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Biomedical subjects

D Blache

Publications and source records attributed to D Blache.

At least 91 records · Page 5Linked to original sources
PubMed1991

Hepatic lipase activity during oral and parenteral 17 beta-estradiol replacement therapy: high-density lipoprotein increase may not be antiatherogenic.

OBJECTIVE: Hepatic lipase activity is involved in the removal of cholesterol and phospholipid from plasma high-density lipoprotein (HDL) to the liver. Changes in hepatic lipase are responsible for some lipoprotein modifications observed during estrogen administration (i.e., increased HDL levels). The aim of this study was to compare the effects of alternative routes of administration of estrogen on hepatic lipase activity and lipoprotein metabolism. DESIGN, PATIENTS: The effects of oral and parenteral estradiol (E2) replacement therapy on post-heparin hepatic lipase were studied in the short-term (6 weeks) in postmenopausal women. INTERVENTIONS: Ten patients received 2 mg/d or oral micronized E2 and 10 patients 3 mg/d of percutaneous E2. RESULTS: Both treatments significantly increased plasma E2 levels. Hepatic lipase decreased (-33%) significantly (P less than 0.05), and the phospholipids and free cholesterol content of HDL and HDL3 increased significantly (P less than 0.05) during oral but not during parenteral treatment. CONCLUSIONS: The short-term pharmacological effect of E2 on hepatic lipase depends on the route of administration. The inhibition of this enzyme may reduce the removal of HDL-cholesterol by the liver. The expected vascular benefits of such a pharmacological increase in HDL are questionable.

Administration, Cutaneous↗
PubMed1989

Effects of organic and inorganic Ca antagonists on rat platelet arachidonic acid metabolism in the presence of Ca2+, Sr2+ and Ba2+.

The effects of Ca-antagonists on the thrombin-induced mobilization of radiolabeled arachidonate preincorporated into rat platelets as well as the subsequent formation of labeled cyclooxygenase and lipoxygenase products were analyzed in the presence of either Ca2+ or Ca2+-substitutes, Sr2+ and Ba2+. Results indicate that following thrombin stimulation (0.2 U/ml) in the presence of Ca2+, nitrendipine (Nit), Cd2+ or Mn2+ reduced the release of arachidonate and the biosynthesis of thromboxane B2. Inhibition of arachidonic acid release and metabolism were also obtained by both Nit and Cd2+ in the presence of Sr2+ and Ba2+. Results from studies with a semi-purified phospholipase A2 fraction prepared from rat platelets indicated that the activity was almost unaffected by Nit or Cd2+. From these findings, we concluded that inhibition of platelet-induced release and metabolism of arachidonic acid by the Ca-antagonists tested require intact platelets. These data support the hypothesis of an interaction of these agents at an unknown surface membrane level.

Animals↗
PubMed1988

Biological effects of oxysterols on platelet function.

We have studied the in vitro exposure to various oxysterols on isolated rat platelets. The oxysterols (1-20 uM) were incubated either dissolved in methanol or as albumin-bound complexes. Aggregation (analyzed by turbidimetry) was measured after stimulation by thrombin or ADP. Serotonin secretion (analyzed by voltammetry) was measured after thombin stimulation. We found that platelet aggregation and serotonin secretion could be either not significantly affected (7B-hydroxy cholesterol) or potentiated (22S-hydroxy cholesterol, 3,5,6-hydroxy cholestan triol) or inhibited (25-hydroxy- and 7-oxo cholesterol), after in vitro incubation with different oxysterols. Our data indicate that a modulation of the platelet behavior occurs after in vitro incubation with different oxysterols, some derivatives acting as inhibitors and others as potentiators. These results provide new interesting information regarding the role of these sterols in cell membrane structure and function in relation to pathology.

Animals↗
PubMed1987

Rat platelet arachidonate metabolism in the presence of Ca2+, Sr2+ and Ba2+: studies using intact platelets and semi-purified phospholipase A2.

