The bioavailability and pharmacodynamics of different concentrations of omega-3 acid ethyl esters.

Omega-3 fatty acids have a long history of use as dietary supplements and more recently for therapeutic applications as prescription pharmaceuticals. Achieving a high concentration is critical for developing convenient, practical therapeutic formulations. The objective of the study was to explore the uptake and effects of different concentrations of omega-3 acid ethyl esters. Three different omega-3 concentrations were investigated in a clinical study with 101 subjects. All participants were dosed for 14 days with 5.1g per day of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) ethyl esters provided in three concentrations: 62.5%, 80% and 85% of total fatty acids. Key endpoints of the study were serum phospholipids and standard fasting lipid panels at day 14. Although administered the same quantity of omega-3 fatty acids, the patients taking the more concentrated formulations had higher levels of EPA/DHA in serum phospholipids and greater reductions in serum triglyceride and VLDL cholesterol levels. Total and non-HDL cholesterol were significantly reduced from baseline with all three formulations. In conclusion the concentration of omega-3 fatty acids of the formulations studied had independent effects on the uptake and effect outcomes during short-term administration. Very high concentrations of omega-3 acid ethyl esters (80%) appear to have higher uptake and are more potent for reducing triglycerides (TGs) and VLDL-cholesterol than formulations with lower concentrations.

The effect of aliphatic adenine analogues on S-adenosylhomocysteine and S-adenosylhomocysteine hydrolase in intact rat hepatocytes.

The aliphatic adenine analogues, D-eritadenine, L-eritadenine, L-threoeritadenine, and 9-(S)-(2,3-dihydroxypropyl)adenine [(S)DHPA] function as inhibitors/inactivators of purified S-adenosylhomocysteine (AdoHcy) hydrolase, but these compounds did not induce reduction of enzyme-bound NAD+. D-Eritadenine, L-eritadenine, (S)DHPA, and L-threo-eritadenine inactivated AdoHcy hydrolase in hepatocytes, and the efficiency decreased in the order mentioned. Concurrently, there was an increase in the AdoHcy content. The accumulation of AdoHcy in the presence of (S)DHPA was more pronounced than would be expected from the inactivation of enzyme activity, suggesting that this compound may function as a reversible inhibitor as well. Furthermore, the inactivation of the intracellular enzyme by (S)DHPA is remarkable in the light of the fact that this compound induces no inactivation of purified AdoHcy hydrolase, but merely functions as an inhibitor of the enzyme. At low concentration of D-eritadenine (less than 6 microM), a distinct lag period could be demonstrated before accumulation of AdoHcy occurred. This suggests that the AdoHcy hydrolase activity must be decreased below a certain level to cause an increase in cellular AdoHcy. None of the analogues tested completely inactivated AdoHcy hydrolase and a residual enzyme activity was observed. The adenosine deaminase inhibitor, 2'-deoxycoformycin, did not potentiate the effect of these compounds on AdoHcy catabolism. The inactive enzyme formed in the presence of aliphatic adenine analogues was not reactivated under conditions where the inactivation induced by 9-beta-D-arabinofuranosyladenine was reversible.

Inactivation and reactivation of intracellular S-adenosylhomocysteinase in the presence of nucleoside analogues in rat hepatocytes.

Several nucleoside analogues, some of which are potent inactivators of isolated S-adenosylhomocysteinase (AdoHcyase), were tested with respect to their effect on intracellular AdoHcyase and S-adenosylhomocysteine (AdoHcy) catabolism in intact rat hepatocytes. Among the analogues tested, 9-beta-D-arabinofuranosyladenine and 9-beta-D-arabinofuranosyl-3-deazaadenine were the most potent inactivators of intracellular AdoHcyase. Compounds like 2-chloroadenosine and carbocyclic adenosine are extremely efficient inactivators of the isolated enzyme, but these nucleosides exerted only a limited effect on the enzyme in intact hepatocytes. Only a moderate effect was observed with 2-chloro-3-deazaadenosine and 2'-deoxyadenosine, and a high concentration of 5'-deoxy-5'-methylthioadenosine was required to inactivate the enzyme. There was a correlation between inactivation and accumulation of AdoHcy. Carbocyclic-3-deazaadenosine caused no inactivation of AdoHcyase in liver cells but caused a massive accumulation of AdoHcy, suggesting that this analogue functions as a reversible inhibitor of the enzyme. Residual enzyme activity was observed with all the nucleoside analogues tested. The intracellular enzyme inactivated in the presence of 2'-deoxyadenosine and 9-beta-D-arabinofuranosyladenine was readily recovered when the cells were transferred to fresh medium containing adenosine deaminase, but reactivation of the enzyme activity after treatment of the cells with 2-chloroadenosine and 5'-deoxy-5'-methylthioadenosine was also observed. The inactivation of intracellular enzyme induced by 2-chloro-3-deazaadenosine, 9-beta-D-arabinofuranosyl-3-deazaadenine, and carbocyclic adenosine was essentially irreversible under the conditions of the experiments. Factors determining the effect of nucleoside analogues on intracellular AdoHcyase are discussed.

