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Biomedical subjects

K Schmidt

Publications and source records attributed to K Schmidt.

At least 307 records · Page 17Linked to original sources
PubMed1993

Inhibition of nitric oxide synthesis by methylene blue.

Methylene blue appears to inhibit nitric oxide-stimulated soluble guanylyl cyclase and has been widely used for inhibition of cGMP-mediated processes. We report here that endothelium-dependent relaxation of isolated blood vessels and NO synthase-dependent cGMP formation in cultured endothelial cells were both markedly more sensitive to inhibition by methylene blue than effects induced by direct activation of soluble guanylyl cyclase. These discrepancies were also observed when superoxide dismutase (SOD) was present to protect NO from inactivation by superoxide anion. Subsequent experiments showed that formation of L-citrulline by purified NO synthase was completely inhibited by 30 microM methylene blue (IC50 = 5.3 and 9.2 microM in the absence and presence of SOD, respectively), whereas guanylyl cyclase stimulated by S-nitrosoglutathione was far less sensitive to the drug (50% inhibition at approximately 60 microM, and maximal inhibition of 72% at 1 mM methylene blue). Experimental evidence indicated that oxidation of NADPH, tetrahydrobiopterin or reduced flavins does not account for the inhibitory effects of methylene blue. Our data suggest that methylene blue acts as a direct inhibitor of NO synthase and is a much less specific and potent inhibitor of guanylyl cyclase than hitherto assumed.

Amino Acid Oxidoreductases↗
PubMed1993

Pteridine biosynthesis in human endothelial cells. Impact on nitric oxide-mediated formation of cyclic GMP.

Stimulation of nitric oxide (NO) synthase in endothelial cells by Ca2+ influx leads to increased intracellular levels of cGMP. NO synthase from various sources is known to use tetrahydrobiopterin, flavins, and NADPH as cofactors. We studied the effect of interferon-gamma, tumor necrosis factor-alpha, and lipopolysaccharide on tetrahydrobiopterin biosynthetic activities in human umbilical vein endothelial cells (HUVEC). These stimuli led to an up to 40-fold increase of GTP cyclohydrolase I (EC 3.5.4.16) activity and to increased accumulation of neopterin and tetrahydrobiopterin in HUVEC. Further enzyme activities of tetrahydrobiopterin biosynthesis, i.e. 6-pyruvoyl tetrahydropterin synthase and sepiapterin reductase (EC 1.1.1.153), remained unchanged. NO synthase activity in protein fractions from homogenates of cells treated with interferon-gamma plus tumor necrosis factor-alpha was not influenced as compared with untreated controls. However, interferon-gamma alone or in combination with tumor necrosis factor-alpha significantly increased intracellular cGMP formation in intact HUVEC by 50 and 80%, respectively. These stimuli increased intracellular tetrahydrobiopterin concentrations up to 14-fold. NO-triggered cGMP formation was similarly increased by incubation of otherwise untreated cells with sepiapterin, leading to elevated intracellular tetrahydrobiopterin levels. Thus, cytokines indirectly stimulate the activity of constitutive NO synthase in HUVEC by upregulating production of the cofactor tetrahydrobiopterin.

Amino Acid Oxidoreductases↗
PubMed1993

Bromocriptine treatment over 12 years in acromegaly: effect on glucose tolerance and insulin secretion.

