title: Saikosaponin d induces cell death through caspase-3-dependent, caspase-3-independent and mitochondrial pathways in mammalian hepatic stellate cells.
abstract: Saikosaponin d (SSd) is one of the main active triterpene saponins in Bupleurum falcatum. It has a steroid-like structure, and is reported to have pharmacological activities, including liver protection in rat, cell cycle arrest and apoptosis induction in several cancer cell lines. However, the biological functions and molecular mechanisms of mammalian cells under SSd treatment are still unclear.
  
  
  
  The cytotoxicity and apoptosis of hepatic stellate cells (HSCs) upon SSd treatment were discovered by MTT assay, colony formation assay and flow cytometry. The collage I/III, caspase activity and apoptotic related genes were examined by quantitative PCR, Western blotting, immunofluorescence and ELISA. The mitochondrial functions were monitored by flow cytometry, MitoTracker staining, ATP production and XF24 bioenergetic assay.
  
  
  
  This study found that SSd triggers cell death via an apoptosis path. An example of this path might be typical apoptotic morphology, increased sub-G1 phase cell population, inhibition of cell proliferation and activation of caspase-3 and caspase-9. However, the apoptotic effects induced by SSd are partially blocked by the caspase-3 inhibitor, Z-DEVD-FMK, suggesting that SSd may trigger both HSC-T6 and LX-2 cell apoptosis through caspase-3-dependent and independent pathways. We also found that SSd can trigger BAX and BAK translocation from the cytosol to the mitochondria, resulting in mitochondrial function inhibition, membrane potential disruption. Finally, SSd also increases the release of apoptotic factors.
  
  
  
  The overall analytical data indicate that SSd-elicited cell death may occur through caspase-3-dependent, caspase-3-independent and mitochondrial pathways in mammalian HSCs, and thus can delay the formation of liver fibrosis by reducing the level of HSCs.
authors.affiliation: Department of Gastroenterology and Hepatology, E-DA Hospital, Kaohsiung, Taiwan.|Graduate Institute of Integrated Medicine, China Medical University, Taichung, Taiwan.
authors.forename: Ming-Feng
authors.initials: MF
authors.lastname: Chen
authors.affiliation: Department of Obstetrics and Gynecology, E-Da Hospital, I-Shou University, Kaohsiung, Taiwan.|Department of Obstetrics and Gynecology, University of South Florida, College of Medicine, Tampa, FL, USA.
authors.forename: S Joseph
authors.initials: SJ
authors.lastname: Huang
authors.affiliation: Department of Pathology, Kaohsiung Chang Gung Memorial Hospital, Chang Gung University College of Medicine, Kaohsiung, Taiwan.|Tissue Bank and Biobank, Kaohsiung Chang Gung Memorial Hospital, Kaohsiung, Taiwan.
authors.forename: Chao-Cheng
authors.initials: CC
authors.lastname: Huang
authors.affiliation: Department of Microbiology, Soochow University, Shihlin, Taipei, Taiwan.
authors.forename: Pei-Shan
authors.initials: PS
authors.lastname: Liu
authors.affiliation: Departments of Obstetrics & Gynecology, Chang Bing Show Chwan Memorial Hospital, Lukang Zhen, Changhua County, Taiwan.
authors.forename: Kun-I
authors.initials: KI
authors.lastname: Lin
authors.affiliation: School of Pharmacy, Kaohsiung Medical University, Kaohsiung, Taiwan.
authors.forename: Ching-Wen
authors.initials: CW
authors.lastname: Liu
authors.affiliation: Department of Biological Science & Technology, I-SHOU University, Kaohsiung, Taiwan.
authors.forename: Wen-Chuan
authors.initials: WC
authors.lastname: Hsieh
authors.affiliation: Department of Medical Research, E-Da Hospital, Kaohsiung, Taiwan. her2neu24@gmail.com.|Department of Medical Research, Cell Therapy and Research Center, E-Da Hospital, I-Shou University, No.6, Yida Road, Jiaosu Village, Yanchao District, Kaohsiung City, 82445, Taiwan, People's Republic of China. her2neu24@gmail.com.
authors.forename: Li-Yen
authors.initials: LY
authors.lastname: Shiu
authors.affiliation: Institute for Translational Research in Biomedicine, Kaohsiung Chang Gung Memorial Hospital, 123 Ta-Pei Road, Niaosong District, Kaohsiung City, Taiwan, People's Republic of China. chench7@gmail.com.|Department of Applied Chemistry, National Chi Nan University, Nantou, Taiwan. chench7@gmail.com.|Center for Infectious Disease and Cancer Research, Kaohsiung Medical University, Kaohsiung, Taiwan. chench7@gmail.com.
authors.forename: Chang-Han
authors.initials: CH
authors.lastname: Chen
mesh_terms: D000818:Animals; D000972:Antineoplastic Agents, Phytogenic / Q000494:pharmacology* / Q000627:therapeutic use; D017209:Apoptosis / Q000187:drug effects*; D029972:Bupleurum / Q000737:chemistry; D053148:Caspase 3 / Q000378:metabolism*; D053453:Caspase 9 / Q000378:metabolism*; D061945:Caspase Inhibitors / Q000494:pharmacology; D002460:Cell Line; D049109:Cell Proliferation / Q000187:drug effects; D004365:Drugs, Chinese Herbal / Q000494:pharmacology / Q000627:therapeutic use; D055166:Hepatic Stellate Cells / Q000187:drug effects* / Q000378:metabolism; D006801:Humans; D008103:Liver Cirrhosis / Q000188:drug therapy; D008928:Mitochondria / Q000187:drug effects* / Q000378:metabolism; D009828:Oleanolic Acid / Q000031:analogs & derivatives* / Q000494:pharmacology / Q000627:therapeutic use; D009842:Oligopeptides / Q000494:pharmacology; D051381:Rats; D012503:Saponins / Q000494:pharmacology* / Q000627:therapeutic use; D014315:Triterpenes / Q000494:pharmacology* / Q000627:therapeutic use
chemical_list: D000972:Antineoplastic Agents, Phytogenic; D061945:Caspase Inhibitors; D004365:Drugs, Chinese Herbal; D009842:Oligopeptides; D012503:Saponins; D014315:Triterpenes; C110772:benzoylcarbonyl-aspartyl-glutamyl-valyl-aspartyl-fluoromethyl ketone; D009828:Oleanolic Acid; C505407:CASP3 protein, human; C506433:CASP9 protein, human; C505410:Casp3 protein, rat; C506435:Casp9 protein, rat; D053148:Caspase 3; D053453:Caspase 9; C025759:saikosaponin D