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Biomedical subjects

A Bast

Publications and source records attributed to A Bast.

At least 19 recordsLinked to original sources
PubMed1992

Atypical molecular pharmacology of a new long-acting beta 2-adrenoceptor agonist, TA 2005.

The molecular pharmacology of a new putative long-acting bronchodilator TA 2005 (8-hydroxy-5-[(1R)-1-hydroxy-2-[N-[(1R)-2-(p-methoxy-phenyl)- 1-methylethyl]amino]ethyl]carbostyril hydrochloride) has been compared with that of the reference compounds isoprenaline and salbutamol in both methacholine (3 x 10(-6) M) precontracted guinea pig tracheal smooth muscle relaxation and in bovine trapezium muscle binding experiments. TA 2005 appeared very potent compared with isoprenaline and salbutamol (pD2 values of 9.29 vs. 7.65 and 7.10 respectively). For isoprenaline and salbutamol a shallow displacement curve was observed and addition of the non-hydrolysable GTP analogue guanylyl-imidodiphosphate (GppNHp) gave a rightward shift (pKd,high and pKd,low values of 7.3 and 6.1 vs. 7.0 and 5.4 respectively). For TA 2005 a steep displacement curve was found with only one binding state even without GppNHp (pKd,high value of 8.2). The long duration of action of TA 2005 might be explained by tight binding of this compound to the beta 2-adrenoceptor. The extent of tight binding for TA 2005 was extremely large. The molecular basis of the tight agonist binding phenomenon for TA 2005 seems to be of different origin than for isoprenaline. It is hypothesized that a different mechanism of activation of the beta 2-adrenoceptor may be involved for TA 2005.

Adrenergic beta-Agonists↗
PubMed1992

The first radiolabeled histamine H3 receptor antagonist, [125I]iodophenpropit: saturable and reversible binding to rat cortex membranes.

We describe the binding to rat cortex membranes of [125I]iodophenpropit, the first radiolabeled histamine H3 antagonist. The binding of [125I]iodophenpropit is selective, saturable, readily reversible, and of high affinity (KD 0.32 nM; Bmax 209 fmol/mg of protein). Specific binding at a concentration of 0.3 nM accounted for 45-55% of the total binding. [125I]Iodophenpropit seems to fulfill the criteria for a suitable ligand for histamine H3 receptor binding studies.

Animals↗
PubMed1992

Fully automated determination of a new anthracycline N-l-leucyldoxorubicin and six metabolites in plasma by high-performance liquid chromatography with on-line sample handling.

N-l-Leucyldoxorubicin (Leu-Dox) was developed as a prodrug of doxorubicin (Dox) in order to diminish the cardiotoxic side-effect associated with repeated anthracycline treatment. To study the pharmacokinetics of Leu-Dox, Dox and other metabolites a sensitive and selective assay was needed. Leu-Dox and six of its known metabolites were extracted from plasma using an in-line reversed-phase precolumn (40-50 microns C8 particles). The trapped analytes were subsequently flushed to the analytical column (3 microns C18) using 0.5 ml of phosphate buffer (pH 3.5)-acetonitrile (2:1, v/v), which also served as the isocratic mobile phase. Within 12 min, a clean baseline-resolved chromatogram is obtained by fluorescence detection. Recoveries were almost quantitative and highly reproducible, with standard deviations less than or equal to 5.4% and less than or equal to 2.7% at spiked concentrations of 10 and 100 nM. Using 300 microliters of plasma, detection limits ranged from 0.3 to 0.8 nM at a signal-to-noise ratio of 3. The calibration curves were linear from 1 to 300 nM (r2 greater than or equal to 0.999) for each of the seven compounds. The between-day accuracy was in the range 91-99% and 99-105% at 10 and 100 nM, respectively, with standard deviations of 1-4%. Application of the assay to the analysis of plasma from patients after administration of Leu-Dox proved successful.

Antibiotics, Antineoplastic↗
PubMed1992

Desensitization of histamine H1 receptor-mediated cyclic GMP production in guinea-pig lung.

Histamine H1 receptor-mediated production of cGMP in guinea-pig lung tissue becomes rapidly desensitized after previous exposure to histamine. This desensitization is clearly concentration dependent and appears to be homologous. Responses to histamine are also inhibited by previous treatment with phorbol 12,13-dibutyrate. Yet, the time course of the inhibition is considerably slower and the maximal inhibition is significantly less compared to receptor desensitization. Moreover, the effects of the phorbol ester are not confined to H1 receptor responses. Since the effects of receptor desensitization are also not prevented by several protein kinase C inhibitors, the development of homologous H1 receptor desensitization is not dependent upon protein kinase C activation, but is caused by a yet unidentified mechanism.

