title: Casticin induces caspase-mediated apoptosis via activation of mitochondrial pathway and upregulation of DR5 in human lung cancer cells.
abstract: OBJECTIVE
  To assess if casticin induces caspase-mediated apoptosis via activation of mitochondrial pathway and upregulation of DR5 in human lung cancer cells.
  
  
  METHODS
  Human non-small-cell lung carcinoma cell lines H460, A549 and H157 were cultured in vitro. The cytotoxic activities were determined using MTT assay. The apoptotic cells death was examined by flow cytometry using PI staining and DNA agarose gel electrophoresis. The activities of caspase-3, -8 and -9 were measured via ELISA. Cellular fractionation was determined by flow cytometry to assess release of cytochrome c and the mitochondrial transmembrane potential. Bcl-2/Bcl-XL/XIAP/Bid/DR5 and DR4 proteins were analyzed using western blot.
  
  
  RESULTS
  The concentrations required for a 50% decrease in cell growth (IC(50)) ranged from 1.8 to 3.2 μM. Casticin induced rapid apoptosis and triggered a series of effects associated with apoptosis by way of mitochondrial pathway, including the depolarization of the mitochondrial membrane, release of cytochrome c from mitochondria, activation of procaspase-9 and -3, and increase of DNA fragments. Moreover, the pan caspase inhibitor zVAD-FMK and the caspase-3 inhibitor zDEVD-FMK suppressed casticin-induced apoptosis. In addition, casticin induced XIAP and Bcl-XL down-regulation, Bax upregulation and Bid clearage. In H157 cell line, casticin increased expression of DR5 at protein levels but not affect the expression of DR4. The pretreatment with DR5/Fc chimera protein effectively attenuated casticin-induced apoptosis in H157 cells. No correlation was found between cell sensitivity to casticin and that to p53 status, suggesting that casticin induce a p53-independent apoptosis.
  
  
  CONCLUSIONS
  Our results demonstrate that casticin induces caspase-mediated apoptosis via activation of mitochondrial pathway and upregulation of DR5 in human lung cancer cells.
authors.affiliation: Department of Pharmacology of Medical College, Hunan Normal University, Changsha, Hunan 410013, China. zhouyuan1122@126.com
authors.forename: Yuan
authors.initials: Y
authors.lastname: Zhou
authors.forename: Yi
authors.initials: Y
authors.lastname: Peng
authors.forename: Qi-Qi
authors.initials: QQ
authors.lastname: Mao
authors.forename: Xia
authors.initials: X
authors.lastname: Li
authors.forename: Ming-Wu
authors.initials: MW
authors.lastname: Chen
authors.forename: Jing
authors.initials: J
authors.lastname: Su
authors.forename: Li
authors.initials: L
authors.lastname: Tian
authors.forename: Nai-Quan
authors.initials: NQ
authors.lastname: Mao
authors.forename: Ling-Zhi
authors.initials: LZ
authors.lastname: Long
authors.forename: Mei-Fang
authors.initials: MF
authors.lastname: Quan
authors.forename: Fei
authors.initials: F
authors.lastname: Liu
authors.forename: Su-Fang
authors.initials: SF
authors.lastname: Zhou
authors.forename: Yong-Xiang
authors.initials: YX
authors.lastname: Zhao
mesh_terms: D017209:Apoptosis / Q000187:drug effects*; D002289:Carcinoma, Non-Small-Cell Lung / Q000188:drug therapy* / Q000201:enzymology / Q000378:metabolism* / Q000473:pathology; D020169:Caspases / Q000378:metabolism*; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D045304:Cytochromes c / Q000378:metabolism; D005419:Flavonoids / Q000494:pharmacology*; D006801:Humans; D008175:Lung Neoplasms / Q000188:drug therapy* / Q000201:enzymology / Q000378:metabolism* / Q000473:pathology; D008928:Mitochondria / Q000187:drug effects* / Q000201:enzymology / Q000378:metabolism; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D053220:Receptors, TNF-Related Apoptosis-Inducing Ligand / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects; D015854:Up-Regulation / Q000187:drug effects
chemical_list: D005419:Flavonoids; D019253:Proto-Oncogene Proteins c-bcl-2; D053220:Receptors, TNF-Related Apoptosis-Inducing Ligand; C105190:TNFRSF10A protein, human; C054133:casticin; D045304:Cytochromes c; D020169:Caspases