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Biomedical subjects

H Kühn

Publications and source records attributed to H Kühn.

At least 127 records · Page 7Linked to original sources
PubMed1990

Temporal relationship between force, ATPase activity, and myosin phosphorylation during a contraction/relaxation cycle in a skinned smooth muscle.

The temporal relationship between myosin phosphorylation, contractile force and ATPase activity was studied in skinned preparations from the guinea-pig Taenia coli. When free Calcium concentration [( Ca2+]) was increased from pCa (-log[Ca2+]) 9 to pCa 4.5 at low calmodulin concentration (0.05 microM), ATPase activity and myosin light-chain phosphorylation rose quickly, while the increase in force and stiffness was delayed. The time-course of tension increase was faster at higher calmodulin concentrations (5 microM), although the maximal level of phosphorylation was unchanged. Lowering the calcium concentration from pCa 4.5 to pCa 9 at the plateau of contraction caused a rapid decrease in ATPase activity and in myosin phosphorylation, while force and stiffness decayed more slowly. The force decay could be accelerated by inorganic phosphate. These results suggest that, during contraction, force may be produced actively by phosphorylated and ATP-splitting crossbridges, but may be maintained by dephosphorylated crossbridges which cycle slowly. However, force could also be modulated by calmodulin and inorganic phosphate in a manner not involving an alteration in the extent of myosin phosphorylation.

Adenosine Triphosphatases↗
PubMed1990

Occurrence of 9- and 13-keto-octadecadienoic acid in biological membranes oxygenated by the reticulocyte lipoxygenase.

Membranes of intact rabbit reticulocytes and rat liver mitochondrial membranes oxygenated by the pure reticulocyte lipoxygenase contain 13-keto-9Z,11E-octadecadienoic acid and 9-keto-10E,12Z-octadecadienoic acid. In mitochondrial membranes not treated with lipoxygenase and in rabbit erythrocyte membranes these products were not detected. The chemical structure of the compounds has been identified by cochromatography with authentic standards on various types of HPLC columns, by uv and ir spectroscopy and GC/MS. In the membranes of rabbit reticulocytes up to 2% of the linoleate residues are present as its 9- and 13-keto derivatives. Most of the keto compounds (up to 90%) are esterified in the membrane ester lipids, only about 10% were found in the free fatty acid fraction. It is proposed that the keto dienoic fatty acids are formed via decomposition of hydroperoxy polyenoic fatty acids originating from the oxygenation of the membrane lipids by the reticulocyte lipoxygenase.

Animals↗
PubMed1990

Metabolism of polyenoic fatty acids by rabbit reticulocytes. Intracellular action of the erythroid lipoxygenase on membrane lipids.

Rabbit reticulocytes metabolize exogenous polyenoic fatty acids via three different pathways. (i) incorporation into the cellular ester lipids, predominantly into phospholipids, (ii) beta-oxidation forming CO2 and (iii) lipoxygenase reaction. The lipoxygenase pathway contributes to about 30% to the metabolism of exogenously added linoleic acid. The endogenous substrates of the lipoxygenase pathway are not only the free polyenoic fatty acids bound to the cellular membranes but also the membrane phospholipids. The lipoxygenase products detected in the membrane lipids have been isolated and their complete chemical structure has been identified as 13-hydroxy-9Z,11E-octadecadienoic acid. 9-hydroxy-10E,12Z-octadecadienoic acid, their all E isomers and 15-hydroxy-5Z,8Z,11Z,13E-eicosatetraenoic acid. Subcellular fractionation of different cellular membranes indicated that these products occur in both the mitochondrial membranes and the plasma membrane. By quantitative HPLC analysis it has been shown that the mitochondrial membranes contain about 3 times more oxygenated fatty acids than the plasma membranes; one out of ten linoleic acid residues in the mitochondrial membranes is present as hydroxylated derivative. The pattern of lipoxygenase products detected in the mitochondrial membranes was much more unspecific than that of plasma membranes. These data are discussed in the light of the involvement of the lipoxygenase pathway in the degradation of mitochondria during the maturation of red blood cells.