Animals↗
PubMed1992

Analysis and pharmacokinetics of N-l-leucyldoxorubucin and metabolites in tissues of tumor-bearing BALB/c mice.

Leucyldoxorubicin (Leu-Dox) was developed as a prodrug of doxorubicin (Dox) with the aim of lowering the cardiotoxicity and improving the therapeutic index produced by Dox. To support the preclinical findings on its antitumor activity and cardiotoxicity, concentrations of Leu-Dox and its metabolites were determined in plasma, heart, and tumor after the administration of Leu-Dox to tumor-bearing mice. A liquid-liquid extraction procedure employing a chloroform/2-propanol/dimethylsulfoxide (DMSO) mixture was developed. By means of high-performance liquid chromatography (HPLC) with fluorescence detection, Leu-Dox and six of its metabolites could be assayed in the tissues with high sensitivity. Detection limits ranged from 0.01 nmol/g tissue for the aglycons to 0.06 nmol/g for Dox. Recoveries were in the range of 82%-110%, and calibration curves were linear over the concentration range tested (0.1-10 nmol/g tissue, r > or = 0.998). Concentration versus time curves were constructed for plasma, heart, and tumor over the first 72 h, and areas under the curves (AUCs) for the first 48 h were determined by the trapezoidal rule. Dox was rapidly formed from Leu-Dox, reaching peak levels in plasma within 5 min and in tissues within 1 h after i.v. administration of Leu-Dox (12 mg/kg). The elimination of Leu-Dox was also fast as illustrated by final half-lives of 1.1, 0.8, and 0.9 h in the plasma, heart, and tumor, respectively. For Dox, the final half-lives were 16.7 h in plasma, 15.3 h in heart tissues, and 27.4 h in tumor tissues. AUC values determined for Leu-Dox and Dox were 221 and 51 nmol ml-1 min, 443 and 4,262 nmol g-1 min, and 153 and 1,466 nmol g-1 min in the plasma, heart, and tumor, respectively. Comparison of these values with those obtained after an equimolar dose of Dox indicated 26%, 30%, and 16% of Leu-Dox appeared as Dox in the plasma, heart, and tumor, respectively. Thus, not only is the plasma compartment not representative for calculations of the conversion of Leu-Dox into Dox in tissue, but differences in its appearance also exist between the tissue compartments. The AUC values found for Dox in the heart may explain the reduced cardiotoxicity elicited by Leu-Dox as compared with Dox; however, the values obtained for Dox in the insensitive murine colon tumor cannot explain the enhanced antitumor activity exerted by Leu-Dox in the sensitive human tumor xenografts in nude mice.

Animals↗
PubMed1992

Role of the epithelium in the control of intestinal motility: implications for intestinal damage after anoxia and reoxygenation.

A vibration technique was used to dislocate the epithelium from the rat small intestine, in order to study the possible regulatory role of the epithelium on intestinal motility. Complete removal of the epithelium led to a slightly potentiated contraction of the longitudinal smooth muscle by the muscarinic agonist methacholine (pD2. 6.5 +/- 0.1 vs. 6.2 +/- 0.2). The maximal beta-adrenergic response expressed relative to the relaxation by 0.5 mM dibutyryl cyclic AMP increased from 55.9 +/- 9.0% to 72.6 +/- 9.1% by this treatment. Efforts were made to relate these observations to the endothelium-dependent relaxation in blood vessels, but no indication was found for a similar mechanism in the small intestine. Not only mechanical dislocation can be employed to affect the mucosal layer, but also intestinal ischemia has been reported to lead to mucosal damage. In this study we mimicked ischemia by applying in vitro anoxia and subsequent reoxygenation to isolated intestinal segments. When intestinal segments are isolated and kept in physiological buffer, xanthine dehydrogenase is converted slowly to xanthine oxidase, irrespective of whether the buffer is oxygenated or not. No evidence was found for oxygen radical damage after anoxia and reoxygenation. However, the intestinal mucosa was damaged both after normoxia, and after anoxia and reoxygenation. Anoxia and subsequent reoxygenation did not affect muscarinic contraction, but slightly increased the beta-adrenergic relaxation, which partly correlates with the effects of mechanical dislocation of the epithelium. The increased sensitivity of the smooth muscle after epithelial damage might be involved in motility changes during intestinal inflammatory diseases.

