1-O-alkyl-2-O-acetyl-sn-glycero-3-phosphocholines: influence on aggregation and [3H]serotonin release of human thrombocytes.

1-O-Alkyl-2-O-acetyl-sn-glycero-3-phosphocholines (platelet activating factor, PAF) aggregate human thrombocytes in a concentration dependent fashion. After a short lag-phase, maximum aggregation is reached within 2 min. PAF releases serotonin from human thrombocytes within 1 min. Indomethacin and creatine phosphate (CP)/creatine phosphokinase (CPK) are able to inhibit the second phase of the aggregation by PAF, while xylocain reduces both the first and second phase of aggregation of human thrombocytes. Hirudine neither influences the first nor the second phase of aggregation by PAF.

Saturated and unsaturated 1-0-alkyl-2-0-acetoyl-sn-glycero-3-phosphocholines and 2-lyso derivatives from ratfish liver oil. Effect on adenosine 3':5'-cyclic monophosphate concentration in hepatocyte suspensions.

1-0-Alkyl-2-0-acetoyl-sn-glycero-3-phosphocholines, containing saturated or unsaturated alkyl moieties and their 2-lyso derivatives, prepared from ratfish liver oil (Hydrolagus colliei), were studied for their in vitro effect on cellular adenosine 3':5'-cyclic monophosphate (cAMP) concentration in freshly isolated hepatocytes from adult rats. We report here for the first time that semisynthetic platelet-activating factor (PAF), the saturated and unsaturated compound, from 10(-6) M to 10(-12) M, is effective in lowering cAMP concentrations by 50% after 4 h of incubation. This effect becomes apparent after 30 min. Concentrations from 10(-12) M up to 10(-20) M did not influence cAMP levels. The 2-lyso derivatives (10(-8) M) of both compounds are equally active as compared to their 2-0-acetoyl derivatives, in lowering cAMP levels after 4 h of incubation. We discuss the existence of a transacetylating enzyme, generating the active form in rat liver cells.

Taurocholate inhibits the glucocorticoid-induced rise of 3-hydroxy-3-methylglutaryl-CoA reductase in primary culture of hepatocytes.

The influence of taurocholate, the major bile acid of the rat, on 3-hydroxy-3-methylglutaryl-CoA reductase [mevalonate: NADP+ oxidoreductase (acylating CoA); EC 1.1.1.34], the regulatory enzyme of cholesterol synthesis, was studied in primary cultures of rat hepatocytes. The basal activity of the enzyme was not altered by adding up to 10 microM taurocholate to the culture medium. On the contrary, 1nM to 10 microM taurocholate caused a dose-dependent inhibition of enzyme activity within 6 h if added simultaneously with 10 microM dexamethasone. Because this glucocorticoid causes a cycloheximide-sensitive rise of 3-hydroxy-3-methylglutaryl-CoA reductase activity in this system the results are taken as evidence that bile salts inhibit the synthesis of the enzyme. The induction of tyrosine transaminase (L-tyrosine:2-oxoglutarate aminotransferase; EC 2.6.1.5) by dexamethasone was not influenced by taurocholate, which demonstrates that the glucocorticoid sensitivity of the cells was not impaired by the bile salt. It is concluded that there is a direct control of hepatic cholesterol synthesis by bile salts.

[Thrombocyte function in essential thrombocythemia and reactive thrombocytosis]. / Untersuchungen zur Thrombozytenfunktion bei essentieller Thrombozythämie und reaktiver Thrombozytose.

