title: Inhibition of apical Cl-/OH- exchange activity in Caco-2 cells by phorbol esters is mediated by PKCepsilon.
abstract: The present studies were undertaken to examine the possible regulation of apical membrane Cl-/OH- exchanger in Caco-2 cells by protein kinase C (PKC). The effect of the phorbol ester phorbol 12-myristate 13-acetate (PMA), an in vitro PKC agonist, on OH- gradient-driven 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS)-sensitive 36Cl uptake in Caco-2 cells was assessed. The results demonstrated that PMA decreased apical Cl-/OH- exchanger activity via phosphatidylinositol 3-kinase (PI3-kinase)-mediated activation of PKCepsilon. The data consistent with these conclusions are as follows: 1) short-term treatment of cells for 1-2 h with PMA (100 nM) significantly decreased Cl-/OH- exchange activity compared with control (4alpha-PMA); 2) pretreatment of cells with specific PKC inhibitors chelerythrine chloride, calphostin C, and GF-109203X completely blocked the inhibition of Cl-/OH- exchange activity by PMA; 3) specific inhibitors for PKCepsilon (Ro-318220) but not PKCalpha (Go-6976) significantly blocked the PMA-mediated inhibition; 4) specific PI3-kinase inhibitors wortmannin and LY-294002 significantly attenuated the inhibitory effect of PMA; and 5) PI3-kinase activators IRS-1 peptide and phosphatidylinositol 3,4,5-trisphosphate [PI(3,4,5)P(3)] mimicked the effects of PMA. These findings provide the first evidence for PKCepsilon-mediated inhibition of Cl-/OH- exchange activity in Caco-2 cells and indicate the involvement of the PI3-kinase-mediated pathways in the regulation of Cl- absorption in intestinal epithelial cells.
authors.affiliation: Section of Digestive and Liver Diseases, Department of Medicine, University of Illinois at Chicago and West Side Department of Veterans Affairs Medical Center, Chicago, Illinois 60612, USA.
authors.forename: Seema
authors.initials: S
authors.lastname: Saksena
authors.forename: Ravinder K
authors.initials: RK
authors.lastname: Gill
authors.forename: Irfan A
authors.initials: IA
authors.lastname: Syed
authors.forename: Sangeeta
authors.initials: S
authors.lastname: Tyagi
authors.forename: Waddah A
authors.initials: WA
authors.lastname: Alrefai
authors.forename: K
authors.initials: K
authors.lastname: Ramaswamy
authors.forename: Pradeep K
authors.initials: PK
authors.lastname: Dudeja
mesh_terms: D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid / Q000494:pharmacology; D000730:Androstadienes / Q000494:pharmacology; D001692:Biological Transport / Q000187:drug effects / Q000502:physiology; D018938:Caco-2 Cells; D002273:Carcinogens / Q000494:pharmacology*; D027963:Chloride-Bicarbonate Antiporters / Q000037:antagonists & inhibitors* / Q000378:metabolism; D002712:Chlorides / Q000378:metabolism; D002867:Chromones / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D055504:Insulin Receptor Substrate Proteins; D007527:Isoenzymes / Q000378:metabolism*; D009025:Morpholines / Q000494:pharmacology; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D018129:Phosphatidylinositol Phosphates / Q000494:pharmacology; D000081082:Phosphoinositide-3 Kinase Inhibitors; D010750:Phosphoproteins / Q000494:pharmacology; D011493:Protein Kinase C / Q000378:metabolism*; D051744:Protein Kinase C-epsilon; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*; D000077191:Wortmannin
chemical_list: D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid; D000730:Androstadienes; D002273:Carcinogens; D027963:Chloride-Bicarbonate Antiporters; D002712:Chlorides; D002867:Chromones; D004791:Enzyme Inhibitors; D055504:Insulin Receptor Substrate Proteins; D007527:Isoenzymes; D009025:Morpholines; D019869:Phosphatidylinositol 3-Kinases; D018129:Phosphatidylinositol Phosphates; D000081082:Phosphoinositide-3 Kinase Inhibitors; D010750:Phosphoproteins; D011493:Protein Kinase C; D051744:Protein Kinase C-epsilon; D013755:Tetradecanoylphorbol Acetate; D000077191:Wortmannin; C526292:IRS1 protein, human; C060974:phosphatidylinositol 3,4,5-triphosphate; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; C497255:PRKCE protein, human