Animals↗
PubMed1992

Monitoring of oxidative free radical damage in vivo: analytical aspects.

Free radical damage is an important factor in many pathological and toxicological processes. During the last decade a wide range of methods has been developed to determine free radical damage in various biological fluids and at various stages of development. This review offers an overview of the state of the art of monitoring free radical damage in vivo, with special emphasis on the analytical aspects of non-invasive methods.

Animals↗
PubMed1992

Effect of oxidative stress on receptors and signal transmission.

Reactive oxygen metabolites affect binding of ligands to membrane receptors and also coupling of receptors to G-proteins and effector enzymes. Peroxidation of membrane lipids may lead to a lowered receptor density and also will alter the viscosity of the plasma membrane, which affects receptor coupling. Reactive oxygen species may also interact with thiol/disulfide moieties on receptor proteins or on other factors in the receptor system, which is responsible for alterations in receptor binding or coupling. Moreover, lipid peroxidation is associated with the phospholipase A2 pathway, which might indirectly affect receptor function. Moreover, oxidative stress may lead to a disturbance in cellular Ca(2+)-homeostasis. This might be related to an effect on Ca(2+)-mobilizing receptors, but there is also evidence for a decreased Ca(2+)-sequestration by ATPases. In addition, peroxidation of membrane lipids increases membrane permeability to Ca2+. Finally, reactive oxygen species interfere with actions of nitric oxide, thus affecting another pharmacological messenger system.

Animals↗
PubMed1992

Glutathione mobilization during cerebral ischemia and reperfusion in the rat.

1. Cerebral ischemia applied for 15 min and followed by a 30 min reperfusion did not change the glutathione (GSH) levels and beta-adrenoceptor density (Bmax) in brain cortex. 2. A significant increase in erythrocyte-lysate GSH concentration (vs control) and a significant decrease of Bmax values in erythrocyte membranes (vs control) was found at the same time. 3. Pretreatment with the alpha-adrenoceptor antagonist phentolamine (5 mg/kg i.p.) prevented the erythrocyte GSH increase but not the decrease of Bmax value. Pretreatment with the beta-antagonist propranolol (2 mg/kg i.p.) did not influence the increase in erythrocyte GSH but circumvented the decrease of Bmax.

Animals↗
PubMed1992

Role of reactive oxygen species in intestinal diseases.

It is well known that reactive oxygen metabolites are generated during several pathologies, and that they are able to disturb many cellular processes and eventually lead to cellular injury. After intestinal ischemia, reactive oxygen species are produced when the ischemic tissue is reperfused. The enzyme xanthine oxidase is thought to play a key role in this process. As a result of this oxygen radical production, the permeability of the endothelium and the mucosa increases, allowing infiltration of inflammatory leukocytes into the ischemic area. Moreover, reactive oxygen species are also indirectly involved in leukocyte activation. In turn, these inflammatory cells respond with the production of oxygen radicals, which play an important role in the development of tissue injury. Thus, intestinal ischemia and reperfusion evokes an inflammatory response. Also during chronic intestinal inflammatory diseases, reactive oxygen metabolites are proposed to play an important role in the pathology. Scavenging of reactive oxygen species will thus be beneficial in these disorders.

Animals↗
PubMed1992

Short-term desensitization of the histamine H1 receptor in human HeLa cells: involvement of protein kinase C dependent and independent pathways.

