cytochrome-c-t and mahanine

1 Mahanine, a naturally occurring carbazole alkaloid in some Asian vegetables, has been shown to exhibit antimutagenicity, antimicrobial activity, cytotoxicity, and other biological properties. 2 In the present study, we investigated the effect of mahanine on the activation of the apoptotic pathway in human leukemia U937 cells. Various end points were used to screen for apoptosis: Morphological changes in cells, the relative numbers of viable and apoptotic cells; translocation of membrane bound phosphatidylserine and DNA analysis. 3 We found that mahanine-induced apoptosis in U937 cells involved activation of caspases, including caspase-3, release of cytochrome c into cytosol, loss of mitochondrial membrane permeability, and decreased levels of cellular ATP. 4 Pretreatment of cells with cyclosporine A, prior to/concomitant with exposure to mahanine, effectively prevented the deleterious effects of the alkaloid on cellular integrity and viability. 5 As mitochondrial permeability is known to be important in the regulation of cytochrome c release, our observations indicate that mitochondria are the principal target of mahanine. More specifically, we propose that mahanine causes the mitochondrial membranes to lose their permeability, resulting in caspase-3 activation and apoptosis.

Topics: Alkaloids; Antineoplastic Agents, Phytogenic; Apoptosis; Carbazoles; Caspase 3; Caspases; Cell Proliferation; Cell Survival; Cyclosporine; Cytochromes c; DNA Fragmentation; Enzyme Activation; Humans; Immunosuppressive Agents; Ion Channels; Mitochondria; Mitochondrial Membrane Transport Proteins; Mitochondrial Permeability Transition Pore; Nucleosomes; Protein Transport; Rutaceae; U937 Cells

Mahanine, a carbazole alkaloid occurs in the edible part of Micromelum minutum, Murraya koenigii and related species has been found to induce apoptosis in human myeloid cancer cell (HL-60). Concentration of 10 microM mahanine caused a complete inhibition of cell proliferation and the induction of apoptosis in a time dependent manner. Mahanine-induced cell death was characterized with the changes in nuclear morphology, DNA fragmentation, activation of caspase like activities, poly(ADP-ribose) polymerase cleavage, release of cytochrome c into cytosol and stimulation of reactive oxygen species generation. The cell death was completely prevented by a pancaspase inhibitor benzyloxycarbonyl-L-aspart-1-yl-[(2,6-dichlorobenzoyl)oxy]methane (Z-Asp-CH(2)-DCB). Mahanine activated various caspases such as caspase-3, -6, -8 and -9 (like) activities but not caspase-1 like activity. More than 70% cell survival was observed in the presence of a caspase-3 inhibitor. In addition, co-treatment of cyclosporin A markedly increased the survival of mahanine-treated HL-60 cells. Flow cytometric analysis revealed that mahanine decreased the mitochondrial membrane potential of intact cells, and disrupted cell cycle progression by increasing the number of cells in sub-diploid region, concomitantly with the decrease of cells in diploid phases, particularly at late hours of apoptosis. The overall results suggest that mahanine down regulates cell survival factors by activation of caspase-3 through mitochondrial dependent pathway, and disrupts cell cycle progression.

Topics: Antineoplastic Agents; Apoptosis; Aspartic Acid; Carbazoles; Caspase 3; Caspase Inhibitors; Caspases; Cell Cycle; Cell Division; Cysteine Proteinase Inhibitors; Cytochromes c; Cytosol; DNA; DNA Fragmentation; HL-60 Cells; Humans; Leukemia; Mitochondria; Oligopeptides; Poly (ADP-Ribose) Polymerase-1; Poly(ADP-ribose) Polymerases; Protease Inhibitors; Proteins; Reactive Oxygen Species