Search Results - (Author, Cooperation:Siekmann)

0022-4731

Elsevier Journal Backfiles on ScienceDirect 1907 - 2002

Biology

Chemistry and Pharmacology

Electronic Resource

1618-2650

Springer Online Journal Archives 1860-2000

Chemistry and Pharmacology

Electronic Resource

1618-2650

Springer Online Journal Archives 1860-2000

Chemistry and Pharmacology

Electronic Resource

1618-2650

Best. von Östriol, Östradiol-17β ; Massenspektrometrie/Isotopenverdünnung

Springer Online Journal Archives 1860-2000

Chemistry and Pharmacology

Electronic Resource

1618-2650

Nachw. von 17α-Äthinylöstradiol-17β in Plasma ; Massenspektrometrie/Isotopenverdünnung

Springer Online Journal Archives 1860-2000

Chemistry and Pharmacology

Electronic Resource

1052-9306

Chemistry ; Analytical Chemistry and Spectroscopy

Wiley InterScience Backfile Collection 1832-2000

Chemistry and Pharmacology

The highly specific and accurate technique of isotope dilution mass spectrometry has been used for the measurement of 17α-ethynyloestradiol-17β and norethisterone in serum. Serum samples were obtained from female volunteers who received 2.5 mg lynestrenol and 50 μg 17α-ethynyloestradiol-17β in two different galenical preparations. The determination of total 17α-ethynyloestradiol-17β (conjugated and non-conjugated) was carried out by the following procedure: (1) adsorption of the steroids from 1 ml serum to Amberlite XAD-2; (2) enzyme hydrolysis of the conjugated steroid; (3) addition of 1 ng [6,7-3H2] 17α-ethynyloestradiol-17β as internal standard; (4) column chromatography on Sephadex LH 20; (5) derivative formation with heptafluorobutyric anhydride; (6) isotope dilution mass spectrometry at m/z 474 and 478 using a glass capillary gas-liquid chromatography column. For the measurement of norethisterone, which is the major metabolite of lynestrenol, 1 ng [7-3H]norethisterone was added to 0.5 ml serum. The labelled and the non-labelled steroids were extracted and purified by column chromatography on Sephadex LH 20. The norethisterone was reacted to form the 3-enol-17β-trimethylsilyl ether of norethisterone and [3H]norethisterone. For isotope dilution mass spectrometry the derivative was injected into the glass capillary column which was coupled to the mass spectrometer. The instrument was adjusted to m/z 442 and 444, corresponding to the molecular ions of the ether derivatives of norethisterone and [7-3H]norethisterone. Accuracy was achieved by the use of the highly specific technique of mass spectrometry and the exact control of recovery using the isotope dilution principle. The precision was 4.5% (CV) for the determination of 17α-ethynyloestradiol-17β and 2.5% (CV) for norethisterone. The lower limit of detection was at 20 pg ml-1 for both methods.

6 Ill.

Electronic Resource

0306-042X

Chemistry ; Analytical Chemistry and Spectroscopy

Wiley InterScience Backfile Collection 1832-2000

Chemistry and Pharmacology

The technique of isotope dilution mass spectrometry was used for the measurement of equiline and oestrone as well as the sulphate esters of the two phenolic steroids in serum. Serum samples were obtained from an ovariectomized woman who received a tablet of 1.25 mg of various conjugated equine oestrogens. For the measurement of the non-conjugated oestrogens 20 pg (2,4,16,16-2H4)equiline and 100 pg (6,7-3H2)oestrone were added to serum samples of 0.5-4.0 ml. The steroids were extracted with 15 ml ether and separated from each other by column chromatography on Sephadex LH-20. This was followed by derivative formation with heptafluorobutyric anhydride. The ester derivatives of the two steroids were injected into a SE-52 capillary column which was coupled to a mass spectrometer. For the recording of equiline and the corresponding isotopically labelled equiline the instrument was adjusted to m/z 464 and 468, respectively. For the measurement of oestrone and (6,7-3H2)oestrone the m/z values 466 and 470 were monitored continuously. The amounts of equiline and oestrone in the serum samples were calculated from the isotope ratios measured by selected ion monitoring. For the determination of the sulphate esters of the oestrogenic steroids the serum samples (0.5-4.0 ml) from which the non-conjugated steroids had been removed by ether extraction were treated with 15 ml methanol. The serum proteins were sedimented by centrifugation and the methanolic supernatant was evaporated to dryness. The sulphate esters of the phenolic steroids were then hydrolysed by enzymatic cleavage. After an incubation period of 48 h, 50 pg (2,4,16,16-2H4)equiline and 150 pg (6,7-3H2)oestrone were added to the mixture. The oestrogens were then extracted with cyclohexane and determined as described for the measurement of the nonconjugated phenolic steroids. The precision of the method for the measurement of equiline in the range from 1.5-50 pg was at 7.3% (CV) and for oestrone in the range from 30-150 pg at 3.6% (CV). The accuracy of the procedure was achieved by the use of the isotope dilution principle in combination with the highly specific technique of selected ion monitoring.