1. In this study we have investigated the effects of short-term exposure of cells to histamine on the subsequent H1 receptor responsiveness in HeLa cells, using Ca2+ fluorescence microscopy and video digital imaging. 2. In HeLa cells, histamine (100 microM) induces an immediate H1 receptor-mediated biphasic elevation of the intracellular Ca2+ concentration ([Ca2+]i) (basal [Ca2+]i: 81 +/- 30 nM, histamine-induced Ca2+ response: first phase: 1135 +/- 79 nM; second phase: 601 +/- 52 nM, n = 11). 3. The histamine H1 receptors on HeLa cells are readily susceptible to desensitization since repetitive exposure of the same group of cells to histamine (100 microM) markedly affected the release and influx component of the induced Ca2+ response (second application of histamine: first phase: 590 +/- 92 nM, second phase: 279 +/- 47 nM; third application of histamine: first phase: 454 +/- 127 nM, second phase: 240 +/- 45 nM, n = 6). Video digital imaging revealed an increase in the lag time between stimulation and monitoring of the Ca2+ response and a reduced increase in [Ca2+]i after desensitization with histamine. 4. Neither the release component of the ATP response (50 microM) nor the caffeine (3 mM)-induced Ca2+ release were found to be affected by desensitization with 100 microM histamine. However, the second phase of the ATP response was significantly reduced after desensitization with histamine (control cells: 516 +/- 33 nM; desensitized cells: 331 +/- 96 nM, n = 4, P < 0.05).5. Activation of protein kinase C (PKC) by phorbol-12-myristate-1 3-acetate was found to inhibit the histamine as well as ATP-induced Ca2" response in a dose-dependent manner.6. In PKC downregulated cells the second phase of the histamine-induced Ca2+ response was significantly elevated, indicating the involvement of PKC in the negative feedback on the Ca2+ influx(control cells: second phase: 601 +/- 52 nM (n = 11); PKC downregulated cells: second phase:890 +/- 90nM, n = I0, P<0.05).7. Homologous desensitization of H, receptor responsiveness was still observed in PKC downregulated cells, implying the rapid activation of a regulatory mechanism other than PKC.8. Based on our experimental data we suggest that short-term desensitization of the histamine H,receptor evolves from two different processes: a selective reduction of the histamine-induced Ca2+ release, mediated by a PKC-independent pathway, and a non-selective inhibition of the receptormediated Ca2+ influx activated by a PKC-dependent pathway.

Adenosine Triphosphate↗
PubMed1992

Intestinal smooth muscle dysfunction after intraperitoneal injection of zymosan in the rat: are oxygen radicals involved?

Zymosan is frequently used as an activator of granulocytes to study inflammatory responses. We used zymosan as a model to understand the mechanisms involved in intestinal inflammatory diseases, and our special interest was focused on the smooth muscle function. Moreover, we investigated the role of oxidative stress in intestinal pathology after inflammatory processes. Intraperitoneal injection of zymosan induces a peritoneal inflammation, characterised by exudate in the peritoneum and peritoneal fibrosis. Three days after injection of zymosan (25-40 mg/100 g) we measured a decreased beta adrenergic smooth muscle response, while the muscarinic receptor-mediated contraction was not significantly affected. Efforts were made to correlate these observations with the development of oxidative stress; however, the intestinal glutathione balance remained undisturbed and no increase in lipid peroxidation products in the intestine was observed. Our conclusion is the peritoneal inflammation will lead to a release of various mediators, which may destroy receptor systems, among which are beta adrenoceptors. There was no evidence of an important role for reactive oxygen metabolites in this effect.

Animals↗
PubMed1992

A simple and rapid in vitro test system for the screening of histamine H3 ligands.

A simple and rapid functional test system for the screening of histamine H3 ligands is described. It is based on the inhibitory effect of histamine H3 agonists on electrically-evoked contractile response of isolated guinea pig intestine. Whole jejunum segments are continuously stimulated maximally (15 V) by electrical pulses with a frequency of 0.1 Hz and a duration of 0.5 msec. The resulting twitches are recorded isotonically (1.0 g) and can be completely abolished by atropine (0.1 mcM).

Acetylcholine↗
PubMed1991

Menadione inhibits the alpha 1-adrenergic receptor-mediated increase in cytosolic free calcium concentration in hepatocytes by inhibiting inositol 1,4,5-trisphosphate-dependent release of calcium from intracellular stores.

In order to establish the mechanism of perturbation of hormonally regulated calcium homeostasis in hepatocytes caused by menadione, the effects of menadione on hepatic alpha 1-adrenergic receptors and on alpha 1-adrenergic receptor-mediated increase in cytosolic free calcium concentration were determined. Menadione had no detectable effect on the alpha 1-adrenergic receptor but significantly inhibited (-)-epinephrine-dependent increases in intracellular free calcium concentration in Quin2 acetoxymethyl ester-loaded hepatocytes. The hormonally induced increase in intracellular free calcium concentration is caused by formation of inositol 1,4,5-trisphosphate (IP3) which binds to a specific receptor and causes a release of intracellular ATP-dependently sequestrated calcium. The IP3-stimulated release of calcium from intracellular pools in hepatocytes was inhibited to a great extent after treatment with menadione. This inhibition could also be observed after treatment of hepatocytes with p-benzoquinone and N-ethylmaleimide and could not be reversed by the thiol-reducing reagent dithiothreitol which indicated covalent binding to an essential free sulfhydryl group. The inhibition of IP3-dependent release of intracellular calcium was accompanied by a large increase in the number of detectable IP3 receptors without any change in the dissociation constant as determined in permeabilized hepatocytes. The increase in IP3 receptors caused by menadione could be reversed by dithiothreitol which suggests the involvement of free sulfhydryl groups. It is concluded that the IP3 receptor plays an important role in the mechanism of menadione-induced perturbation of hormonally regulated calcium homeostasis in rat hepatocytes.