Binding of S-adenosylhomocysteine to various domains of the plasma membrane and to the endoplasmic reticulum from rat liver: relation between binding and phospholipid methyltransferase activity.

S-Adenosylhomocysteine (AdoHcy) binding to various membrane fractions of rat liver was determined at pH 7.4, using an oil centrifugation technique. The highest binding activity was found in the heavy microsomal (M-H) fraction enriched in endoplasmic reticulum, but high binding activity was also observed in the light microsomal fractions enriched in blood sinusoidal membranes (M-L fraction), and the heavy nuclear fraction (N-H fraction) containing the contiguous area. A substantial portion of AdoHcy binding activity in the M-L fraction may be ascribed to contamination of this fraction with endoplasmic reticulum, as indicated by the distribution of NADPH cytochrome c reductase activity. Binding activity was low in the light nuclear (N-L) fraction corresponding to the bile canaliculi. Phospholipid methyltransferase activity was determined in the same membrane fractions under similar conditions (pH 7.4), and in the absence and presence of added phospholipids. The distribution of the enzyme activity was dependent on the presence of exogenous phospholipids, and grossly similar to AdoHcy binding, the highest activities being observed in the M-H and the M-L fractions. The N-H fraction, rich in AdoHcy-binding activity, demonstrated, however, a very low phospholipid methyltransferase activity. It is concluded that AdoHcy-binding activity is not confined to the plasma membranes, and a major fraction of the binding activity resides on membranes derived from the endoplasmic reticulum. Also, the present results add to previous data suggesting that phospholipid methyltransferase does not totally account for the AdoHcy-binding sites on rat liver membranes.

Inhibition of phospholipid methylation in isolated rat hepatocytes by analogues of adenosine and S-adenosylhomocysteine.

Phospholipid methylation in isolated hepatocytes was inhibited in the presence of 3-deazaadenosine (ID50=1.7 microM) 9-beta-D-arabinofuranosyladenine (ID50=6.0 microM), S-tubercidinylhomocysteine (ID50=30 microM), and 5'-deoxy-5'-isobutylthioadenosine (ID50=177 microM). A transient inhibitory effect was observed with adenosine, whereas S-adenosyl-L-homocysteine and Sinefungin were essentially without effect. The inhibition of phospholipid methylation by S-tubercidinylhomocysteine and 9-beta-D-arabinofuranosyladenine showed a lag-phase, whereas the effect of the other inhibitors was apparent within a few minutes. Cells exposed to 9-beta-D-arabinofuranosyladenine or 3-deazaadenosine accumulated large amounts of AdoHcy, and adenosine induced a transient increase in the AdoHcy level. In addition, 3-deazaadenosine served as a precursor for the formation of S-3-deazaadenosylhomocysteine, which accumulated rapidly in cells exposed to this agent. The inhibitory effects of 3-deazaadenosine, 9-beta-arabinofuranosyladenine and adenosine could be explained by the increase in total nucleosidylhomocysteine induced by these agents. In contrast, only a slight (less than 2-fold) increase in S-adenosyl-L-homocysteine content was observed in hepatocytes treated with 5'-deoxy-5'-isobutylthioadenosine, and this metabolic effect could not explain the inhibition of phospholipid methylation induced by this agent. None of the compounds tested reduced the amount nor the specific radioactivity of S-adenosylmethionine. Biological processes determining the inhibitory effects of adenosine, S-adenosyl-L-homocysteine and their analogues on phospholipid methylation in intact cells are discussed.