Biomedical subjects

A Lückhoff

Publications and source records attributed to A Lückhoff.

44 records · Page 3Linked to original sources

PubMed1987

Endothelium-derived relaxant factor inhibits platelet activation.

Experiments were designed to investigate whether platelet activation is modulated by endothelium-derived relaxant factor (EDRF) which has been shown to induce vascular smooth muscle relaxation by direct stimulation of soluble guanylate cyclase. EDRF was released from cultured bovine endothelial cells, grown on microcarrier beads, by stimulation with thimerosal in the presence of indomethacin. EDRF had no effect on the intracellular free calcium concentration (Cai2+, measured with the fluorescent indicator indo-1) of resting washed human platelets but significantly attenuated the thrombin-induced rise of Cai2+ from 896 +/- 99 (SEM) to 509 +/- 48 nmol/l. EDRF significantly increased platelet cyclic GMP levels from 0.25 +/- 0.04 to 2.5 +/- 0.4 pmol/10(8) platelets and reduced the thrombin-induced aggregation to 23 +/- 3% of control. EDRF had no effect on Cai2+, cyclic GMP or aggregation after a 3 min storage interval, but superoxide dismutase (shown to increase stability of the labile factor) significantly augmented the EDRF effects on Cai2+. The antiaggregatory potency of EDRF was completely abolished in the presence of hemoglobin. The results characterize EDRF as a potent cyclic GMP-dependent antiaggregatory factor which may act synergistically in vivo with the cyclic AMP-dependent inhibitory effect of prostacyclin.

Animals↗

PubMed1987

Characterization of vascular relaxant factor released from cultured endothelial cells.

Cultured bovine endothelial cells were grown on microcarrier beads. Columns (0.2 ml) packed with microcarriers were perfused with oxygenated (20% O2) Tyrode's solution containing indomethacin (10 microM), and the effluent was passed through precontracted, endothelium-denuded detector arteries. When the endothelial cells were stimulated with bradykinin (3-100 nM), adenosine 5'-triphosphate (0.3-30 microM), or calcium ionophore A23187 (10-300 nM), they released dose-dependently a nonprostanoid compound that dilated the detector vessel. The factor, probably identical to the endothelium-derived relaxing factor of native endothelium, evoked dilations of the same magnitude in different types of detector vessels (rabbit thoracic aorta, rabbit femoral artery, canine coronary artery). However, this relaxant factor was significantly more effective in arteries precontracted by norepinephrine or serotonin than in arteries precontracted by potassium depolarization. Thus, its dilator action resembles that of the nitrovasodilators. The factor is labile, with an apparent half-life in the range of 20 to 30 seconds. Its dilator potency was inhibited by dithiothreitol (0.2 mM), metyrapone (0.2 mM), nordihydroguaiaretic acid (20 microM), and hemoglobin (1 microM), all of which apparently inactivated the factor. Synthesis or release (or both) of the relaxant factor was abolished by methylene blue (1 microM). High PO2 levels (greater than 400 mm Hg) in the perfusate markedly reduced the release of the relaxant factor from the cultured cells. This study demonstrates that a vascular relaxant factor is released from endothelial cells in monoculture by adenosine 5'-triphosphate, bradykinin, and A23187 and establishes such a culture as a useful tool for analyzing the mechanisms of endothelium-dependent vasomotion.

Adenosine Triphosphate↗

PubMed1986

Increased free calcium in endothelial cells under stimulation with adenine nucleotides.

