cytochrome-c-t and rhein

Rhein, a compound found as a glucoside in the root of rhubarb, is currently a subject of interest for its antitumor properties. The apoptosis of tumor cell lines induced by rhein was observed, and the involvement of mitochondria was established; however, the role of mitochondrial permeability transition (MPT) remains unknown. Here we report that MPT plays an important role in the apoptosis of human hepatocellular carcinoma Hep-G2 cells induced by rhein. After adding rhein to the isolated hepatic mitochondria, swelling effects and the leakage of Ca(2+) were observed. These alterations were suppressed by cyclosporin A (CsA), an MPT inhibitor. Furthermore, in Hep-G2 cells, the decrease of ATP production, the loss of mitochondrial transmembrane potential (MTP), the release of cytochrome c (Cyto c), and the activation of caspase 3 were also observed. These toxic effects of rhein can also be attenuated by CsA as well. Moreover, TUNEL assay confirmed that in the presence of CsA, rhein-induced apoptosis was largely inhibited. These results suggest that MPT plays a critical role in the pathogenesis of Hep-G2 cell injury induced by rhein, and imply that MPT may contribute to the anti-cancer activity of rhein.

Topics: Adenosine Triphosphate; Anthraquinones; Antineoplastic Agents, Phytogenic; Apoptosis; Carcinoma, Hepatocellular; Caspase 3; Cyclosporine; Cytochromes c; Hep G2 Cells; Humans; In Situ Nick-End Labeling; Liver Neoplasms; Membrane Potential, Mitochondrial; Mitochondria; Permeability; Phytotherapy; Plant Extracts; Rheum

Vascular smooth-muscle cell proliferation plays an important role in atherosclerosis and restenosis. Rhein is an active component extracted from rhubarb. In this study, rhein was found to exert potent inhibitory effects against tumor necrosis factor (TNF)-alpha-induced human aortic smooth-muscle cells (HASMCs) proliferation.. These effects were associated with induced apoptosis, including the induction of Annexin V-positive cells, the cleavage of poly(ADP-ribose)polymerase (PARP), and caspases 3, 8 and 9.. Inhibitors of caspases 3, 8 and 9 were efficiently blocked by rhein-induced apoptosis in TNF-alpha-treated HASMCs. In addition, treatment with rhein resulted in the release of cytochrome c into the cytosol, a loss of mitochondrial membrane potential (DeltaPsi(m)), a decrease in Bcl-2 and Bcl-xL and an increase in Bax and Bak expression. However, rhein-mediated apoptosis was blocked by a mitochondrial membrane depolarization inhibitor. These findings indicate that rhein-induced apoptosis occurred via a mitochondrial pathway. Furthermore, the inhibition of mitochondrial membrane depolarization was efficiently blocked by rhein-induced caspase-9 activity, which indicates that the rhein-induced caspase activation signal was downstream of the mitochondrial pathway. Taken together, the results of this study show that rhein inhibits TNF-alpha-induced HASMC proliferation via mitochondria-dependent apoptosis and that rhein has the potential to act as an anti-atherosclerosis agent.

Topics: Anthraquinones; Aorta; Apoptosis; Apoptosis Regulatory Proteins; Bongkrekic Acid; Cardiovascular Agents; Caspase Inhibitors; Caspases; Cell Proliferation; Cells, Cultured; Cysteine Proteinase Inhibitors; Cytochromes c; Dose-Response Relationship, Drug; Humans; Membrane Potential, Mitochondrial; Mitochondria, Muscle; Muscle, Smooth, Vascular; Myocytes, Smooth Muscle; Recombinant Proteins; Tumor Necrosis Factor-alpha

Rhein is an anthraquinone compound enriched in the rhizome of rhubarb, a traditional Chinese medicine herb showing anti-tumor promotion function. In this study, we first reported that rhein could induce apoptosis in human promyelocytic leukemia cells (HL-60), characterized by caspase activation, poly(ADP)ribose polymerase (PARP) cleavage, and DNA fragmentation. The efficacious induction of apoptosis was observed at 100 microM for 6h. Mechanistic analysis demonstrated that rhein induced the loss of mitochondrial membrane potential (DeltaPsi(m)), cytochrome c release from mitochondrion to cytosol, and cleavage of Bid protein. Rhein also induced generation of reactive oxygen species (ROS) and the phosphorylation of c-Jun N-terminal kinase (JNK) and p38 kinase. However, these actions seem not to be associated with the apoptosis induction because antioxidants including N-acetyl cysteine (NAC), Tiron, and catalase did not block rhein-induced apoptosis, although they could block the generation of ROS and the phosphorylation of JNK and p38 kinase. Our data demonstrate that rhein induces apoptosis in HL-60 cells via a ROS-independent mitochondrial death pathway.

Topics: Anthraquinones; Apoptosis; BH3 Interacting Domain Death Agonist Protein; Carrier Proteins; Caspase 3; Caspases; Cytochromes c; DNA Fragmentation; Dose-Response Relationship, Drug; Enzyme Activation; HL-60 Cells; Humans; JNK Mitogen-Activated Protein Kinases; MAP Kinase Kinase 4; Membrane Potentials; Mitochondria; Mitogen-Activated Protein Kinase Kinases; Poly(ADP-ribose) Polymerases; Reactive Oxygen Species