Epi -reevesioside F inhibits Na + /K + -ATPase, causing cytosolic acidification, Bak activation and apoptosis in glioblastoma

// Jui-Ling Hsu 1 , Fan-Lun Liu 1 , Lih-Ching Hsu 1 , Hsun-Shuo Chang 2,3 , Wohn-Jenn Leu 1 , Chia-Chun Yu 1 , Wei-Ling Chang 4 , Ih-Sheng Chen 2,3 , Fan-Lu Kung 1 and Jih-Hwa Guh 1 1 School of Pharmacy, National Taiwan University, Taipei, Taiwan 2 School of Pharmacy, College of Pharmacy, Kaohsiung Medical University, Kaohsiung, Taiwan 3 Graduate Institute of Natural Products, College of Pharmacy, Kaohsiung Medical University, Kaohsiung, Taiwan 4 The Division of Medicinal Chemistry, College of Pharmacy, The Ohio State University, Columbus, OH, USA Correspondence to: Jih-Hwa Guh, email: // Fan-Lu Kung, email: // Keywords : Epi-reevesioside F, intracellular Na + concentration, cytosolic acidification, mitochondrial dysfunction, bak activation Received : March 23, 2015 Accepted : June 04, 2015 Published : June 10, 2015 Abstract Epi -reevesioside F, a new cardiac glycoside isolated from the root of Reevesia formosana , displayed potent activity against glioblastoma cells. Epi -reevesioside F was more potent than ouabain with IC 50 values of 27.3±1.7 vs. 48.7±1.8 nM ( P < 0.001) and 45.0±3.4 vs. 81.3±4.3 nM ( P < 0.001) in glioblastoma T98 and U87 cells, respectively. However, both Epi -reevesioside F and ouabain were ineffective in A172 cells, a glioblastoma cell line with low Na + /K + -ATPase α3 subunit expression. Epi -reevesioside F induced cell cycle arrest at S and G2 phases and apoptosis. It also induced an increase of intracellular concentration of Na + but not Ca 2+ , cleavage and exposure of N-terminus of Bak, loss of mitochondrial membrane potential, inhibition of Akt activity and induction of caspase cascades. Potassium supplements significantly inhibited Epi -reevesioside F-induced effects. Notably, Epi -reevesioside F caused cytosolic acidification that was highly correlated with the anti-proliferative activity. In summary, the data suggest that Epi -reevesioside F inhibits Na + /K + -ATPase, leading to overload of intracellular Na + and cytosolic acidification, Bak activation and loss of mitochondrial membrane potential. The PI3-kinase/Akt pathway is inhibited and caspase-dependent apoptosis is ultimately triggered in Epi -reevesioside F-treated glioblastoma cells.