Activation of AMP-activated protein kinase by a plant-derived dihydroisosteviol in human intestinal epithelial cell.
Our previous study has shown that dihydroisosteviol (DHIS), a derivative of stevioside isolated from Stevia rebaudiana (Bertoni), inhibits cystic fibrosis transmembrane conductance regulator (CFTR)-mediated transepithelial chloride secretion across monolayers of human intestinal epithelial (T84) cells and prevents cholera toxin-induced intestinal fluid secretion in mouse closed loop models. In this study, we aimed to investigate a mechanism by which DHIS inhibits CFTR activity. Apical chloride current measurements in Fisher rat thyroid cells stably transfected with wild-type human CFTR (FRT-CFTR cells) and T84 cells were used to investigate mechanism of CFTR inhibition by DHIS. In addition, effect of DHIS on AMP-activated protein kinase (AMPK) activation was investigated using Western blot analysis. Surprisingly, it was found that DHIS failed to inhibit CFTR-mediated apical chloride current in FRT-CFTR cells. In contrast, DHIS effectively inhibited CFTR-mediated apical chloride current induced by a cell permeable cAMP analog CPT-cAMP and a direct CFTR activator genistein in T84 cell monolayers. Interestingly, this inhibitory effect of DHIS on CFTR was significantly (p<0.05) reduced by pretreatment with compound C, an AMPK inhibitor. AICAR, a known AMPK activator, was able to inhibit CFTR activity in both FRT-CFTR and T84 cells. Western blot analysis showed that DHIS induced AMPK activation in T84 cells, but not in FRT-CFTR cells. Our results indicate that DHIS inhibits CFTR-mediated chloride secretion in T84 cells, in part, by activation of AMPK activity. DHIS therefore represents a novel candidate of AMPK activators.
Department of Physiology, Faculty of Science, Mahidol University, Rama 6 Road, Rajathevi, Bangkok 10400, Thailand. chatchai.mua@mahidol.ac.th
Chatchai
C
Muanprasat
Lalida
L
Sirianant
Sutthipong
S
Sawasvirojwong
Sureeporn
S
Homvisasevongsa
Apichart
A
Suksamrarn
Varanuj
V
Chatsudthipong
D055372:AMP-Activated Protein Kinases / Q000378:metabolism*; D000818:Animals; D002460:Cell Line; D002712:Chlorides / Q000378:metabolism; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000037:antagonists & inhibitors* / Q000378:metabolism; D045786:Diterpenes, Kaurane / Q000494:pharmacology*; D020536:Enzyme Activators / Q000494:pharmacology*; D004847:Epithelial Cells / Q000187:drug effects* / Q000378:metabolism; D006801:Humans; D007413:Intestinal Mucosa / Q000166:cytology; D051381:Rats; D036625:Stevia
C505032:CFTR protein, human; C579654:CFTR protein, rat; D002712:Chlorides; D045786:Diterpenes, Kaurane; D020536:Enzyme Activators; C583718:dihydroisosteviol; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D055372:AMP-Activated Protein Kinases