Tauroursodeoxycholic acid mobilizes α-PKC after uptake in human HepG2 hepatoma cells

Background Thuroursodeoxycholic acid (TUDCA) may exert anticholestatic effects via Ca++- and alpha-protein kinase C (alpha-PKC)-dependent apical vesicular insertion of canalicular transporters in cholestatic hepatocytes (Hepatology 2001; 33: 1206-16). Tauroursodeoxycholic acid is mainly taken up into liver cells by Na+-taurocholate cotransporting polypeptide (Ntcp). Thuroursodeoxycholic acid selectively translocates alpha-PKC, a key mediator of regulated exocytosis, to hepatocellular membranes. It is unclear whether TUDCA exerts its effects on alpha-PKC after carrier-mediated uptake into liver cells or by interaction with extracellular/membraneous structures. Materials and methods Human hepatoblastoma HepG2 cells lacking Ntcp were stably transfected with pcDNA3.1/Ntcp or sham-transfected with pcDNA3.1 [+]. Distribution of alpha-PKC was studied using a Western blotting technique. Uptake of [H-3]taurocholic acid (TCA) was determined radiochemically. Results [H-3] taurocholic acid uptake was approximately 180-fold higher in Ntcp-transfected than in sham-transfected cells. Phorbol 12-myristate 13-acetate (1 mumol L-1; positive control) increased membrane binding of alpha-PKC by 34% in Ntcp-transfected and by 37% in sham-transfected cells. Tauroursodeoxycholic acid (10 mumol L-1) increased membrane-associated alpha-PKC by 19% in Ntcp-transfected, but not in sham-transfected cells (-13%). Taurocholic acid (10 mumol L-1) did not affect the distribution of alpha-PKC. Conclusion Carrier-mediated uptake is a prerequisite for TUDCA-induced translocation of alpha-PKC to hepatocellular membranes.