8 Ill.

Electronic Resource

1471-4159

Blackwell Publishing Journal Backfiles 1879-2005

Medicine

Dehydroepiandrosterone and its sulphate are important factors for vitality, development and functions of the CNS. They were found to be subjects to a series of enzyme-mediated conversions within the rodent CNS. In the present study, we were able to demonstrate for the first time that membrane-associated dehydroepiandrosterone 7α-hydroxylase activity occurs within the human brain. The cytochrome P450 enzyme demonstrated a sharp pH optimum between 7.5 and 8.0 and a mean KM value of 5.4 µm, corresponding with the presence of the oxysterol 7α-hydroxylase CYP7B1. Real-time RT–PCR analysis verified high levels of CYP7B1 mRNA expression in the human CNS. The additionally observed conversion of dehydroepiandrosterone via cytosolic 17β-hydroxysteroid dehydrogenase activity could be ascribed to the activity of an enzyme with a broad pH optimum and an undetectably high KM value. Subsequent experiments with cerebral neocortex and subcortical white matter specimens revealed that 7α-hydroxylase activity is significantly higher in the cerebral neocortex than in the subcortical white matter (p 〈 0.0005), whereas in the subcortical white matter, 17β-hydroxysteroid dehydrogenase activity is significantly higher than in the cerebral neocortex (p 〈 0.0005). No sex differences were observed. In conclusion, the high levels of CYP7B1 mRNA in brain tissue as well as in a variety of other tissues in combination with the ubiquitous presence of 7α-hydroxylase activity in the human temporal lobe led us to assume a neuroprotective function of the enzyme such as regulation of the immune response or counteracting the deleterious effects of neurotoxic glucocorticoids, rather than a distinct brain specific function such as neurostimulation or neuromodulation.

Electronic Resource

1471-4159

Blackwell Publishing Journal Backfiles 1879-2005

Medicine

Dehydroepiandrosterone (DHEA) and its sulfate (DHEAS) are suggested to be important neurosteroids. We investigated steroid sulfatase (STS) in human temporal lobe biopsies in the context of possible cerebral DHEA(S) de novo biosynthesis. Formation of DHEA(S) in mature human brain tissue has not yet been studied. 17α-Hydroxylase/C17-20-lyase and hydroxysteroid sulfotransferase catalyze the formation of DHEA from pregnenolone and the subsequent sulfoconjugation, respectively. Neither their mRNA nor activity were detected, indicating that DHEA(S) are not produced within the human temporal lobe. Conversely, strong activity and mRNA expression of DHEAS desulfating STS was found, twice as high in cerebral neocortex than in subcortical white matter. Cerebral STS resembled the characteristics of the known placental enzyme. Immunohistochemistry revealed STS in adult cortical neurons as well as in fetal and adult Cajal-Retzius cells. Organic anion transporting proteins OATP-A, -B, -D, and -E showed high mRNA expression levels with distinct patterns in cerebral neocortex and subcortical white matter. Although it is not clear whether they are expressed at the blood–brain barrier and facilitate an influx rather than an efflux, they might well be involved in the transport of steroid sulfates from the blood. Therefore, we hypothesize that DHEAS and/or other sulfated 3β-hydroxysteroids might enter the human temporal lobe from the circulation where they would be readily converted via neuronal STS activity.