To document further the involvement of external Ca2+ in the platelet-induced activation process, we have studied the arachidonate metabolism of intact washed rat platelets in the presence of different concentrations of Ca2+, Sr2+ or Ba2+. The thrombin-induced mobilization of radiolabeled arachidonate preincorporated into platelet phospholipids was followed as well as the subsequent formation of labeled cyclooxygenase and lipoxygenase products. Results indicate that upon thrombin stimulation (0.2 U/ml), the release of endogenous arachidonate and the formation of its metabolites are reduced by 50-90% only by omission of Ca2+ as compared to 1 mM Ca2+ in the suspending medium. At higher Ca2+ concentrations (5 mM), the arachidonate mobilization and metabolite formation are inhibited and the data are thus close to those obtained in the absence of Ca2+. In the presence of Sr2+ or Ba2+, the results indicate that these cations can substitute for Ca2+. As for Ca2+, an optimum concentration is found for Sr2+ and Ba2+ (3-5 mM), and higher concentrations inhibit the metabolism of arachidonic acid. As the above data might be compatible with the possible entry of Sr2+ and Ba2+ into platelets upon stimulation, we also studied the activity of a semi-purified preparation of phospholipase A2 from rat platelets. This activity was assayed (pH 9.2) using heat-denatured [3H]arachidonate-prelabeled phospholipids as substrate. The results show that this phospholipase A2 activity was strongly Ca2+-dependent. In addition, we found that unlike Mg2+, Sr2+ and Ba2+ are able to greatly enhance this activity. Relative efficiency (Vmax) was in the order Ca2+ greater than Sr2+ greater than Ba2+. Taken together, these findings suggest that external Ca2+ may play a major role in the regulation of rat platelet activity. Our interpretation is in line with the view that Sr2+ or Ba2+ could enter the platelet through a mechanism common to Ca2+ (a Ca2+ channel). Although direct evidence is awaited from the results of further studies which are in progress, it can reasonably be considered that Sr2+ or Ba2+ might cause platelet-induced activation mimicking a rise in the cytosolic Ca2+ and subsequent activation of Ca2+-dependent enzymes.

Animals↗
PubMed1987

Direct evidence for the modulation of human platelet cytosolic free Ca2+ by intracellular cyclic AMP produced with a photoactivatable derivative.

We have previously reported that intraplatelet "cyclic AMP jumps" produced with newly synthesized photoactivatable cyclic AMP analogue, inhibited washed rat platelet aggregation and serotonin release as induced by thrombin. Using the same approach on human platelets, thrombin-induced platelet aggregation was dose-dependently inhibited only when a flash was delivered. The mechanism of action of intraplatelet cyclic AMP as resulting from photolysis could be by controlling the level of cytosolic Ca2+. In order to test this hypothesis, the same protocol was used on human platelets preloaded with the internal Ca2+ fluorescent indicator, Quin 2, we found that the extent and the rate of the rise of the cytosolic Ca2+ induced by thrombin were dramatically decreased, in the presence of the photoactivatable cyclic AMP, only following photoirradiation. In addition, the flashes were produced, in the presence of photoactivatable cyclic AMP, after the thrombin-induced rise of internal Ca2+ had reached its peak. In these conditions, photoirradiation caused a rapid fall in fluorescence. These experiments provide the first direct evidence that intracellular cyclic AMP is involved in the control of platelet cytosolic Ca2+ by inhibition of its mobilization and by stimulation of its sequestration.

Aminoquinolines↗
PubMed1987

Platelet aggregation and endogenous 5-HT secretion in presence of Ca2+, Sr2+ and Ba2+. Effects of calcium antagonists.

Washed rat platelets aggregation and endogenous serotonin release were studied after thrombin stimulation in the presence of different concentrations of Ca2+, Sr2+ or Ba2+. The extent of platelet aggregation and release was found to depend upon the external concentration of these cations. For all of them, an optimum concentration could be defined. Higher concentrations were shown to inhibit both aggregation and release. Efficiency to support thrombin-induced aggregation was in the order Ca2+ greater than Sr2+ greater than Ba2+. Complete inhibition of aggregation and release induced by thrombin was obtained after a 30 second preincubation with 38 uM nitrendipine, 1 mM Cd2+ or 1 mM Mn2+. Inhibition was obtained in the presence of Ca2+, Sr2+ or Ba2+. These results are consistent with the hypothesis that Sr2+ and Ba2+ are able to support platelet activation acting as Ca2+ substitutes. Following thrombin stimulation, they could penetrate the platelets and mimick a rise in cytoplasmic Ca2+.