Animals↗
PubMed1990

Oxygenation of biological membranes by the reticulocyte lipoxygenase. Lack of stoichiometry between oxygen uptake and product formation.

The oxygenation of different types of biological membranes (rat liver mitochondria, rat liver endoplasmic membranes, inside-out erythrocyte ghosts, right side-out erythrocyte ghosts) was studied with respect to products formed during the reaction. In all cases a very similar product pattern was observed with 15S-hydroperoxy-5Z.8Z.11Z.13E-eicosatetraenoic acid (15-HETE) and 13S-hydroperoxy-9Z,11E-octadecadienoic acid, (13-HODE) being the major products. Comparison of the amount of lipoxygenase products formed with the oxygen uptake measured during the reaction indicated an excessive oxygen uptake. With mitochondrial membranes the oxygen consumption was almost one order of magnitude higher than the amount of the products detected. The origin of the excessive oxygen uptake remains unclear. These data, however, indicate that the oxygen consumption with complex substrates is not a reliable measure for the lipoxygenase activity.

Animals↗
PubMed1990

Oxygenation of mitochondrial membranes by the erythroid lipoxygenase. Consequences for membrane properties.

The reticulocyte lipoxygenase is able to oxygenate mitochondrial membranes. The main products formed during this reaction are 15S-hydroperoxy-5Z,8Z,11Z,13E-eicosatetraenoic acid (15-HETE) and 13S-hydroperoxy-9Z,11E-octadecadienoic acid (13-HODE). The oxygenation of mitochondrial membranes is accompanied by an inactivation of enzymes localized in the outer and inner mitochondrial membranes, by a drastic change of the passive electric properties and by a destruction of the iron-sulfur clusters of the outer mitochondrial membrane. The possible mechanism of the processes leading to these functional changes is discussed.

Animals↗
PubMed1990

Formation of oxygenase and hydroperoxidase products by the pure reticulocyte lipoxygenase.

The pure reticulocyte lipoxygenase oxygenates free arachidonic acid to 15S- and 12S-HPETE in a ratio of about 10:1 15S-H(P)ETE or its methyl ester can be further converted by the lipoxygenase via three different types of reactions: (i) oxygenase reaction, (ii) hydroperoxidase reaction and (iii) leukotriene synthase reaction. Here we summarize the products formed from arachidonic acid by the reticulocyte lipoxygenase via these three types of reactions.

Animals↗
PubMed1990

The linoleic acid metabolite 9DS-hydroxy-10,12(E,Z)-octadecadienoic acid is a strong proinflammatory mediator in an experimental wound healing model of the rat.

Locally administered 9Ds-hydroxy-10,12(E,Z)-octadecadienoic acid (9-HODE) caused a drastic inflammatory response in the experimental model of granulation tissue formation of the rat according to RUDAS (Arzneimittelforsch. 10,226-229, 1960). Three days after implantation of the polyvinyl chloride rings the granulation tissue became inhomogeneous with proliferation islets surrounded by edematous regions containing a diminished number of cells. The number of polymorphonuclear leukocytes and of macrophages was greatly enhanced in the whole tissue, whereas the number of lymphocytes was reduced. After seven days the whole granulation tissue was loosened, and its mass was twice as high as in the control animals. The number of fibroblasts per area unit and the hydroxyproline content were diminished. Linoleic acid and 13Ls-hydroxy-9,11(Z,E)-octadecadienoic acid (13-HODE) caused also some changes in the formation of granulation tissue, but in a different manner, in particular, without accumulation of polymorphonuclear leukocytes and macrophages, indicating the specificity of the effect of 9-HODE. The recruitment of leukocytes was not due to a direct chemotactic action of 9-HODE as shown in an agarose diffusion test comparing the effects of 9-HODE and leukotriene B4. The possible biological importance of the proinflammatory effect of 9-HODE is discussed.

Animals↗
PubMed1990

The erythroid arachidonate 15-lipoxygenase in rat reticulocytes.

