Biomedical subjects

H Heinle

Publications and source records attributed to H Heinle.

46 records · Page 3Linked to original sources

PubMed1976

[The biosynthesis of glutathione in human erythrocytes (author's transl)].

The concentrations of glutathione precursors in human erythrocytes were investigated. 300muM glutamate, 375 muM glycine, and 10muM cysteine were found by automated amino acid analysis. The concentration of 2-aminobutyrate, the precursor of ophthalmic acid, was 15muM. The influence of the activities of endogenous or added glutamyl-cysteine synthetase and glutathione synthetase on the rate of glutathione biosynthesis was measured in membrane-free hemolysates under physiological conditions. The results show that the rate of the overall biosynthesis mainly depends on the formation of the dipeptide glutamyl-cysteine. The effect of glutathione precursor concentrations on the synthesis of the tripeptide was investigated at constant (endogenous) activities of the synthesizing enzymes. The rate was not enhanced by addition of glutamate and/or glycine unless cysteine or glutamyl-cysteine was also added. It is concluded that the concentration of cysteine limits the actual rate of the glutamyl-cysteine-synthetase reaction in vivo. No cysteine or bis(glutamyl)cystine was detected in human hemolysate; however, these disulfides were converted to glutathione. This indicates that erythrocytes have an appropriate system for their reduction, since the disulfides themselves are not substrates for the glutathione-synthesizing enzymes. Studies with intact human red cells indicate that the uptake of cysteine is the rate-determining step in the biosynthesis of glutathione.

Amino Acids↗

PubMed1976

[Does a modified gamma-glutamyl cycle exist in human erythrocytes (author's transl)].

The first step in the biosynthesis of glutathione is the formation of gamma-glutamyl-cysteine by the enzyme glutamyl-cysteine synthetase. Since this enzyme is not specific for cysteine, different gamma-glutamylamino acids may be formed in vivo which represent potential substrates for the enzymes gamma-glutamylcyclotransferase; in this way 5-oxo-L-proline and free amino acid are formed. We investigated in membrane-free hemolysate the competition between the biosynthesis of glutathione or ophthalmic acid and the degradation of gamma-glutamyl peptides by measuring the formation of 5-oxoproline. The endogenous rate of 5-oxoproline production was 0.13 muM/min. This increased to 2muM/min after addition of 2-aminobutyrate, and to 10muM/min after addition of glutamate and 2-aminobutyrate to hemolysate. Addition of cysteine resulted in an increased oxoproline production only under conditions where glutamyl-cysteine accumulated. In addition, it was shown that for glutamyl-2-aminobutyrate the degradation to 5-oxoproline is faster than the utilization for the tripeptide synthesis. This was not the case for glutamyl-cysteine. Since membrane-free hemolysate (which lacks gamma-glutamyltransferase) is able to produce 5-oxoproline starting from glutamate, it is concluded that this 5-oxoprolinent amino acid transport via a modified gamma-glutamyl cycle.

Aminobutyrates↗

PubMed1975

[Biosynthesis of glutathione. VI. Ocurrence and level of gamma-glutamylcysteine in human red cells (author's transl)].

The concentration of the glutathione precursor gamma-glutamylcysteine in hemolysates of human red cells was determined using three different methods. a) The [14C]N-ethylmaleimide derivatives of the intracellular sulfhydryl compounds were separated on a cation exchange column. The amount of [14C]N-ethylmaleimide glutamycysteine was radiochemically determined. b) With purified glutathione synthetase, an enzymatic endpoint metabolite determination was performed with [14C]glycine as one substrate. The amount of labelled glutathione formed from the gamma-glutamylcysteine present was measured radiochemically.

Cysteine↗

PubMedDate not supplied

Luminol-enhanced chemiluminescence after reaction of hydroperoxides with opsonized zymosan.

Luminol-enhanced chemiluminescence (LEC) is very sensitive in detecting free radicals but relatively insensitive for hydroperoxides (hydrogen peroxide, tert-butyl hydroperoxide). However, in the presence of opsonized zymosan (often used for stimulation of phagocytic cells) hydroperoxides also induce LEC, suggesting that free radicals are produced under these conditions. Therefore careful interpretation with respect to the nature of the reactive species is necessary when LEC is used for characterization of zymosan-induced phagocytosis. We studied the properties of zymosan-induced LEC under different test conditions and with various inhibitors. Typical radical scavengers, e.g. nordihydroguaiaretic acid and superoxide dismutase, are strong inhibitors, indicating the importance of the superoxide anion. This system is useful for drug testing with respect to antioxidative or radical scavenging activity.

Antioxidants↗

PubMedDate not supplied

Metabolic changes in modified smooth muscle cells of rabbit carotid arteries.

In the common carotid artery of rabbits, intimal myocyte proliferations were induced by daily repeated local electrical stimulation of the vessel wall in combination with a cholesterol containing diet given for 4 weeks. Some biochemical parameters of the morphologically modified intimal smooth muscle cells were studied and compared with those of samples obtained from nonstimulated tunica media of the contralateral carotid artery. The results show that in relation to dry weight both alkali extractable protein and DNA content of the proliferates are increased to about 125%. In the proliferates, the in-vivo tissue concentrations of glucose and glycogen are only 50-70% of the normal values, whereas the concentration of lactate is increased to about 160%. In-vitro incubation experiments of excised tissues samples from the intimal proliferates and normal media indicate that under an optimal supply of substrates the glucose uptake and lactate production of the proliferates are increased to 140% and 150%, respectively. This results provides evidence for an increased capacity of glycolysis in the proliferates, which in vivo may lead to a decrease in glucose concentration and to an increased concentration of lactate. This investigation shows that modified smooth muscle cells proliferating in the arterial intima exhibit an activated metabolism as seen in other models of arteriosclerosis.

