title: Involvement of P53 and Bax/Bad triggering apoptosis in thioacetamide-induced hepatic epithelial cells.
abstract: OBJECTIVE
  Thioacetamide (TAA) has been used in studying liver fibrosis and cirrhosis, however, the mechanisms of TAA-induced apoptosis in liver are still unclear. The hepatic epithelial cell line clone 9 was cultured and treated with TAA to investigate the causes of cell death.
  
  
  METHODS
  The cell viability of TAA-induced clone 9 cells was determined using MTT assay. Total cellular GSH in TAA-induced clone 9 cells was measured using a slight modification of the Tietze assay. The activity of caspase 3 in TAA-induced clone 9 cells was monitored by the cleavage of DEVD-p-nitroanaline. TUNEL assay and flow cytometry were applied for the determination of DNA fragmentation and the proportion of apoptosis in TAA-induced clone 9 cells, respectively. The alterations of caspase 3, Bad, Bax and Phospho-P53 contents in TAA-induced clone 9 cells were measured by Western blot.
  
  
  RESULTS
  The experimental data indicated that TAA caused rat hepatic epithelial cell line clone 9 cell death in a dose-and time-dependent manner; 60% of the cells died (MTT assay) within 24 h after 100 mg/L TAA was applied. Apoptotic cell percentage (TUNEL assay) and caspase 3 activities were highest after 100 mg/L TAA was added for 8 h. The release of GSH and the elevation in caspase content after TAA treatment resulted in clone 9 cell apoptosis via oxidative stress and a caspase-dependent mechanism. The phospho-p53, Bax and Bad protein expressions in clone 9 cells were increased after TAA treatment.
  
  
  CONCLUSIONS
  These results reveal that TAA activates p53, increases caspase 3, Bax and Bad protein contents, perhaps causing the release of cytochrome c from mitochondria and the disintegration of membranes, leading to apoptosis of cells.
authors.affiliation: Institute of Biotechnology, National Dong Hwa University, Hualien, Taiwan, China.
authors.forename: Li-Hsuen
authors.initials: LH
authors.lastname: Chen
authors.forename: Chia-Yu
authors.initials: CY
authors.lastname: Hsu
authors.forename: Ching-Feng
authors.initials: CF
authors.lastname: Weng
mesh_terms: D000818:Animals; D017209:Apoptosis; D002470:Cell Survival; D042783:Endothelial Cells / Q000378:metabolism* / Q000473:pathology*; D005355:Fibrosis; D005786:Gene Expression Regulation; D016158:Genes, p53; D006801:Humans; D008099:Liver / Q000187:drug effects* / Q000473:pathology*; D051381:Rats; D013778:Tetrazolium Salts / Q000494:pharmacology; D013844:Thiazoles / Q000494:pharmacology; D013853:Thioacetamide / Q000494:pharmacology*; D016159:Tumor Suppressor Protein p53 / Q000502:physiology*; D051028:bcl-2-Associated X Protein / Q000502:physiology*; D051025:bcl-Associated Death Protein / Q000502:physiology*
chemical_list: D013778:Tetrazolium Salts; D013844:Thiazoles; D016159:Tumor Suppressor Protein p53; D051028:bcl-2-Associated X Protein; D051025:bcl-Associated Death Protein; D013853:Thioacetamide; C022616:thiazolyl blue