Animals↗
PubMed1987

The effect of calcium channel blockers on blood platelet function, especially calcium uptake.

The effects of organic and inorganic calcium antagonists on washed platelets from rat and human have been studied. Platelet aggregation was assessed by turbidimetry. Endogenous serotonin release was measured on the same sample by means of electrochemically treated carbon fiber electrodes. The organic calcium antagonist, nitrendipine, and the inorganic calcium channel blockers (Co2+, Mn2+, Cd2+, La3+) drastically inhibited rat and human platelet aggregation induced by thrombin, ADP or adrenaline in the presence of 0.32 mM Ca2+. In our conditions, the thrombin-induced release of endogenous serotonin was found to be external Ca2+-dependent and completely inhibited by 20 microM nitrendipine or 1 mM Cd2+. In addition, Ba2+ or Sr2+ ions can be substituted for Ca2+ to bring about platelet aggregation as well as endogenous serotonin secretion. In Ba2+ or Sr2+-containing media, rat platelet aggregation and/or serotonin secretion can be inhibited by either nitrendipine or Cd2+. Finally, we have also studied the thrombin- and external Ca2+-dependence of radiolabeled calcium uptake by rat platelets. We found that the thrombin-induced 45Ca uptake was inhibited by either 18 microM nitrendipine or 1 mM Cd2+. These results provide strong evidence for the existence of an influx of divalent cations (Ca2+, Sr2+, Ba2+) triggering platelet function. They also suggest, although they do not prove, that the translocation of these cations occurs through an agonist-operated channel as proposed by Hallam and Rink (FEBS Lett. 186 (1986) 175-179).

Animals↗
PubMed1986

Direct evidence for the inhibition of platelet aggregation and release by intracellular cyclic AMP produced with a new photoactivatable derivative.

An increase in platelet cyclic AMP (cAMP) via stimulation of adenylate cyclase is thought to be the underlying mechanism by which potent prostaglandins i.e. PGD2, PGI2, inhibit platelet functions. We report here new and direct evidence for the inhibitory effects of cAMP on platelet aggregation and serotonin release. Washed platelets from rat were incubated with a new photoactivatable cAMP analogue (4,5-dimethoxy-2-nitrobenzyl ester); this compound is almost physiologically inert before irradiation and liberates free cAMP ("cAMP jumps") following light flashes. A single flash, delivered after 2 min incubation in 100-200 microM of the analogue, dramatically inhibited thrombin-induced aggregation, as compared with controls. Endogenous serotonin release, measured in the same samples by means of an electrochemically treated carbon electrode was undetectable after the cAMP jump. Pre-irradiated solutions added to platelets had no effect. The kinetics of the flash-induced effects were also studied. From these results we can conclude that: i) the photoactivatable cAMP derivative has to permeate through the platelet membrane; ii) the analogue remains photolabile; and, iii) intracellular cAMP, resulting from photolysis dramatically inhibits platelet aggregation and serotonin release. It is possible that cAMP exerts its effects by regulating cytoplasmic free calcium concentration and/or other actions affecting platelet activation.

Animals↗
PubMed1985

Endogenous platelet serotonin release monitored during aggregation by means of a new electrochemical technique.

Differential pulse voltammetry combined with electrochemically treated carbon fibre microelectrodes was used to monitor endogenous serotonin release occurring during platelet aggregation. After platelet stimulation by thrombin, an oxidation peak was recorded at +280 mV. HPLC analyses performed with fluorimetric detection have shown that this released electroactive compound was essentially serotonin. Moreover, serotonin measurements in the same samples by the technique reported here and by fluorimetry were found to be very similar (1.15 +/- 0.30 microM and 1.17 +/- 0.15 (mean +/- S.D., n = 6), respectively). Extracellular serotonin concentrations could be estimated either directly during aggregation or in supernatants obtained from stimulated or lysed platelets. Maximal serotonin concentrations have been found to be 6.93 +/- 0.37 and 3.28 +/- 0.39 nmol/10(9) platelets from rat and human, respectively. Using the reported procedure, we have observed that no serotonin was released from thrombin-stimulated platelets prepared from rats treated with reserpine. Our new technique represents a selective and performant tool for rapid determination of endogenous serotonin platelet secretion.

Animals↗
PubMed1984

A sensitive method for the routine determination of plasma nicotine by flame ionization gas-liquid chromatography.

