Protein kinase B/Akt mediates cAMP- and cell swelling-stimulated Na+/taurocholate cotransport and Ntcp translocation.
Cyclic AMP and cell swelling stimulate hepatic Na+/TC cotransport and Ntcp translocation via the phosphoinositide 3-kinase signaling pathway. To determine the downstream target of the phosphoinositide 3-kinase action, we examined the role of protein kinase B (PKB)/Akt using SB203580 in hepatocytes as well as by transfection with a dominant negative (DN-PKB) or a constitutively active (CA-PKB) form of PKB in HuH-Ntcp cells. Both cAMP and cell swelling stimulated p38 mitogen-activated protein (MAP) kinase as well as PKB activity. Although 100 microm SB203580 inhibited cell swelling- and 8-chlorophenylthio-cAMP-induced activation of both p38 MAP kinase and PKB, 1 microm SB203580 inhibited activation of p38 MAP kinase, but not of PKB, in hepatocytes. 100 microm, but not 1 microm SB203580, inhibited cell swelling- and cAMP-induced increases in taurocholate (TC) uptake and Ntcp translocation in hepatocytes. TC uptake in HuH-Ntcp cells was more than 90% dependent on extracellular Na+. Cyclic AMP and cell swelling increased TC uptake by 50-100% and PKB activity 2-4-fold in HuH-Ntcp cells transfected with the empty vector and failed to increase PKB activity, TC uptake, and Ntcp translocation in DN-PKB-transfected HuH-Ntcp cells. Transfection with CA-PKB increased PKB activity, TC uptake, and Ntcp translocation in HuH-Ntcp cells compared with cells transfected with the empty vector. In contrast, transfection with DN-PKB did not affect basal PKB activity, TC uptake, or Ntcp translocation. Taken together, these results strongly suggest that cell swelling and cAMP-mediated stimulation of hepatic Na+/TC cotransport and Ntcp translocation requires activation of PKB and is mediated at least in part via a phosphoinositide 3-kinase/PKB-signaling pathway.
Webster
Cynthia R L
CR
Department of Clinical Sciences, Tufts University School of Veterinary Medicine, North Grafton, Massachusetts 01536, USA.
Srinivasulu
Usha
U
Ananthanarayanan
Meenakshisundaram
M
Suchy
Frederick J
FJ
Anwer
M Sawkat
MS
D000818:Animals; D002352:Carrier Proteins / Q000378:metabolism*; D002478:Cells, Cultured; D002756:Cholagogues and Choleretics / Q000494:pharmacology; D000242:Cyclic AMP / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D022781:Hepatocytes / Q000378:metabolism*; D007093:Imidazoles / Q000494:pharmacology; D008099:Liver / Q000166:cytology; D026901:Membrane Transport Proteins; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D029363:Organic Anion Transporters, Sodium-Dependent; D011485:Protein Binding; D017346:Protein Serine-Threonine Kinases; D021381:Protein Transport; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D051057:Proto-Oncogene Proteins c-akt; D011725:Pyridines / Q000494:pharmacology; D051381:Rats; D015398:Signal Transduction; D012964:Sodium / Q000378:metabolism*; D027981:Symporters; D013656:Taurocholic Acid / Q000378:metabolism*; D014162:Transfection; D048051:p38 Mitogen-Activated Protein Kinases
D002352:Carrier Proteins; D002756:Cholagogues and Choleretics; D004791:Enzyme Inhibitors; D007093:Imidazoles; D026901:Membrane Transport Proteins; D029363:Organic Anion Transporters, Sodium-Dependent; D011518:Proto-Oncogene Proteins; D011725:Pyridines; D027981:Symporters; C071908:sodium-bile acid cotransporter; D013656:Taurocholic Acid; D012964:Sodium; D000242:Cyclic AMP; C494920:Akt1 protein, rat; D017346:Protein Serine-Threonine Kinases; D051057:Proto-Oncogene Proteins c-akt; D020928:Mitogen-Activated Protein Kinases; D048051:p38 Mitogen-Activated Protein Kinases; C093642:SB 203580