinase from isoprenaline-treated cells was increased to approx. 100 muM, compared with approx. 25 muM for receptor of control cells. (5) We conclude from the data that catecholamine treatment of rat adipocytes modulates the kinase activity of the insulin receptor by increasing its Km for ATP and that this is part of the mechanism leading to insulin-resistance in these cells.

Adenosine Triphosphate↗
PubMed1985

Insulin receptor kinase in human skeletal muscle.

Receptor-associated protein kinase activity has been shown in all primary target tissues of insulin action in the rat and a function of insulin receptor phosphorylation in signal transmission was proposed. Insulin receptor phosphorylation so far has not been demonstrated in human target tissues of insulin. We describe here insulin receptor kinase activity in human skeletal muscle. Insulin (10(-8) mol/l) stimulates the phosphorylation of a 95-kDa protein from skeletal muscle 2-fold. The phosphoprotein is quantitatively immunoprecipitated with insulin receptor antibody identifying it as the beta-subunit of the insulin receptor. The insulin stimulation of phosphorylation is detectable also at physiological insulin concentrations (10(-9) mol/l) showing that receptor phosphorylation could be involved in insulin action in human skeletal muscle as well.

Animals↗