Blood of 16 patients with essential thrombocythemia (ET), 9 patients with reactive thrombocytosis (RT) and 13 healthy persons was used for platelet aggregation studies. When the aggregation was induced with adenosine diphosphate (0.01 microM), collagen (0.1 micrograms/ml) or platelet activating factor (PAF 0.5 microM) the plasma of the patients with ET showed significantly decreased aggregation (35%-44% of the value for the control groups). Independent of inhibitors of platelet aggregation, thrombin (0.05 U/ml) caused similar aggregation in healthy controls and patients with ET; patients with RT showed an increase aggregation. Adrenalin-induced aggregation discriminated best between patients with ET and controls. Adrenalin in concentrations ranging from 0.01 micrograms/ml to 100 micrograms/ml caused comparable dose-related amounts of aggregation in healthy controls and patients with RT. Over the whole concentration range, patients with ET showed significantly decreased aggregation (28%-34% of the value for the control groups). This difference proved to be independent of the influence of inhibitors of platelet aggregation. Though concentrations of alpha1-acid glycoprotein never reached inhibitory levels in the plasma of patients with ET (n = 12) they were significantly higher compared with those in normal plasma (n = 12). Fibrinogen concentrations in plasma of ET-patients (n = 12) were in the normal range. Cellular adenosine 3'-5'-cyclic monophosphate concentrations in ET (n = 10) are comparable with normal values (n = 5). The significance of the results for diagnosis and better pathophysiological understanding of ET is discussed.

Effects of a thio analog of platelet-activating factor on platelet aggregation and adenosine 3',5'-monophosphate concentration in hepatocyte suspensions and in platelets. A comparison with the naturally occurring compound.

2-Acetyl-S-octadecyl-rac-1-thioglycero-3-phosphocholine, the thio analog of platelet-activating factor, in concentrations of 10(-6) M to 10(-12) M in the medium, lowers cAMP-concentrations in rat hepatocytes. In a concentration of 10(-4) M, the thio analog aggregates human platelets irreversibly, above 10(-5) M aggregation shows a reversible course. Compared with the saturated or unsaturated ether analogs, the thio compound shows less platelet-aggregating potency. We have correlated this difference in platelet aggregation with reduced cAMP-depressing activity of the thio analog (compared with saturated and unsaturated 2-acetyl-1-O-alkyl-sn-glycero-3-phosphocholines).

Influence of fatty acids on microsomal diacylglycerol acyltransferase activity in primary cultures of hepatocytes.

The regulation of triacylglycerol synthesis by various long-chain fatty acids was studied using primary cultures of hepatocytes. The activity of diacylglycerol acyltransferase was measured after 72-h incubation with various fatty acids in equimolar concentrations (0.5 mM): oleate, linoleate, linolenate, palmitate, stearate and arachidonate increased enzyme activity to 191%, 161%, 148%, 124%, 116% and 114%, respectively, compared to controls. Cellular triacylglycerol levels showed the same changes, suggesting a regulatory function of diacylglycerol acyltransferase in triacylglycerol synthesis. Elaidate lowered the enzyme activity by 57%. The addition of different fatty acids to the incubation medium did not influence the distribution of enzyme activity in the 30 000 x g pellet which was routinely discarded and the microsomal fraction (105 000 x g), where the enzyme was predominantly localized (91%). The release of lactate dehydrogenase into the culture medium of hepatocyte monolayers as well as ketone body production was not affected by any of the fatty acids.

Influence of fatty acids on cholesterol synthesis of hepatocytes in monolayer culture.

The short-term (6-hour) and long-term (72-hour) influences of a wide spectrum of fatty acids on cholesterogenesis in monolayer cultures of rat hepatocytes were studied. A 6-hour addition of 0.5 mmol/liter of oleate to the culture medium raised 3-hydroxy-3-methylglutaryl-CoA reductase [mevalonate:NADP+ oxidoreductase (CoA-acylating), EC 1.1.1.34] activity by 62%. Octanoate, palmitate, stearate, linoleate, arachidonate and linolenate did not change enzyme activity significantly under these circumstances. A 72-hour incubation led to a 162% rise of enzyme activity by oleate and a 33% lowering by arachidonate, the other long-chain fatty acids having no significant effect (linoleate, linolenate, palmitate and stearate). These modulations of enzyme activity were paralleled by comparable changes of cholesterogenesis as measured by incorporation of [1-14C]acetate into cholesterol. The results are compatible with the concept that the response of hepatic cholesterogenesis to dietary triglycerides in vivo (observed by earlier investigators) is due to influences of the triglyceride fatty acids on hepatocyte 3-hydroxy-3-methylglutaryl-CoA reductase activity.