Rat reticulocytes contain an arachidonate 15-lipoxygenase which attacks submitochondrial particles in an identical way as the corresponding enzyme from rabbit reticulocytes. It shows immunological cross-reactivity with a polyclonal antiserum against the rabbit reticulocyte lipoxygenase. It differs from the rabbit enzyme with respect to some kinetic properties and its isoelectric point.

Animals↗
PubMed1989

Occurrence of free and esterified lipoxygenase products in leaves of Glechoma hederacea L. and other Labiatae.

Leaves of Glechoma hederacea L. and other Labiatae contain (9S,10E,12Z,15Z)-9-hydroxy-10,12,15-octadecatrienoic acid, (10E,12Z,15Z)-9-oxo-10,12,15-octadecatrienoic acid, (9S,10E,12Z)-9-hydroxy-10,12-octadecadienoic acid and (10E,12Z)-9-oxo-10,12-octadecadienoic acid in a ratio of 71/14/12/3 (by mass), predominantly esterified in the membrane ester lipids. The leaves contain the highest level of these products, whereas only small amounts were found in the stalk and the roots. The chemical structures of these compounds were established by ultraviolet and infrared spectroscopy, by co-chromatography with authentic standards on various types of HPLC columns including chiral-phase HPLC and gas chromatography/mass spectrometry. The stereochemical specificity indicates the enzymatic origin of the products, most probably via a lipoxygenase reaction. Freshly harvested specimens of G. hederacea L. contain only small amounts of hydroxy-polyenoic fatty acids. Air-drying causes a strong increase in the content of free and esterified (9S,10E,12Z,15Z)-9-hydroxy-10,12,15-octadecatrienoic acid. Up to 80% of the hydroxy fatty acids of the total lipid extracts were esterified in the cellular lipids. The data presented indicate that lipoxygenase products occur in the cellular ester lipids of G. hederacea L. and other Labiatae. The results are discussed in the light of a possible involvement of the lipoxygenase pathway in the natural senescence of leaves.

Chemical Phenomena↗
PubMed1989

Oxygenation of mitochondrial membranes by the reticulocyte lipoxygenase. Action on monoamine oxidase activities A and B.

Incubation of isolated rat liver mitochondria with the pure rabbit reticulocyte lipoxygenase caused a time-dependent inactivation of the monoamine oxidase activities A and B. Furthermore, a conversion of the monoamine oxidase into a diamine oxidase was observed. The inactivation kinetics for both monoamine oxidase activities A and B showed a biphasic behaviour; a reversible short-term inhibition during the first 5 min of incubation was followed by an irreversible inactivation of the enzyme. The kinetic studies suggest that the slow irreversible inactivation of the monoamine oxidase activities is due to secondary reactions subsequent to the initial attack of the lipoxygenase on the mitochondrial outer membrane. During the interaction of the lipoxygenase with the mitochondria, only about 1.5% of the polyenoic fatty acids present in the mitochondrial membranes were oxygenated. The predominant products formed during the interaction of the lipoxygenase with the mitochondrial membranes are (13S)-hydro(pero)xy-9Z,11E-octadecadienoic acid and (15S)-hydro(pero)xy-5,8,11,13(Z,Z,Z,E)-eicosatetraenoic acid.

Animals↗
PubMed1989

Occurrence of the erythroid cell specific arachidonate 15-lipoxygenase in human reticulocytes.

Human reticulocytes obtained from patients suffering from various haemolytic disorders convert exogenous [1-14C]-arachidonic acid to 15-hydroxy-5,8,11,13(Z,Z,Z,E)-eicosatetraenoic acid (15-HETE). Immunological studies (dot blot, Western blot) indicated that human reticulocytes contain a lipoxygenase which cross-reacts with a polyclonal antiserum against the rabbit reticulocyte lipoxygenase. Northern blotting with a cloned lipoxygenase cDNA probe shows that the specific mRNA is also present. Reaction of the lipoxygenase with submitochondrial particles caused inactivation of respiratory enzymes. The occurrence of an erythroid cell specific lipoxygenase of similar type in reticulocytes of various mammals and man suggests the general role of this enzyme in the maturational degradation of mitochondria.