It is not known whether the beneficial effect of bromocriptine on glucose homeostasis in acromegaly is limited by a certain duration of therapy. To elucidate this problem, oral glucose tolerance tests were performed in 12 acromegaly patients before bromocriptine medication, under therapy (15.0 +/- 6.8 mg/day for 12 +/- 3 years), and during a 2-week drug withdrawal after long-term treatment. Initially altered glucose tolerance was normalized in 4 of 5 patients under bromocriptine therapy. During drug withdrawal the mean fasting glucose level and the mean glucose concentration at 120 min after oral glucose load increased from 5.05 +/- 0.61 to 5.77 +/- 0.78 mmol/l and from 5.61 +/- 2.05 to 7.55 +/- 3.05 mmol/l, respectively. A deterioration in glucose homeostasis was observed in 9 patients, and impaired glucose tolerance was ameliorated (but not to normal range) in 2 when bromocriptine was withdrawn. The proportion of alterations in glucose tolerance during drug withdrawal corresponded to that before the beginning of long-term bromocriptine treatment. Impaired glucose tolerance, observed in 2 patients under bromocriptine treatment, seemed to be compensated because a distinct elevation of glycosylated hemoglobin A1c was not observed. Bromocriptine led to a significant decrease in basal as well as glucose-stimulated insulin levels, and growth hormone secretion during oral glucose load was reduced in all 12 patients.(ABSTRACT TRUNCATED AT 250 WORDS)

Acromegaly↗
PubMed1993

Megadose glucocorticoid therapy: investigations in rat anaphylactic shock and experimental allergic encephalomyelitis.

Glucocorticoids are the most potent antiinflammatory drugs. Large doses of dexamethasone and other glucocorticoids are widely used in the clinic but the mechanism of the beneficial effects of megadoses still remains to be explained. We tested the effects of a dexamethasone (DEX) megadose therapy in a rat model of anaphylactic shock. By combining DEX with the potent glucocorticoid receptor antagonist RU 486 we looked for evidence of a non-specific action of the glucocorticoid at high doses. A dose of 10 mg/kg DEX given 2 h before challenge protected the heart against the symptoms of cardiac anaphylaxis: heart rate was improved by 22%, ventricular contractility by 13% and coronary flow by 5%. Administration of 20 mg/kg RU 486, 30 min before dexamethasone, reduced the beneficial effect of DEX by about 30%, although this failed to reach statistical significance. We found no dose-response relationship for the high dexamethasone doses of 0.5-10 mg/kg. In experimental allergic encephalomyelitis (EAE) there was a surprising toxic action of DEX after daily doses of 0.25 mg/kg for 5 days, and also following single administration of 2.5 or 10 mg/kg. A single dose of 1.25 mg/kg, given on the day of immunization alone or with additional doses at weekly intervals for 3 weeks, caused a strong inhibition of the EAE. In summary we conclude that the present data of cardiac anaphylaxis hardly point to extra glucocorticoid megadose effects. The present dexamethasone effects in cardiac anaphylaxis and in EAE have to be cleared up by further experiments.

Anaphylaxis↗
PubMed1993

Analgesic effect of the somatostatin analogue octreotide in two acromegalic patients: a double-blind study with long-term follow-up.

Two acromegalic patients with severe headache were treated with the somatostatin analogue, octreotide (Sandostatin). A double-blind study of octreotide versus placebo in which pain intensity was measured using a visual analogue scale (VAS) was performed initially with these patients. A rapid (within 4-15 min) pain relief occurred lasting 2-8.5 h after injection of 100 micrograms of octreotide, an effect that was not reversed by intravenous (i.v.) naloxone. These 2 acromegalic patients then received treatment for 71 and 82 months, respectively, with doses starting at 500 micrograms/day and 1500 micrograms/day, respectively, without evidence of either tolerance or dependence, although the effect of octreotide on headache appears to be selective. No unwanted sedative effect has been observed. A screening procedure with injection of 50 micrograms of subcutaneous (s.c.) octreotide was performed in 11 other patients with chronic severe pain associated with various conditions. Only 3 patients (2 with diabetic polyneuropathy and 1 with bone pain associated with myelodysplastic syndrome) reported more than 50% pain relief. In the insulin-dependent diabetic patients the double-blind check was not performed due to the risk of octreotide-induced hypoglycemia. In the patient with bone pain the same double-blind check as in the acromegalic patients could not confirm the analgesic effect. It may thus be concluded that octreotide appears to be useful for the treatment of both chronic and acute severe painful conditions in acromegalic patients. However, since its analgesic effect in our patients was confined to headaches only, further controlled studies must be carried out in order to determine appropriate target groups.