A gas chromatographic procedure for the analysis of nicotine in plasma, which uses quinoline as an internal standard, is reported. The nicotine is extracted with diethyl ether, concentrated without any evaporation, thus avoiding losses, and analyzed without derivatization. The recovery is 83.2 +/- 6.1% (n = 6). Although the analysis is carried out with a classical flame ionization detector, the detection limit is 0.1 ng/ml. Linearity is observed up to 100 ng/ml. The results of the precision analysis performed in the working range indicate a good reproducibility: a coefficient of variation of 5.2% is obtained for within-run analysis and 10.5 to 4.5% for nicotine values from 2.9 to 19.1 ng/ml for day analysis. Since a single run (the limitative step) lasts less than 15 min this improved procedure allows a great number of samples to be processed per day.

Animals↗
PubMed1984

Vitamin E prevents the platelet abnormalities induced by estrogen in rat.

Platelet lipid biosynthesis in relation to aggregation has been studied in female rats treated with ethynylestradiol and fed laboratory chow or a vitamin E-deficient diet. In both normal and vitamin E-deficient rats, administration of ethynylestradiol highly significantly (p less than .001) increased the biosynthesis of total lipids but mostly of lanosterol (+ dihydrolanosterol) by thirteen-fold in normal rats and by nine-fold in vitamin E-deficient rats. The increased lipid synthesis was associated with a higher response of platelets to thrombin-induced aggregation. Concomitant administration of alpha-tocopherol acetate in both normal and vitamin E-deficient rats depressed markedly the enhanced lipid synthesis and aggregation induced by estrogen. Administration of ethynylestradiol lowered considerably the level of vitamin E in plasma but not in platelets. Treatment by tocopherol partly corrected the low plasma level of vitamin E resulting from estrogen administration. In vitro addition of lanosterol to platelets highly significantly increased the response of platelets to thrombin- and ADP-induced aggregation. This hyperaggregability was almost entirely inhibited by preincubation of platelets with tocopherol acetate. In the present in vivo and in vitro studies, alpha-tocopherol was able to neutralize most of the adverse effects of estrogen on blood platelets.

Animals↗
PubMed1984

Platelet function after cigarette smoking in relation to nicotine and carbon monoxide.

In two sets of experiments involving 10 smokers, we followed the acute effect of inhaling smoke from cigarettes with five different nicotine yields (0.07 to 1.44 mg) on platelet function in relation to blood levels of carboxyhemoglobin and nicotine. Blood was drawn from fasted subjects who had not smoked for 10 hr before and after smoking one cigarette. Depending on the cigarette, the increase in platelet aggregation to thrombin, adenosine diphosphate, collagen, and epinephrine 10 min after smoking ranged from 0% to approximately 80% for the cigarettes with the higher nicotine yields. Blood nicotine levels increased from 112% to 644%. Clotting activity of platelet-rich plasma (PRP) and platelets rose by 16% with the cigarettes with the highest nicotine contents. Platelet activity correlated with blood nicotine levels but not with carboxyhemoglobin levels. Nicotine diluted in saline solution and added in vitro to PRP from six other subjects 2 min before the aggregation or clotting test at levels after smoking (10 and 20 ng/ml) induced a rise in platelet reactivity of the same order as that after smoking cigarettes. Data suggest that in some cigarettes, nornicotine and substances contained in tar may contribute to the effect of cigarette smoking on platelets.

Adenosine Diphosphate↗
PubMed1983

Simple, reproducible procedure for selective measurement of lipoprotein lipase and hepatic lipase.

Existing methods for determining the release of lipoprotein lipase (EC 3.1.1.34) and hepatic lipase (EC 3.1.1.3) into plasma after heparin injection give highly variable results, primarily traceable to errors in the isolation of labeled oleate from the substrate, triolein. Methods involving anion-exchange resin to bind oleate show high variability and have a low yield. Introducing a strong base in the last step of the assay may spuriously increase the counts from oleate, whereas a detergent such as Triton X-100, used to minimize this problem, has a strong quenching effect. We report a simple and rapid method in which we eliminate rather than correct for the sources of variation. The substrate, tri[1-14C]oleoyl-glycerol, is sonicated under strictly standardized conditions with gum arabic, 50 g/L. Incubation is stopped by addition of a benzene/chloroform/methanol mixture and NaOH, 0.2 mol/L. Labeled oleic acid is extracted with hexane after acidification of the alkaline aqueous (upper) phase, so that no alkali is introduced into the scintillation liquid. For lipoprotein lipase measurement, hepatic lipase is inactivated by a specific antiserum, whereas hepatic lipase is measured after lipoprotein lipase is inactivated by NaCl, 1.0 mol/L. The method is efficient and specific, and quenching and chemiluminescence artifacts are avoided.