Anemia↗
PubMed1989

Kinetics, binding constant, and activation energy of the 48-kDa protein-rhodopsin complex by extra-metarhodopsin II.

We have found that the 48-kDa protein (or S-antigen 48k) of the rod photoreceptor enhances the light-induced formation of the photoproduct metarhodopsin II (MII) from prephosphorylated rhodopsin. The effect is analogous to the known enhancement of MII (extra-MII) that results from selective interaction of MII with G-protein. We have determined some parameters of the MII-48k interaction by measuring the extra-MII absorption change induced by the 48-kDa protein. The amplitude saturation yields a dissociation constant for the MII-48k complex on the order of 50 nM. At the technical limit of these measurements, 13.7 degrees C and 12 microM 48-kDa protein, we find a rate of 2.3 s-1 for formation of the 48k-MII complex. Extrapolation of these values to cellular conditions yields an occupation time of phosphorylated MII by 48k less than 200 ms. This is short compared to estimated rates of phosphorylation. The temperature dependence of the MII-48k formation rate is very high (Q10 for 5 degrees C/15 degrees C = 9-10). The related Arrhenius activation energy (165 kJ mol-1) is correspondingly high and indicates a considerable transient chemical change during the binding process.

Animals↗
PubMed1989

[Diagnosis of salmonellosis].

The determination of clonal identity of bacterial isolates of Salmonella, in particular epidemiological important serovars of Salmonella, is an unequivocal prerequisite for the study of the epidemic process of salmonellosis. This is demonstrated for the serovar S. typhimurium by means of classical as well as molecular methods for epidemiological subdifferentiation.

Animals↗
PubMed1989

[Epidemiology of salmonellosis].

The diagnostically and in regard to surveillance rather comprehensively recorded salmonellosis bear the character of a model of zoonotic diarrhoeal diseases. Whereas those serovars predominantly adapted to human beings or animal species are without any importance in this connection, such serovars with own epizootic processes in different animal species with or without causing illness are able to be transferred to human beings via foods of animal origin and to act as etiological agents of gastroenteritis. Some strains of them equipped by special properties often encoded by plasmids can be transferred from man to man, too. In GDR in 1987 S. enteritidis (34.0%), S. typhimurium (17.9%), S. manhattan (7.7%) and S. agona (7.4%) amounted to two thirds of all the salmonella isolates in connection with human enteritis. For analyzing epizootic-epidemic processes in the case of rare serovars the identity of serovars is sufficient in first approximation, in the case of frequently isolated serovars the complex typing of the concerned strains by using epidemiological laboratory methods is required.

Animals↗
PubMed1988

Lipoxygenase catalyzed oxygenation of hydroxy fatty acids to lipoxins.

The pure lipoxygenases from rabbit reticulocytes and soybeans convert a variety of substrates (arachidonic acid, 15-HPETE, 15-HETE, 5-HETE, various DiHETE isomers) to trihydroxy eicosanoids containing a conjugated tetraene system (lipoxins). In general, the methyl esters are better substrates for lipoxin formation than are the free acids. Lipoxygenase inhibitors (5,8,11,14-eicosatetraynoic acid, nordihydroguaiaretic acid) strongly inhibit the lipoxin formation. The complete stereochemistry of the lipoxin B formed from 15S-HETE methyl ester has been established by co-chromatography with authentic standards on various types of HPLC columns, by GC/MS analysis, by gas liquid chromatography of the ozonolysis fragments of the menthoxy carbonyl derivatives and 1H-NMR studies. The molar absorption coefficient of the conjugated tetraenes was measured as epsilon 301 = 53,000. The lipoxins formed from 15-HETE and various DiHETE isomers are formed exclusively via the oxygenation pathway as shown by experiments under an 17O2 atmosphere and/or by anaerobic incubations. Our results indicate that lipoxins can be synthesized via lipoxygenase-catalyzed sequential oxygenation of polyenoic fatty acids and their hydro(pero)xy derivatives.

Animals↗