title: Cisplatin-induced apoptosis in Hep3B cells: mitochondria-dependent and -independent pathways.
abstract: Human hepatoma cell lines undergo apoptosis after treatment with cisplatin (CP), by mechanisms that are not fully understood, although our previous study demonstrated that Fas-dependent or -independent pathways are involved. To elucidate the mechanisms of CP-induced apoptosis in Hep3B cells, which are Fas- and p53-negative, we investigated mitochondria associated pathways, the involvement of NF-kappaB, and p73 activation. Results of Western blot and flow cytometry assay revealed that the translocation of Bax, resulted in the loss of mitochondrial membrane potential (Deltaphi(m)) and the efflux of cytochrome c and of second mitochondria-derived activator of caspase/DIABLO from mitochondria into the cytosol. Caspase-3, -8 and -9 were activated by CP treatment, however, CP-induced apoptosis was not completely blocked by pretreating with the pan-caspase inhibitor, benzyloxycarbonyl-valinyl-alaninyl-aspartyl-(O-methyl)-fluoromethylketone, indicating that caspase-independent apoptotic pathways might also be involved. RNase protection assay confirmed that NF-kappaB downregulation leading to the suppression of its target genes, such as XIAP and TRAF2, and p73 accumulation were also observed in Hep3B cells treated with CP. CP-induced apoptosis was inhibited to some extent by transiently overexpressed p73 dominant negative and XIAP, but not by p73DN or XIAP alone. In conclusion, this study demonstrates that CP-induced apoptosis in Hep3B cells is associated with mitochondrial dysregulation, NF-kappaB downregulation and p73 accumulation.
authors.affiliation: Department of Microbiology and Brain Korea 21 Project for Medical Sciences, Institute for Immunology and Immunological Diseases, Seoul, South Korea. jhpark5277@yumc.yonsei.ac.kr
authors.forename: Ji Su
authors.initials: JS
authors.lastname: Kim
authors.forename: Jae Myun
authors.initials: JM
authors.lastname: Lee
authors.forename: Yong-Joon
authors.initials: YJ
authors.lastname: Chwae
authors.forename: Yeon Hyang
authors.initials: YH
authors.lastname: Kim
authors.forename: Jung Hwan
authors.initials: JH
authors.lastname: Lee
authors.forename: Kunhong
authors.initials: K
authors.lastname: Kim
authors.forename: Tae Ho
authors.initials: TH
authors.lastname: Lee
authors.forename: Se Jong
authors.initials: SJ
authors.lastname: Kim
authors.forename: Jeon Han
authors.initials: JH
authors.lastname: Park
mesh_terms: D000970:Antineoplastic Agents / Q000494:pharmacology*; D017209:Apoptosis; D051017:Apoptosis Regulatory Proteins; D001692:Biological Transport; D002352:Carrier Proteins / Q000378:metabolism; D020169:Caspases / Q000378:metabolism; D002945:Cisplatin / Q000494:pharmacology*; D045304:Cytochromes c / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism; D015536:Down-Regulation; D016147:Genes, Tumor Suppressor; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D008928:Mitochondria / Q000187:drug effects* / Q000378:metabolism; D024101:Mitochondrial Proteins / Q000378:metabolism; D016328:NF-kappa B / Q000378:metabolism; D009687:Nuclear Proteins / Q000378:metabolism; D011506:Proteins / Q000378:metabolism; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D019253:Proto-Oncogene Proteins c-bcl-2; D014407:Tumor Cells, Cultured; D000072160:Tumor Protein p73; D025521:Tumor Suppressor Proteins; D051636:X-Linked Inhibitor of Apoptosis Protein; D051028:bcl-2-Associated X Protein
chemical_list: D000970:Antineoplastic Agents; D051017:Apoptosis Regulatory Proteins; C494826:BAX protein, human; D002352:Carrier Proteins; C412403:DIABLO protein, human; D004268:DNA-Binding Proteins; D047908:Intracellular Signaling Peptides and Proteins; D024101:Mitochondrial Proteins; D016328:NF-kappa B; D009687:Nuclear Proteins; D011506:Proteins; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; C000606460:TP73 protein, human; D000072160:Tumor Protein p73; D025521:Tumor Suppressor Proteins; D051636:X-Linked Inhibitor of Apoptosis Protein; C496866:XIAP protein, human; D051028:bcl-2-Associated X Protein; D045304:Cytochromes c; D020169:Caspases; D002945:Cisplatin