Animals↗

PubMedDate not supplied

Significance of glucose for mechanical activity, flavin and pyridine nucleotide oxidation-reduction changes in isolated rat portal veins under ACh-stimulation.

The influence of extracellular glucose on isometric concentrations induced by acetylcholine was investigated under aerobic conditions in rat portal veins. In order to estimate the contractile potency of acetylcholine (ACh), dose-response curves of ACh and nor-epinephrine (NE) were compared. Simultaneously with the mechanical activity, changes in the redox states of pyridine nucleotides (PN) and flavoproteins were detected by a surface-fluorimetric method. Tissue concentrations of glucose, lactate, pyruvate, and ATP were determined parallel to ACh-induced contractions at corresponding intervals. In response to ACh-induced contractions, PN and flavoproteins were oxidized in vessels supplied with glucose. Following 6 h of glucose withdrawal, a small reduction or no change in the redox state was observed; ACh-induced contractions and tissue concentrations of glucose, lactate, pyruvate, and ATP were decreased. When glucose was added, the contraction force was restituted, and the tissue concentrations of glucose, lactate and PN were increased again. The ATP concentration remained decreased. The results suggest that isometric contractions of portal veins are significantly influenced by aerobic glycolysis. A direct effect of glucose on the excitability of vascular smooth muscle is discussed.

Acetylcholine↗

PubMedDate not supplied

Metabolic effects of direct current stimulation on cultured vascular smooth muscle cells.

Vascular smooth muscle cells from rabbit arteries were grown in tissue culture and stimulated by DC impulses (1 mA, 1 V, 10 Hz, 1 ms/imp). Scanning microscopic examination disclosed that in stimulated cultures the cell surface was enlarged by numerous microvilli. This was interpreted as being indicative of an increase in cell activity. Cellular metabolism was characterized by analyzing the incubation medium for glucose, glutamate/glutamine, and lactate. When compared to unstimulated controls, stimulation caused an increase in the uptake of glucose and glutamine as well as an increased lactate production. The enhancing effect on metabolism was prevented when the "calcium antagonist" verapamil was present (5 X 10(-6) M). Although the exact mechanism by which DC stimulation influences the cells remains obscure, this finding indicates an important mediating role of Ca2+ ions.

Animals↗

PubMedDate not supplied

Oxidation of plasmalogens produces highly effective modulators of macrophage function.

Model derivatives of plasmalogens and chemically synthesized oxidative degradation products as found e.g. during oxidation of low density lipoproteins show strong effects on phagocytosis induced secretion of reactive oxygen species of macrophages which was measured by luminol-enhanced chemiluminescence. Whereas a plasmalogen epoxide showed enhancing effects in submicromolar range, inhibition was found with higher concentrations as well as with alpha-hydroxyaldehydes. The substances showed only little effects on the non-cellular ROS-dependent chemiluminescence of the reaction between hydrogen peroxide and opsonized zymosan and no cytotoxic effects under the assay conditions used. These results show that oxidative modification and degradation of plasmalogens occuring also under pathophysiological situations in vivo produces effective modulators of macrophage function which could be important; e.g. during inflammation or atherogenesis.

Animals↗

PubMedDate not supplied

Increased contractile responses of isolated arteriosclerotic rabbit carotid arteries to various vasoactive stimuli.

Clinical observations indicate that atherosclerotic vessels are prone to develop vasospasm. It was assumed that this property is related to hypercholesterolemia, but the basic mechanisms are still unknown. To investigate further mechanical stimulation experiments were performed with segments of rabbit carotid arteries. Arteriosclerotic lesions were induced in vivo in these arteries by application of DC impulses either in hyper- or normo- cholesterolemic animals. Contractions were evoked by noradrenaline, KCl or hydrogen peroxide. The results show that all the arteriosclerotic segments were hypersensitive to noradrenaline and hydrogen peroxide, compared with the corresponding controls. An increase in contraction force was also found upon application of KCl to lipid-containing plaques. However, in arteriosclerotic segments obtained from normally fed animals, the contraction response to KCl was lower than in controls. This indicates that complex alterations in the contractile properties occur in smooth muscle during atherogenesis which cannot be explained solely on the basis of the influence of cholesterol on the arterial cells.

Animals↗

PubMedDate not supplied

The degradation of glutathione derivatives in the rat kidney.

Single proximal rat kidney tubules and Henle loops were microperfused with labelled S-substituted glutathione derivatives which are physiological mercapturic acid precursors. S-methyl-glutathione as well as the non-permeating bromo-sulfaleinglutathione adduct were degraded in the proximal convolution with a half-life of about 3.5 sec. The findings demonstrate that the brush-border membrane-bound renal gamma-glutamyl transpeptidase acts on luminal substrates and is involved in mercapturic acid synthesis and splitting of extracellular glutathione.

Animals↗