title: Dioscin suppresses hepatocellular carcinoma tumor growth by inducing apoptosis and regulation of TP53, BAX, BCL2 and cleaved CASP3.
abstract: BACKGROUND
  Hepatocellular carcinoma (HCC) is the most commonly diagnosed malignancy of the liver, occurs frequently in the setting of chronic liver injury. Although multiple therapeutic approaches are available, the prognosis of patients with HCC remains poor. Dioscin is a natural steroid saponin that presents in various plants. The anti-cancer and anti-fibrotic effects have been extensively reported. However, the effect of dioscin on HCC remains unclear. We aimed to investigate the anti-HCC properties of dioscin in vitro and in vivo.
  
  
  METHODS
  MTT (3-(4,5-dimethylthiazol-2-yl)- 2,5-diphenyl-tetrazolium bromide) assay was used to analyze the growth inhibition activity of Dioscin in human cell lines, Bel-7402, HepG2, Lovo, and EAhy926. Antitumor activity through induction of apoptosis was evaluated by flow cytometry using Annexin-V and propidium iodide (PI) staining, laser scanning confocal microscopy (LSCM) analysis with Hochest33342 and PI labeling, and DNA fragmentation analysis. The expression of apoptosis-related proteins tumor protein p53 (TP53), BCL2-associated X protein (BAX), B-Cell CLL/Lymphoma 2 (BCL2) and Caspase 3 (CASP3) was measured by Western blot. Nude mice bearing Bel-7402 were administered intraperitoneally at different doses of dioscin and 5-FU (5-Fluorouracil) treatment was used as a control. Tumor volume and tumor weight of each mouse were then measured.
  
  
  RESULTS
  We demonstrated that Dioscin inhibited proliferation of HCC cell lines in a dose-dependent manner. Dioscin also significantly induced morphological changes during death by apoptosis and increased DNA damage of Bel-7402 cells. Moreover, we demonstrated that Dioscin displayed anticancer activity via up-regulating expression of TP53, BAX and CASP3 protein, as well as down-regulating BCL2 in Bel-7402 cells. Notably, the in vivo anticancer activity of Dioscin was further assessed and achieved greater inhibition efficiency at the concentration increased to 24mg/kg/day than 5-FU at dose of 10mg/kg/day in nude mice bearing Bel-7402 cells.
  
  
  CONCLUSIONS
  Dioscin inhibited tumor growth via inducing apoptosis, which was accompanied by altered expression of apoptotic pathway proteins, such as TP53, BAX, BCL2 and CASP3. Our findings indicate that further evaluation of dioscin as a novel therapeutic approach for HCC is warranted.
authors.affiliation: School of Fundamental Medical Sciences, Guangzhou University of Chinese Medicine, Guangzhou 510006, China.
authors.forename: Guangxian
authors.initials: G
authors.lastname: Zhang
authors.affiliation: Department of General Surgery, The Second People's Hospital of Guangdong Province, Guangzhou 510317, China.
authors.forename: Xiancheng
authors.initials: X
authors.lastname: Zeng
authors.affiliation: School of Fundamental Medical Sciences, Guangzhou University of Chinese Medicine, Guangzhou 510006, China.
authors.forename: Ren
authors.initials: R
authors.lastname: Zhang
authors.affiliation: School of Fundamental Medical Sciences, Guangzhou University of Chinese Medicine, Guangzhou 510006, China.
authors.forename: Juan
authors.initials: J
authors.lastname: Liu
authors.affiliation: Department of Internal Medicine, Division of Rheumatology, Allergy and Clinical Immunology, University of California Davis, School of Medicine, Davis, CA 95616, USA.
authors.forename: Weici
authors.initials: W
authors.lastname: Zhang
authors.affiliation: Department of General Surgery, The Second People's Hospital of Guangdong Province, Guangzhou 510317, China.
authors.forename: Yujun
authors.initials: Y
authors.lastname: Zhao
authors.affiliation: School of Fundamental Medical Sciences, Guangzhou University of Chinese Medicine, Guangzhou 510006, China.
authors.forename: Xiaoyuan
authors.initials: X
authors.lastname: Zhang
authors.affiliation: School of Fundamental Medical Sciences, Guangzhou University of Chinese Medicine, Guangzhou 510006, China.
authors.forename: Zhixue
authors.initials: Z
authors.lastname: Wu
authors.affiliation: School of Fundamental Medical Sciences, Guangzhou University of Chinese Medicine, Guangzhou 510006, China.
authors.forename: Yuhui
authors.initials: Y
authors.lastname: Tan
authors.affiliation: School of Fundamental Medical Sciences, Guangzhou University of Chinese Medicine, Guangzhou 510006, China.
authors.forename: Yingya
authors.initials: Y
authors.lastname: Wu
authors.affiliation: School of Fundamental Medical Sciences, Guangzhou University of Chinese Medicine, Guangzhou 510006, China. Electronic address: dubiaoyan@gzucm.edu.cn.
authors.forename: Biaoyan
authors.initials: B
authors.lastname: Du
mesh_terms: D000818:Animals; D000964:Antimetabolites, Antineoplastic / Q000494:pharmacology; D000972:Antineoplastic Agents, Phytogenic / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects*; D006528:Carcinoma, Hepatocellular / Q000188:drug therapy* / Q000473:pathology; D053148:Caspase 3 / Q000096:biosynthesis / Q000235:genetics; D045744:Cell Line, Tumor; D053938:DNA Fragmentation / Q000187:drug effects; D004144:Diosgenin / Q000031:analogs & derivatives* / Q000494:pharmacology; D004354:Drug Screening Assays, Antitumor; D005472:Fluorouracil / Q000494:pharmacology; D006801:Humans; D008113:Liver Neoplasms / Q000188:drug therapy* / Q000473:pathology; D008297:Male; D051379:Mice; D008819:Mice, Nude; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000096:biosynthesis / Q000235:genetics; D013778:Tetrazolium Salts; D013844:Thiazoles; D016159:Tumor Suppressor Protein p53 / Q000096:biosynthesis / Q000235:genetics; D051028:bcl-2-Associated X Protein / Q000096:biosynthesis / Q000235:genetics
chemical_list: D000964:Antimetabolites, Antineoplastic; D000972:Antineoplastic Agents, Phytogenic; C494826:BAX protein, human; C000595707:BCL2 protein, human; D019253:Proto-Oncogene Proteins c-bcl-2; C495901:TP53 protein, human; D013778:Tetrazolium Salts; D013844:Thiazoles; D016159:Tumor Suppressor Protein p53; D051028:bcl-2-Associated X Protein; C019357:dioscin; C505407:CASP3 protein, human; D053148:Caspase 3; C022616:thiazolyl blue; D004144:Diosgenin; D005472:Fluorouracil
keywords: Apoptosis; Dioscin; Hepatocellular carcinoma