Mechanism of inhibition of taurolithocholate‐induced retrieval of plasma membrane MRP2 by cyclic AMP and tauroursodeoxycholate

摘要

Taurolithocholate (TLC) produces cholestasis by inhibiting biliary solute secretion in part by retrieving MRP2 from the plasma membrane (PM). Tauroursodeoxycholate (TUDC) and cAMP reverse TLC‐induced cholestasis by inhibiting TLC‐induced retrieval of MRP2. However, cellular mechanisms for this reversal are incompletely understood. Recently, we reported that TLC decreases PM‐MRP2 by activating PKCε followed by phosphorylation of myristoylated alanine‐rich C kinase substrate (MARCKS). Thus, cAMP and TUDC may reverse TLC‐induced cholestasis by inhibiting the style="fixed-case">TLC/ style="fixed-case">PKC ε/ style="fixed-case">MARCKS phosphorylation pathway. We tested this hypothesis by determining whether style="fixed-case">TUDC and/or style="fixed-case">cAMP inhibit style="fixed-case">TLC‐induced activation of style="fixed-case">PKCε and phosphorylation of style="fixed-case">MARCKS. Studies were conducted in HuH‐ style="fixed-case">NTCP cell line and rat hepatocytes. Activation of style="fixed-case">PKC ε was determined from the translocation of style="fixed-case">PKC ε to style="fixed-case">PM using a biotinylation method. Phosphorylation of style="fixed-case">MARCKS was determined by immunoblotting with a phospho‐ style="fixed-case">MARCKS antibody. style="fixed-case">TLC, but not style="fixed-case">cAMP and style="fixed-case">TUDC, activated style="fixed-case">PKC ε and increased style="fixed-case">MARCKS phosphorylation in HuH‐ style="fixed-case">NTCP as well in rat hepatocytes. Treatment with style="fixed-case">TUDC or style="fixed-case">cAMP inhibited style="fixed-case">TLC‐induced activation of style="fixed-case">PKCε and increases in style="fixed-case">MARCKS phosphorylation in both cell types. Based on these results, we conclude that the reversal of style="fixed-case">TLC‐induced cholestasis by style="fixed-case">cAMP and style="fixed-case">TUDC involves, at least in part, inhibition of style="fixed-case">TLC‐mediated activation of the style="fixed-case">PKCε/ style="fixed-case">MARCKS phosphorylation pathway.