The release of vasodilating substances from the vascular endothelium has been postulated to depend on a rise in the level of intracellular free calcium (Cai++). We measured Cai++ in intact monolayers of calf endothelial cells, grown in culture, that were loaded with the fluorescent calcium indicator quin 2. Fluorescence (excitation wavelength 340 nm, emission wavelength 492 nm) was calibrated by raising Cai++ to a maximum with the calcium ionophore ionomycin (0.1 microM) and by lowering it to a minimum with ionomycin plus manganese (0.4 mM), which quenches quin 2 fluorescence completely. Loss of fluorescent dye from the cells was calculated from fluorescence at the isosbestic excitation wavelength (365 nm). Resting Cai++ was 71 +/- 3 (SEM) nM. ATP (adenosine-5'-triphosphate) raised Cai++ dose-dependently and reversibly to 458 +/- 60 nM at a concentration of 10 microM, and at 0.1 mM to values close to those that occurred under ionomycin. ADP (A-5'-PP) and AMP (A-5'-P) had smaller effects with a maximal Cai++ of 287 +/- 72 nM at 30 microM ADP and 176 +/- 17 nM at 0.1 mM AMP. At these concentrations, ADP and AMP attenuated significantly the increase of Cai++ under ATP (10 microM). Adenosine (0.1 or 0.3 mM) and acetylcholine (0.1 to 30 microM) enhanced Cai++ inconsistently, by a maximum of 50 nM. These effects were abolished by theophylline and atropine, respectively. In the absence of extracellular calcium, ATP still raised Cai++, although endothelial responsiveness declined after repetitive stimulations. We conclude that activation of purinergic receptors increases intracellular free calcium in endothelial cells, and that this increase is probably an essential trigger for synthesis of prostacyclin and the labile endothelium-derived relaxant factor.

Acetylcholine↗

PubMed1986

Measuring cytosolic free calcium concentration in endothelial cells with indo-1: the pitfall of using the ratio of two fluorescence intensities recorded at different wavelengths.

Indo-1 is a new fluorescent indicator of the intracellular free calcium concentration Cai++. Indo-1 may be used in a similar manner as its predecessor quin2 but offers the principal advantage that the Ca++ saturated form of the Ca++ chelator has a emission maximum different in wavelength from that of free indo-1 (400 nm versus 483 nm). Therefore, the ratio of the fluorescence intensity F emitted at 400 nm to that of the fluorescence intensity G emitted at 483 nm (or 500 nm) should be a measure of Cai++ independent of the total amount of intracellular dye. However, when indo-1 is loaded into endothelial cells (grown in culture on quartz coverslips) by incubation with the acetoxymethylester of indo-1 (indo-1/AM), the ester in not completely hydrolysed to indo-1 intracellularly. Fluorescence emitted by uncleaved indo-1/AM at wavelengths 483-500 nm interferes with the fluorescence of indo-1. Ester fluorescence is influenced not only by ester concentration but by the fluorescence emitted at 400 nm by Ca++ bound indo-1 as well. Therefore, the ratio F/G cannot reliably evaluate increases in Cai++ in endothelial cells although F/G would indicate a basal Cai++ constant with time. By contrast, the fluorescence F is a sensitive parameter of the intracellular concentration of Ca++ bound indo-1, in particular when the excitation wavelength is set to 332 nm. F was used to measure resting Cai++ in endothelial cells (132 +/- 22 nM; n = 22) and to demonstrate dose-dependent and reversible increases in Cai++ in response to stimulation with bradykinin.

Animals↗

PubMed1985

Role of the endothelium in the vasomotor effects of angiotensin I in isolated arteries with or without inhibition by MK 422.

We investigated in the isolated rabbit aorta the ability of the endothelium to attenuate the vasoconstrictor effects of angiotensin (ANG) I and II. After preincubation with enalaprilat, which inhibited the conversion of ANG I to ANG II by 80%, the contractile response to ANG I (10(-8) to 10(-6) mol/l) was significantly greater in aortae which were endothelium-denuded compared with endothelium-intact segments. There was no such difference for ANG II (10(-10) to 3 x 10(-8) mol/l). We conclude that an endothelium-mediated dilatation is part of the net vasomotor action of ANG I.

Angiotensin I↗

PubMed1984

Hormonal regulation of electrolyte and water transport in the colon.

The colon participates in water and electrolyte homeostasis by the absorption of sodium (Na) and water as well as by potassium (K) secretion. The primary step of colonic transport is the active Na transport via a transcellular route. Steroidal hormones considerably increase Na absorption by utilizing two mechanisms: (1) passive Na entry into the cells in enhanced by an increased membrane permeability; (2) active transport capacity is increased by a stimulation of ATPase synthesis. Mineralocorticoid versus glucocorticoid actions of steroids have not yet been clearly differentiated; parallel influences are possible. Active chloride (Cl) secretion is found in the colon under certain pathological conditions and is induced by a number of factors, e.g., hormones produced by pancreas tumors. Cellular events involve a rise of intracellular cAMP and calcium (Ca) concentrations, and altered Cl permeabilities. Functional changes of colonic epithelial cells caused by hormones assume a significant role in the etiology of diarrhea, as well as in compensatory processes by which an intestinal loss of electrolytes and water is prevented.

