Characterization of the growth-inhibitory and apoptosis-inducing activities of a triterpene saponin, securioside B against BAC1.2F5 macrophages

Abstract

BACKGROUND: Since the growth state of macrophages in local pathological sites is considered a factor that regulates the processes of many disease, such as tumors, inflammation, and atherosclerosis, the substances that regulate macrophage growth or survival may be useful for disease control. We previously reported that securiosides A and B, novel triterpene saponins, exerted macrophage-oriented cytotoxicity in the presence of a L-cell-conditioned medium containing macrophage colony-stimulating factor (M-CSF), while the compounds did not exhibit an effect on macrophages in the absence of the growth-stimulating factors. Aim: This study was undertaken to characterize the growth-inhibitory and the apoptosis-inducing activities of securioside B, focusing on the effects of the macrophage-growth factor(s), and to examine the implication of a mitochondria pathway in apoptosis induction. Methods: The effect of securioside B on a murine macrophage cell line (BAC1.2F5) was examined by MTT assay and lactose dehydrogenase release assay in the presence of L-cell-conditioned medium, M-CSF, or granulocyte-macrophage CSF (GM-CSF). Results: Securioside B inhibited the growth of the cells stimulated by recombinant M-CSF or GM-CSF, but it scarcely induced cytolysis of the cells under the same conditions. Moreover, securioside B did not induce cell death when the compound only was added to the cells. On the other hand, the compound extensively induced apoptotic cell death in the presence of L-cell-conditioned medium, suggesting that apoptosis induction by securioside B requires the additional factor(s) present in L-cell-conditioned medium. Securioside B plus L-cell-conditioned medium induced the activation of caspase-3 and caspase-9, but not caspase-8. In addition, cytochrome c release from the mitochondria into the cytosol, and disrupted mitochondria membrane potential, was also observed in the apoptotic BAC1.2F5 cells. Conclusion: These data suggest that securioside B has growth-inhibitory activity against growth factor-stimulated macrophages, and that it induces apoptotic macrophage death through the activation of a mitochondrial pathway in the presence of L-cell-conditioned medium.