title: Salinomycin inhibits proliferation and induces apoptosis of human hepatocellular carcinoma cells in vitro and in vivo.
abstract: The anti-tumor antibiotic salinomycin (Sal) was recently identified as a selective inhibitor of breast cancer stem cells; however, the effect of Sal on hepatocellular carcinoma (HCC) is not clear. This study aimed to determine the anti-tumor efficacy and mechanism of Sal on HCC. HCC cell lines (HepG2, SMMC-7721, and BEL-7402) were treated with Sal. Cell doubling time was determinated by drawing growth curve, cell viability was evaluated using the Cell Counting Kit 8. The fraction of CD133(+) cell subpopulations was assessed by flow cytometry. We found that Sal inhibits proliferation and decreases PCNA levels as well as the proportion of HCC CD133(+)cell subpopulations in HCC cells. Cell cycle was analyzed using flow cytometry and showed that Sal caused cell cycle arrest of the various HCC cell lines in different phases. Cell apoptosis was evaluated using flow cytometry and Hoechst 33342 staining. Sal induced apoptosis as characterized by an increase in the Bax/Bcl-2 ratio. Several signaling pathways were selected for further mechanistic analyses using real time-PCR and Western blot assays. Compared to control, β-catenin expression is significantly down-regulated upon Sal addition. The Ca(2+) concentration in HCC cells was examined by flow cytometry and higher Ca(2+) concentrations were observed in Sal treatment groups. The anti-tumor effect of Sal was further verified in vivo using the hepatoma orthotopic tumor model and the data obtained showed that the size of liver tumors in Sal-treated groups decreased compared to controls. Immunohistochemistry and TUNEL staining also demonstrated that Sal inhibits proliferation and induces apoptosis in vivo. Finally, the role of Sal on in vivo Wnt/β-catenin signaling was evaluated by Western blot and immunohistochemistry. This study demonstrates Sal inhibits proliferation and induces apoptosis of HCC cells in vitro and in vivo and one potential mechanism is inhibition of Wnt/β-catenin signaling via increased intracellular Ca(2+) levels.
authors.affiliation: Department of Gastroenterology, Shanghai tenth People's Hospital, Tongji University of Medicine, Shanghai, People's Republic of China.
authors.forename: Fan
authors.initials: F
authors.lastname: Wang
authors.forename: Lei
authors.initials: L
authors.lastname: He
authors.forename: Wei-Qi
authors.initials: WQ
authors.lastname: Dai
authors.forename: Ya-Ping
authors.initials: YP
authors.lastname: Xu
authors.forename: Dong
authors.initials: D
authors.lastname: Wu
authors.forename: Chun-Lei
authors.initials: CL
authors.lastname: Lin
authors.forename: Shu-Mei
authors.initials: SM
authors.lastname: Wu
authors.forename: Ping
authors.initials: P
authors.lastname: Cheng
authors.forename: Yan
authors.initials: Y
authors.lastname: Zhang
authors.forename: Miao
authors.initials: M
authors.lastname: Shen
authors.forename: Chen-Feng
authors.initials: CF
authors.lastname: Wang
authors.forename: Jie
authors.initials: J
authors.lastname: Lu
authors.forename: Ying-Qun
authors.initials: YQ
authors.lastname: Zhou
authors.forename: Xuan-Fu
authors.initials: XF
authors.lastname: Xu
authors.forename: Ling
authors.initials: L
authors.lastname: Xu
authors.forename: Chuan-Yong
authors.initials: CY
authors.lastname: Guo
mesh_terms: D000071916:AC133 Antigen; D000818:Animals; D015703:Antigens, CD / Q000378:metabolism; D000970:Antineoplastic Agents / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects*; D002118:Calcium / Q000378:metabolism; D006528:Carcinoma, Hepatocellular / Q000473:pathology*; D059447:Cell Cycle Checkpoints / Q000187:drug effects; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D006023:Glycoproteins / Q000378:metabolism; D006801:Humans; D042541:Intracellular Space / Q000187:drug effects / Q000378:metabolism; D008113:Liver Neoplasms / Q000473:pathology*; D008297:Male; D051379:Mice; D010455:Peptides / Q000378:metabolism; D011714:Pyrans / Q000494:pharmacology*; D015398:Signal Transduction / Q000187:drug effects; D051153:Wnt Proteins / Q000378:metabolism; D023041:Xenograft Model Antitumor Assays; D051176:beta Catenin / Q000378:metabolism; D000095662:Polyether Polyketides
chemical_list: D000071916:AC133 Antigen; D015703:Antigens, CD; D000970:Antineoplastic Agents; D002118:Calcium; D006023:Glycoproteins; D010455:Peptides; D011714:Pyrans; D051153:Wnt Proteins; D051176:beta Catenin; C000606023:PROM1 protein, human; C000606024:Prom1 protein, mouse; C010327:salinomycin; D000095662:Polyether Polyketides