Acromegaly↗
PubMed1993

Synthesis of the Escherichia coli K-12 nucleoid-associated DNA-binding protein H-NS is subjected to growth-phase control and autoregulation.

Mutations in the structural gene (hns) for the Escherichia coli nucleoid-associated DNA-binding protein H-NS cause highly pleiotropic effects on gene expression, site-specific recombination, transposition of phage Mu, the stability of the genetic material and the topological state of the DNA. We have investigated the regulation of hns expression and found that hns transcription is subjected to stationary phase induction and negative autoregulation. A set of hns-lacZ protein and operon fusions was constructed in vitro and integrated in single copy into the attB site of the bacterial genome. Quantification of beta-galactosidase activity along the bacterial growth curve showed that hns expression increases approximately 10-fold in stationary phase compared with exponentially growing cells. Immunological detection of the H-NS protein in growing and stationary phase cells supported the genetic data and showed that H-NS synthesis varies with growth phase. In addition, primer extension experiments demonstrated that the amount of hns mRNA is elevated in stationary phase cultures and that hns transcription is directed by a unique promoter functioning in both log and stationary phase. Disruption of the hns gene by an insertion mutation led to the derepression (approximately fourfold) of the expression of an hns-lacZ operon fusion integrated at the attB site, showing that hns transcription is subjected to negative regulation by its own gene product. Autoregulation of hns expression is particularly pronounced in log phase. Both stationary phase control and autoregulation of hns transcription are associated with a 130 bp fragment that contains the hns promoter. In order to study the interaction of H-NS with its own regulatory region, we developed an efficient overproduction procedure and a simple purification scheme for H-NS. DNA gel retardation assays showed that the H-NS protein can preferentially interact with a restriction fragment carrying the hns promoter. This restriction fragment showed features of curved DNA as judged by two-dimensional polyacrylamide gel electrophoresis performed at 4 degrees C and 60 degrees C.

Amino Acid Sequence↗
PubMed1993

Importance of cerebral pan-angiography for detection of multiple aneurysms in patients with aneurysmal subarachnoid haemorrhage.

In the Danish Aneurysm Study 948 patients had a ruptured intracranial aneurysm (RA) revealed by cerebral angiography. Unilateral carotid angiography (UCA) with or without vertebral angiography (VA) demonstrated the RA in 206 patients, and 16 (7.8%) unruptured aneurysms (UA) were disclosed by UCA and 1 UA by VA. In 740 patients with at least bilateral carotid angiography (BCA) 164 UA (22.2%) were disclosed by BCA and 10 by VA. I.e. the number of UA demonstrated in patients with RA is not only doubled by BCA compared to ipsilateral UCA, but tripled demonstrating that an additional UA is more likely situated on the opposite side of the RA. Pan-angiography (or at least BCA) is therefore recommended in patients with RA.

Adolescent↗
PubMed1993

Hypercholesterolemia is associated with a reduced response of smooth muscle guanylyl cyclase to nitrovasodilators.

A diminished relaxant response of atherosclerotic arteries to nitrovasodilators has been frequently observed in advanced stages of hypercholesterolemia. In the present study, we investigated whether this effect might be a result of reduced activity of smooth muscle guanylyl cyclase. Experimental atherosclerosis was induced by feeding rabbits a cholesterol-rich diet (1%) over a period of 4 months. Aortas were removed and homogenized, and guanylyl cyclase activity was measured in the 100,000 g supernatants. Sodium nitroprusside, which stimulated cyclic GMP (cGMP) formation in control tissues almost 200-fold (from 3 to 585 pmol cGMP.mg-1 x min-1), increased enzyme activities in atherosclerotic aortas only approximately 90-fold (from 3 to 257 pmol cGMP.mg-1 x min-1). Similarly, the maximal stimulatory effects of S-nitroso-glutathione were reduced from 200-fold (controls) to 114-fold in atherosclerotic tissues. Basal guanylyl cyclase activities were identical in both atherosclerotic and control vessels. Hypercholesterolemia also reduced the activity of smooth muscle adenylyl cyclase. In control aortas, basal and NaF-stimulated enzyme activities were 24 and 349 pmol cAMP.mg-1 x min-1, respectively, whereas cAMP formation was reduced in atherosclerotic aortas to 7 (basal) and 96 (NaF) pmol cAMP.mg-1.min-1. The stimulatory effect of NaF (approximately 14-fold) remained unchanged. Since adenylyl and guanylyl cyclase have important functions in regulating vascular tone, reduced activities of both enzymes may contribute to the diminished relaxant and/or enhanced vasoconstricting effects of vasoactive compounds in atherosclerotic blood vessels.