Adrenergic alpha-Antagonists↗
PubMed1991

Oxidants and antioxidants: state of the art.

Reactive oxygen species are regarded as merely pernicious. This is incorrect for they play a pivotal role in many physiologic reactions, such as cytochrome P450-mediated oxidations, regulation of the tone of smooth muscle, and killing of microorganisms. An imbalance in oxidant-antioxidant activity is involved in many free radical-mediated pathologies, e.g., ischemia-reperfusion and asthma. In an attempt to alleviate these pathologies with antioxidants, it should be noted that these compounds are neither specific nor mere antioxidants. Associated with antioxidant activity is a pro-oxidant action. In the development of new antioxidant therapies, the important question of how these drugs are incorporated in or commensurate with existing integrated physiologic radical-defense systems should be addressed.

Animals↗
PubMed1991

Simple and sensitive quantification of anthracyclines in mouse atrial tissue using high-performance liquid chromatography and fluorescence detection.

Anthracyclines are very effective against soft tissue sarcomas, with cardiotoxicity being an important side effect after repeated administration. To estimate the relative cardiotoxicity of various anthracyclines and their metabolites, we developed an isolated mouse left atrium model. To relate an effect of doxorubicin, 4'-epidoxorubicin and their four main metabolites (doxorubicinol, epidoxorubicinol and the aglycons 7-deoxydoxorubicinon and 7-deoxydoxorubicinolon) to concentrations in the tissue instead of the incubation bath, a method of quantifying the anthracyclines in small tissue samples was developed. Atria were homogenized by sonication followed by extraction of the anthracyclines with methanol. The extract was directly analyzed by high-performance liquid chromatography with fluorescence detection. Recoveries for the six compounds tested ranged from 67.5% for 4'-epidoxorubicin to 100.6% for 7-deoxydoxorubinol aglycon with coefficients of variation of 2-3% at two spiked concentrations (0.1 and 1 nmol/mg of tissue). The calibration plots were linear (r2 greater than 0.996) over the concentration range tested (0.05-1 nmol/mg wet weight). The limits of detection (4-10 pmol/mg of tissue) were low enough to allow the determination of the anthracyclines at all relevant tissue concentrations.

Animals↗
PubMed1991

Histamine H1-receptor-mediated cyclic GMP production in guinea-pig lung tissue is an L-arginine-dependent process.

Histamine produces a rapid and massive increase of the c-GMP level of guinea-pig lung tissue. The EC50 value for this in vitro response is found to be 27 microM and the c-GMP level is maximally 9-fold elevated by 100 microM histamine. The response is stereoselectively inhibited by the enantiomers of chlorpheniramine, indicating H1-receptor involvement. Preincubation of lung tissue with 200 microM NCDC, a phospholipase C inhibitor, reduces the histamine (100 microM) responses to 16 +/- 3% (N = 6) of the control c-GMP production. Inhibition of protein kinase C by 50 microM H-7 does not significantly attenuate the H1-receptor response, whereas omittance of extracellular Ca2+ results in almost complete inhibition of the c-GMP production. The histamine-induced c-GMP response is inhibited by hemoglobin, methylene blue and the antioxidants butylated hydroxytoluene and nordihydroguaretic acid, indicating the involvement of a nitric oxide-dependent activation of soluble guanylate cyclase. This suggestion is supported by the concentration-dependent inhibition of the c-GMP production by NG-monomethyl-L-arginine (NMA). At a concentration of 20 microM NMA the histamine (100 microM) response is inhibited to 34 +/- 8% (N = 6) of the control response. This inhibition is reversed to 127 +/- 20% (N = 6) by the exogenous addition of 1 mM L-arginine. These findings show that after an initial H1-receptor-mediated, phospholipase C-dependent, Ca(2+)-mobilization the enzymatic conversion of L-arginine to nitric oxide is stimulated. This nitric oxide production is finally responsible for the activation of soluble guanylate cyclase, leading to the production of c-GMP.

Animals↗