title: Sodium butyrate induces apoptosis in human hepatoma cells by a mitochondria/caspase pathway, associated with degradation of beta-catenin, pRb and Bcl-XL.
abstract: Butyrate can promote programmed cell death in a number of tumour cells in vitro. This paper provides evidence that butyrate induces apoptosis in human hepatoma HuH-6 and HepG2 cells but is ineffective in Chang liver cells, an immortalised non-tumour cell line. In both HuH-6 and HepG2 cells, apoptosis appeared after a lag period of approximately 16 h and increased rapidly during the second day of treatment. In particular, the effect was stronger in HuH-6 cells, which were, therefore, chosen for ascertaining the mechanism of butyrate action. In HuH-6 cells, beta-catenin seemed to exert an important protective role against apoptosis, since pretreatment with beta-catenin antisense ODN reduced the content of beta-catenin and anticipated the onset of apoptosis at 8 h of exposure to butyrate. Moreover, in HuH-6 cells, butyrate induced loss of mitochondrial membrane potential, release of cytochrome c from mitochondria, activation of caspase 9 and caspase 3, and degradation of poly(ADP-ribose) polymerase. In addition, during the second day of treatment, beta-catenin, pRb, and cyclins D and E were diminished and the phosphorylated form of pRb disappeared. Also, the content of the anti-apoptotic factor Bcl-XL fell markedly during this period, while that of the pro-apoptotic factor Bcl-Xs increased. These effects were accompanied by an increase in both Bcl-XL and Bcl-Xs mRNA transcripts, as ascertained by reverse transcriptase-polymerase chain reaction. Our results suggest that caspases have a crucial role in butyrate-induced apoptosis. This conclusion is supported by the observation that the inhibitors of caspases, benzyloxy carbonyl-Val-Ala-Asp-fluoromethylketone and benzyloxy carbonyl-Asp-Glu-Val-Asp-fluoromethylketone, prevented apoptosis and the decrease in Bcl-XL, pRb, cyclins and beta-catenin. These effects were most probably responsible for the increased sensitivity of the cells to butyrate-induced apoptosis, which was observed on the second day of treatment.
authors.affiliation: Dipartimento di Biologia Cellulare e dello Sviluppo, Sezione di Biochimica, Università di Palermo, Policlinico, Via del Vespro 129, 90127 Palermo, Italy.
authors.forename: S
authors.initials: S
authors.lastname: Emanuele
authors.forename: A
authors.initials: A
authors.lastname: D'Anneo
authors.forename: G
authors.initials: G
authors.lastname: Bellavia
authors.forename: B
authors.initials: B
authors.lastname: Vassallo
authors.forename: M
authors.initials: M
authors.lastname: Lauricella
authors.forename: A
authors.initials: A
authors.lastname: De Blasio
authors.forename: R
authors.initials: R
authors.lastname: Vento
authors.forename: G
authors.initials: G
authors.lastname: Tesoriere
mesh_terms: D017209:Apoptosis / Q000187:drug effects; D015153:Blotting, Western / Q000379:methods; D002087:Butyrates / Q000494:pharmacology*; D006528:Carcinoma, Hepatocellular / Q000473:pathology*; D020169:Caspases / Q000378:metabolism; D002460:Cell Line / Q000187:drug effects; D056741:Cyclin D; D019927:Cyclin E / Q000378:metabolism; D016213:Cyclins / Q000378:metabolism; D003598:Cytoskeletal Proteins / Q000378:metabolism; D006801:Humans; D008113:Liver Neoplasms / Q000473:pathology*; D008564:Membrane Potentials / Q000187:drug effects; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015534:Trans-Activators / Q000378:metabolism; D051020:bcl-X Protein; D051176:beta Catenin
chemical_list: C494811:BCL2L1 protein, human; D002087:Butyrates; C495270:CTNNB1 protein, human; D056741:Cyclin D; D019927:Cyclin E; D016213:Cyclins; D003598:Cytoskeletal Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D015534:Trans-Activators; D051020:bcl-X Protein; D051176:beta Catenin; D011065:Poly(ADP-ribose) Polymerases; D020169:Caspases