Adrenal Cortex Hormones↗

PubMed1983

Aldosterone on sodium transport of rat distal colon in long-term adrenalectomy during acute and chronic substitution.

1. The influence of aldosterone upon water and sodium transport properties of the distal colon was studied in long-term adrenalectomy (11-29 days).2. Six groups of rats were used: I, normal (control); II, adrenalectomized; III, adrenalectomized, acutely substituted with aldosterone (200 mug/kg 4 h); IV, adrenalectomized rats receiving aldosterone simultaneously with the specific inhibitor spironolactone (40 mg/kg within 4 h); V, adrenalectomized, substituted chronically with aldosterone (2 x 75 mug/kg day); VI, adrenalectomized, substituted chronically with dexamethasone (120 mug/kg day).3. Distal colon segments were perfused in vivo with isotonic Ringer solution. In addition, a hypotonic electrolyte solution (Na(+) 111 mM) was used in groups I and II.4. In adrenalectomy (group II), net water absorption (J(v)) was significantly decreased from (normal) 54.4 mul/h cm(2)+/-10.5 (n = 9) to 41.2 mul/h cm(2)+/-7.3 (n = 4), and net Na(+) absorption (J(Na)) was decreased from 13.6 mumol/h cm(2)+3.5 to 8.5 mumol/h cm(2)+/-0.9 (isotonic perfusate). Similarly, J(v) was decreased from 54.0 mul/h cm(2)+/-8.3 (n = 4) to 37.3 mul/h cm(2)+/-4.2 (n = 7), and J(Na) from 8.6 mumol/h cm(2)+/-2.1 to 4.2 mumol/h cm(2)+/-2.1 (hypotonic perfusate).5. Acute aldosterone substitution in adrenalectomy (III) had no effect upon J(v) (37.1 mul/h cm(2)+/-10.3; n = 5) but increased J(Na) to 10.3 mumol/h cm(2)+/-0.3.6. The luminal Na(+) steady-state concentration was higher in group II (11.2 mmol l(-1)+/-3.6; n = 6) than in group I (3.3 mmol l(-1)+/-1.4; n = 29). Acute aldosterone substitution restored this value to normal (3.0 mmol l(-1)+/-1.2; n = 4). The aldosterone effect was partly blocked by spironolactone: the Na(+) steady-state concentration was 6.4 mmol/l+/-0.6 (n = 3) in group IV.7. At the steady-state luminal Na(+) concentration, the osmotically driven net water fluxes were not different in groups I and II, indicating that the hydraulic permeability coefficient is not altered in adrenalectomy.8. In group V, J(v) (54.9 mul/h cm(2)+/-10.9; n = 7) and J(Na) (11.9 mumol/h cm(2)+/-1.7; n = 6) were not significantly different from normal.9. In group VI, J(v) (37.3 mul/h cm(2)+/-6.0; n = 5) and J(Na) (8.0 mumol/h cm(2)+/-1.4) were not significantly different from group II.10. The mineralocorticoid effects of aldosterone in long-term adrenalectomy appear to represent the principal determining factors of colonic J(v) and J(Na).

Adrenalectomy↗

PubMed1981

Hydraulic permeability coefficient and sodium steady-state luminal concentration of the in vivo perfused rat distal colon.

The distal colon (rat) was perfused in vivo at low rates (1-2 ml/h). Dialyzed polyethyleneglycol 4,000 (PEG) was used to vary the luminal osmotic activity (pi eff). Perfusate sodium concentration of 3.7 mmol 1-1 +/- 0.9 (SD) resulted in an effluent Na+ of 3.3 mmol 1-1 +/- 1.4. Potassium concentrations of 15 or 25 mmol 1-1 remained unaltered. At this cationic steady-state, transmural net water flux (Jv) was linearly correlated (r = -0.96, n = 24) to the logarithm of pi eff. Jv was zero at 634 mosm 1-1. Lp (microliters h-1 cm-2 mosm-1 l) was 0.195 at pi eff 85 mosm 1-1 and 0.046 at 1,050 mosm 1-1. These data characterize the distal colon as an epithelium with high capacity for salt conservation.

Animals↗