Ouabain induces apoptotic cell death in human prostate DU 145 cancer cells through DNA damage and TRAIL pathways.

Abstract

Ouabain, a cardiotonic steroid and specific Na+ /K+ -ATPase inhibitor, has a potential to induce cancer cell apoptosis but the mechanisms of apoptosis induced by ouabain are not fully understand. The aim of this study was to investigate the cytotoxic effects of ouabain on human prostate cancer DU 145 cells in vitro. Cell morphological changes were examined by phase contrast microscopy. Cell viability, cell cycle distribution, cell apoptosis, DNA damage, the production of ROS and Ca2+ , and mitochondrial membrane potential (ΔΨm ) were measured by flow cytometry assay. Results indicated that ouabain induced cell morphological changes, decreased total cell viability, induced G0/G1 phase arrest, DNA damage, and cell apoptosis, increased ROS and Ca2+ production, but decreased the levels of ΔΨm in DU 145 cells. Ouabain also increased the activities of caspase-3, -8, and -9. Western blotting was used for measuring the alterations of apoptosis-associated protein expressions in DU 145 cells and results indicated that ouabain increased the expression of DNA damage associated proteins (pATMSer1981 , p-H2A.XSer139 , and p-p53Ser15 ) and ER-stress-associated proteins (Grp78, ATF6β, p-PERKThr981 , PERK, eIF2A, GADD153, CaMKIIβ, and caspase-4) in time-dependently. Furthermore, ouabain increased apoptosis-associated proteins (DR4, DR5, Fas, Fas Ligand, and FADD), TRAIL pathway, which related to extrinsic pathway, promoted the pro-apoptotic protein Bax, increased apoptotic-associated proteins, such as cytochrome c, AIF, Endo G, caspase-3, -8, and -9, but reduced anti-apoptotic protein Bcl-2 and Bcl-x in DU 145 cells. In conclusion, we may suggest that ouabain decreased cell viability and induced apoptotic cell death may via caspase-dependent and mitochondria-dependent pathways in human prostate cancer DU 145 cells.

• Authors: Yi-Ming Chang, Yung-Luen Shih, Chao-Ping Chen, Ko-Lin Liu, Mei-Hui Lee, Ming-Zhe Lee, Hsin-Tu Hou, Hsieh-Chou Huang, Hsu-Feng Lu, Shu-Fen Peng, Kuo-Wei Chen, Ming-Yang Yeh, Jing-Gung Chung
• Journal: Environmental toxicology (Environ Toxicol) Vol. 34 Issue 12 Pg. 1329-1339 (Dec 2019) ISSN: 1522-7278 [Electronic] United States
• PMID: 31436044 (Publication Type: Journal Article)
• Copyright: © 2019 Wiley Periodicals, Inc.

Chemical References

• Cardiotonic Agents
• Endoplasmic Reticulum Chaperone BiP
• HSPA5 protein, human
• Proto-Oncogene Proteins c-bcl-2
• Reactive Oxygen Species
• TNF-Related Apoptosis-Inducing Ligand
• bcl-2-Associated X Protein
• Ouabain
• Caspase 3

Topics

• Apoptosis (drug effects)
• Cardiotonic Agents (pharmacology)
• Caspase 3 (metabolism)
• Cell Line, Tumor
• DNA Damage (drug effects)
• Endoplasmic Reticulum Chaperone BiP
• G1 Phase Cell Cycle Checkpoints (drug effects)
• Humans
• Male
• Membrane Potential, Mitochondrial (drug effects)
• Ouabain (pharmacology)
• Prostatic Neoplasms (metabolism, pathology)
• Proto-Oncogene Proteins c-bcl-2 (metabolism)
• Reactive Oxygen Species (metabolism)
• TNF-Related Apoptosis-Inducing Ligand (metabolism)
• bcl-2-Associated X Protein (metabolism)