Adenylyl Cyclases↗
PubMed1993

Clinical features and outcome in females and males with ruptured intracranial saccular aneurysms.

Of 1076 patients with aneurysmal subarachnoid haemorrhage 674 were females and 402 males. No significant differences between females and males were seen as regards age, clinical condition on admission, pre-existing arterial hypertension and number of rebleeds. Angiographically demonstrated vasospasm was seen with a significantly higher incidence in females (p < 0.05) which may possibly explain a significantly poorer outcome in females compared with males (p < 0.05), despite the much higher rate in females of internal carotid artery aneurysms which have a significantly better prognosis compared with aneurysms at other sites.

Adolescent↗
PubMed1993

Novel actions of methylene blue.

Methylene blue has been frequently used as an inhibitor of soluble guanylyl cyclase. We found that endothelium-dependent relaxations of isolated blood vessels were considerably more sensitive to inhibition by methylene blue than relaxation induced by direct activators of soluble guanylyl cyclase. Similar data were obtained in the presence of superoxide dismutase, indicating that the diverse potencies of methylene blue were not due to superoxide-induced inactivation of nitric oxide (NO). Subsequent experiments revealed that methylene blue is an inhibitor of purified NO synthase. Conversion of L-arginine to L-citrulline was inhibited by the dye in a concentration-dependent fashion with half-maximal effects observed at 5.3 microM and 9.2 microM in the absence and presence of superoxide dismutase, respectively. Purified soluble guanylyl cyclase, however, was far less sensitive to methylene blue. When the enzyme was maximally stimulated with S-nitroso-glutathione, cyclic guanosine monophosphate, (cGMP) formation was reduced by 50% at approximately 60 microM methylene blue; 1 mM produced maximal inhibitions of about 70%. Our data indicate that methylene blue is only a poor inhibitor of soluble guanylyl cyclase. The dye seems to act primarily via inhibition of NO synthase, with enzyme-bound heme being a possible target in its inhibitory action.

Amino Acid Oxidoreductases↗
PubMed1993

Characterization of endothelial cell amino acid transport systems involved in the actions of nitric oxide synthase inhibitors.

N omega-Substituted analogues of L-arginine have proven useful as specific inhibitors of nitric oxide formation in various biological systems. In the present study we describe the characteristics of amino acid transporters that mediate uptake of N omega-methyl-L-arginine (L-NMA) and N omega-nitro-L-arginine (L-NNA) into cultured porcine aortic endothelial cells. The transport of L-[14C]NMA showed biphasic kinetics, with Km values of 4 and 368 microM, and was inhibited by L-arginine, L-homoarginine, L-lysine, and L-ornithine but not by L-leucine or L-isoleucine. Similar transport kinetics (Km values of 6 and 609 microM) and substrate specificities were obtained for L-[3H]arginine uptake, indicating that L-arginine and L-NMA are transported by the same system. In contrast to L-arginine and L-NMA transport, uptake of L-[3H]NNA was monophasic (Km = 617 microM) and was inhibited by L-leucine and L-isoleucine but not by L-arginine, L-homoarginine, L-NMA, L-lysine, or L-ornithine. Uptake studies with L-[3H]leucine revealed that the transport of this amino acid occurred in a manner very similar to that of L-[3H]NNA transport, suggesting that the uptake of both compounds may be mediated by the same system. In additional experiments, we determined the effects of L-NMA and L-NNA on the A23187-induced accumulation of intracellular cGMP, to establish to what extent these transport systems are involved in the actions of nitric oxide synthase inhibitors. L-Lysine and L-ornithine, which both inhibited L-NMA uptake, increased the IC50 of L-NMA from 7.8 microM to 57 microM but did not reduce the inhibitory effects of L-NNA. In the presence of L-leucine or L-isoleucine, however, which both inhibited L-NNA uptake, the IC50 of L-NNA was increased from 1.2 microM to 37 microM but the inhibitory actions of L-NMA remained unaffected. These data demonstrate that the endothelial transport systems for L-arginine and L-leucine mediate the biological effects of L-NMA and L-NNA, respectively.