Electronic Resource

1365-2826

Blackwell Publishing Journal Backfiles 1879-2005

Medicine

Sex steroids exert important effects on the central nervous system (CNS). Although the formation of 17β-hydroxysteroid dehydrogenase (17β-HSD) metabolites in the CNS was discovered almost 30 years ago, conclusive studies concerning 17β-HSD activity in the human brain are still lacking. Therefore, we investigated 17β-HSD in vitro activity in human temporal lobe biopsies of 13 women and 13 men using radioactively labelled androstenedione, testosterone, oestrone and 17β-oestradiol and compared it to that in human placenta, liver, testis and prostate. We could demonstrate androgenic and oestrogenic 17β-HSD activities in all tissues under investigation. The reduction of androstenedione and oestrone in brain was NADPH dependent with a broad pH optimum between 6.5 and 9.0, whereas the oxidation of testosterone and 17β-oestradiol was NAD dependent with a pH optimum of ≥9.0. Using optimum cofactors sex differences of brain 17β-HSD activities were not observed. Conversion of androstenedione, testosterone, oestrone and 17β-oestradiol was significantly higher in the subcortical white matter than in the cerebral cortex. We could demonstrate a significant formation of testosterone in the brain tissue of all patients under investigation. Substrate specificity and cofactor requirement patterns as well as pH optima and kinetic properties suggest the occurrence of 17β-HSD type 3 and type 4 in the human temporal lobe.

Electronic Resource

1365-2516

Blackwell Publishing Journal Backfiles 1879-2005

Medicine

A new collagen binding assay has been developed for the determination of the functional activity of human von Willebrand factor based on the following principle: pepsin-digested type III collagen from human placenta was covalently immobilized on a microtitre plate. Binding of collagen to the microtitre plate was carried out in neutral phosphate buffer within 1 h. A collagen concentration of 3 μg mL–1 was sufficient to achieve optimal coating. After drying, the coated microtitre plates remained stable for months without losing their vWF-binding properties and could be incorporated into a ready-to-use kit. The suitability of various types of collagen for use in this assay was evaluated by simulating the binding of vWF to collagen immobilized on the microtitre plate by using surface plasmon resonance technology with collagen immobilized on a sensor chip. vWF was most effectively bound to pepsin-digested type III collagen from human placenta. The assay thus comprises the following steps: (1) serial dilutions of a vWF reference preparation and vWF-containing samples are prepared and bound to a microtitre plate which is precoated with collagen; (2) vWF is detected with a polyclonal antibody; (3) the substrate reaction is photometrically measured with an ELISA reader. This test is highly specific and sensitive for vWF (detection limit: 10 ng mL–1). The collagen binding activity measured corresponds to the degree of multimerization of vWF. The high sensitivity and the short time needed to carry it out may make it useful for clinical diagnosis and for the measurement of the functional activity of vWF in factor concentrates. In certain applications it may represent a suitable replacement for the ristocetin cofactor assay.

Electronic Resource

1365-2516

Blackwell Publishing Journal Backfiles 1879-2005

Medicine

Patients with severe forms of von Willebrand disease (vWD) most frequently require substitution of both factor VIII and von Willebrand factor (vWF). Immunate is a high-purity, double-virus inactivated FVIII/vWF concentrate derived from human plasma. The clinical efficacy of Immunate concerning the management of acute bleeding episodes and surgical prophylaxis in patients with von Willebrand disease (vWD) is being investigated and documented in a prospective, phase III, open-label, single-armed multicenter trial. Data on its pharmacokinetics are collected, and the product's safety with respect to adverse experiences is monitored.

Electronic Resource

1476-4687

Nature Archives 1869 - 2009

Biology

Chemistry and Pharmacology

Medicine

Natural Sciences in General

Physics

[Auszug] Recent evidence indicates that growing blood-vessel sprouts consist of endothelial cells with distinct cell fates and behaviours; however, it is not clear what signals determine these sprout cell characteristics. Here we show that Notch signalling is necessary to restrict angiogenic cell ...

Electronic Resource

0375-9601

Elsevier Journal Backfiles on ScienceDirect 1907 - 2002

Physics

Electronic Resource

0012-0936

German, Dutch and Scandinavian Studies

0016-8831

German, Dutch and Scandinavian Studies

0022-0248

Elsevier Journal Backfiles on ScienceDirect 1907 - 2002

Chemistry and Pharmacology

Geosciences

Physics

Electronic Resource

0020-7462

Elsevier Journal Backfiles on ScienceDirect 1907 - 2002

Mechanical Engineering, Materials Science, Production Engineering, Mining and Metallurgy, Traffic Engineering, Precision Mechanics

Physics

Electronic Resource

0020-7462

Elsevier Journal Backfiles on ScienceDirect 1907 - 2002

Mechanical Engineering, Materials Science, Production Engineering, Mining and Metallurgy, Traffic Engineering, Precision Mechanics

Physics

Electronic Resource

0022-4731

Elsevier Journal Backfiles on ScienceDirect 1907 - 2002

Biology

Chemistry and Pharmacology

Electronic Resource