肝潅流法による胆汁酸の取り込み機構
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The overall transport of bile acids from the blood into the bile is known to be a saturable process, but mechanisms of uptake by hepatocytes and excretion into bile remain to be clarified. In general, bile acid metabolism is regulated by the following three processes: hepatic uptake, intracellular transport and excretion into bile. We investigated the mechanism of radioactive taurocholic acid (TCA) uptake by rat liver and excretion into bile using the flowthrough perfusion technique. Red blood cells and albumin-free Krebs-Henseleit buffer containing TCA were used for perfusion under aeration of 95% 0<SUB>2</SUB>-5% C0<SUB>2</SUB>. Fluid was pumped through the liver at a constantflow rate (24 m<I>l</I>/min at 37°C) . Perf usate samples were collected each 5 min as a fraction from the vena cava inferior. Bile was collected from a catheter inserted in the common bile duct each 5 min as one fraction. We found that bromsulf ophthalein (BSP) has a delay effect on the disappearance of radioactive cholic acid injected<I>in vivo</I>. This result indicated that BSP should affect hepatic uptake of cholic acid and/or excretion into the bile. When rat liver was Perf used with solution containing TCA, it was taken up by a saturable process depending on both the perfusion period and substrate concentration. It was considered that TCA incorporated during 5 min of perfusion should represent the amount of TCA taken up by hepatocytes located very closely to the blood stream. Since TCA uptake for 5 min exhibited a saturable kinesis, the Km obtained by Lineweaver-Burk plot was estimated to be 8.8×10<SUP>-6</SUP>μmol/g liver/min with Vmax 3.96 pmol/g liver. This uptake was inhibited by a low concentration of BSP. It was also found that taurochenodeoxy cholic acid inhibited the uptake in a competitive manner. These results suggested that TCA was taken up by hepatocytes in a carrier-mediated manner. When the TCA concentration in cytosol fraction was measured after 30 min of perfusion, the uptake was found to be a saturable mechanism. The Km of TCA uptake for binding proteins in the cytosol was approximately 1.75×10<SUP>-4</SUP>μmol/g liver. Although BSP inhibited hepatic TCA uptake, it did not alter the amount of TCA excreted into bile ; BSP only delayed the time until the maximal excretion rate was reached.
- 学校法人 昭和大学・昭和医学会の論文
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