Amino Acid Oxidoreductases↗
PubMed1993

Changes in the frontal sinuses of the weasel (Mustela nivalis) in Poland possibly caused by nematodes or trematodes.

The presence of parasitic Nematodes was determined by visual assessment of damage to the 271 weasel skulls (154 males and 117 females). The damages were attributed to Skrjabingylus nasicola (Leuckart, 1842) or Troglotrema acutum (Leuckart, 1842) (on the basis of it's appearance and relevant papers). The frequency of infestation by both parasites was 38%. It was higher in females and also increased along with age. A significant dependence between skull length of adult specimens and infestation rate was found.

Animals↗
PubMed1993

[Reporting and evaluation and a blood alcohol laboratory--automated by using a new databank system].

For the processing and evaluation of a large number of data obtained in the course of blood alcohol determination the use of computerized systems is essential. By applying the software Skylight a data management program was developed under Windows. After entering the case data (name of the person involved, type of offence, time of accident and of blood sampling etc.) and BAC-values the evaluation of the data is automatically performed, e.g. calculation of the mean value, BACs at the time of event (minimum and maximum value). The user is guided by screen menus, which facilitate the entry of data. Reports are created by using predesigned formats retrieved from a report file. The data base system is installed on a local network. Access is permitted by password only to guarantee optimal security of the data.

Accidents, Traffic↗
PubMed1992

Increases in endothelial cyclic AMP levels amplify agonist-induced formation of endothelium-derived relaxing factor (EDRF).

The interaction between intracellular cyclic AMP and agonist-induced endothelium-derived relaxing factor (EDRF) (NO) formation was investigated in pig aortic endothelial cells. Three potent stimulators of adenylate cyclase, namely forskolin, adenosine and isoprenaline, amplified bradykinin- and ATP-induced biosynthesis and release of EDRF. None of the substances by itself affected basal EDRF formation. The effects of forskolin, adenosine and isoprenaline corresponded to an enhanced agonist-induced rise in intracellular free Ca2+ concentration ([Ca2+]i), were mimicked by the membrane-permeable cyclic AMP analogue dibutyryl cyclic AMP and were antagonized by the protein kinase inhibitor N-[2-(methylamino)ethyl]-5-isoquinolinesulphonamide dihydrochloride (H-8). Our data suggest that cyclic AMP-dependent phosphorylation modulates Ca(2+)-signalling and thus the function of endothelial cells. This mechanism may be of particular physiological importance, since it allows a joint regulation of endothelial functions by tissues factors such as bradykinin, which directly affects [Ca2+]i and agonists which affect intracellular cyclic AMP levels.

Adenosine↗
PubMed1992

Brain nitric oxide synthase is a haemoprotein.

Brain nitric oxide (NO) synthase showed pyridine haemochrome spectra typical of ferroprotoporphyrin IX-containing enzymes. The haem content of purified NO synthase was in the range 0.7-0.9 mol/mol of 160 kDa subunit. In the presence of CO, NO, KCN and miconazole, the L-citrulline-forming activity of NO synthase was markedly diminished, demonstrating that enzyme-bound haem is involved in enzymic NO synthesis.

Amino Acid Oxidoreductases↗