Animals↗
PubMed1982

Biosynthesis of platelet lipids in relation to aggregation in women using oral contraceptives.

The platelet lipid biosynthesis in relation to platelet aggregation and lipemia was studied by 14C-acetate and mevalonate incorporation into platelets of seventeen women without medication and of eighteen women using a low estrogen oral contraceptive. The lipid biosynthesis was significantly increased by 59% (mevalonate) and 38% (acetate) in women on oral contraceptives. From mevalonate, lipid synthesis was increased mostly in the lanosterol-dihydrolanosterol fraction (p less than .01). From acetate, lipid synthesis was significantly enhanced in all the lipid classes. In the oral contraceptive group, the response of platelets to thrombin aggregation was only slightly higher, but HDL-cholesterol was significantly lower. However, in the women using oral contraceptives, the percentage of abnormal values in HDL-cholesterol, thrombin-aggregation and acetate incorporation into lanosterol was similar. Thus, more than 40% of the women studied here, using low estrogen oral contraceptives, presented an increase in platelet lipid biosynthesis, especially in the lanosterol-dihydrolanosterol fraction, which was significantly correlated (p less than .05) with the response of their platelets to thrombin-induced aggregation.

Adult↗
PubMed1982

Tissue lipids in acute acrylamide intoxicated rats.

A preliminary survey of tissue lipid composition in acrylamide intoxicated rats is reported. The animals were injected intraperitoneally with acrylamide 50 mg/kg body weight per day for 10 days. Liver cholesterol, mainly in the ester fraction, was decreased in treated rats. When fatty acid composition of liver cholesterol esters was examined, the proportions of linoleate and stearate were found to be decreased and were compensated by the increase of palmitate. Atrophy of epididymal fat pad resulted in severe triglyceride depletion and a relative increase in the proportion of phospholipids and cholesterol. There was also a reduction of linoleate, palmitate and palmitoleate in triglycerides and phospholipids of this tissue. There were, however, only minor changes in the fatty acid profile of the sciatic nerve.

Acrylamides↗
PubMed1982

Plasma lipoprotein lipase and hepatic lipase activities in Friedreich's ataxia.

Plasma triglycerides although within the normal range have been shown to be higher in Friedreich's ataxia than in control subjects. To determine whether this difference could be ascribed to a reduced catabolism of triglyceride-rich lipoproteins, the activities of lipoprotein lipase (LPL) and hepatic triglyceride lipase (HL), released into plasma after an heparin injection, were measured in 13 cases of Friedreich's ataxia and 14 control subjects of comparable signs. LPL was found to be significantly lower in the ataxic patients. Moreover about half of the cases clustered below the normal range for both lipase activities. This subgroup of Friedreich's patients had significantly higher plasma triglycerides than those with normal lipase activities. Further studies are needed to relate these findings to other characteristics of the disease.

Adolescent↗
PubMed1981

[Structure of the heptose region of the lipopolysaccharide from Escherichia coli K12 CR34 (author's transl)].

The heptose region of the lipopolysaccharide of Escherichia coli K12 CR34 was studied. The glucose linked to the heptose II was found to be substituted by a D-galactose and the linear chain of the core polysaccharide has two (1 lead to 3) linked heptoses. The heptose II is substituted by a lateral (1 leads to 7) linked heptose III and heptose I is linked in (1 leads to 5) to 2-deoxy-D-manno-octulosonic acid. The three sugars of the linear chain, heptose I, heptose II and glucose are substituted by phosphate, pyrophosphate or pyrophosphorylethanolamine group linked to C-4 hydroxyl groups. However, in some polysaccharidic chains one or two substituting groups may be absent. This result may explain the heterogeneity in the length of the core polysaccharidic chains.

Carbohydrate Conformation↗