The Impact And Mechanisms Of Silibinin On Human Monocyte Cell Line U937

Silibinin a flavonoligan, is a major constituent of silimarin, and has been recently shown to have pleiotropic anticancer capabilities. In this dissertation, we examined the effects of silibinin in human leukemia U937 cell line, apoptosis and autophagy, investigated the molecular mechanisms of this compound in vitro.The results demonstrate that the proliferation of U937 cells was inhibited by silibinin in a dose- and time-dependent manner. Photomicroscopical observation and lactate dehydrogenase assay showed that silibinin-induced U937 cell death characterized as apoptosis. Monodansylcadaverine (MDC), a fluorescent compound, has been shown to stain autophagosomes. By fluorescent density analysis, silibinin increased MDC positive U937 cells, and the expression of autophagy-related protein, Beclin 1 was increased, indicated that autophagy was augmented.Upon the stimulus of silibinin, the expression of pro-caspase 8, pro-caspase 3 were down-regulated and IC AD was digested. Co-treatment of U937 cells with the caspase 8 inhibitor (z- IETD-fmk), was capable of restoring cell proliferation significantly in silibinin-treated cells. FAK and Src was protein tyrosine kinase (PTK), downstream of integrin. The expression of FAK, Grb2 (a protein downstream of FAK), as well as ERK-MAPK and p38-MAPK were decreased by silibinin. In the presence of PTK inhibitor genistein, silibinin induced U937 cells death more apparently, suggesting that the integrin signal pathway may be blocked by silibinin.Interestingly, we found that the general raise of cytochrome c during apoptosis was not observed, whereas antiapoptotie protein Bcl-2 was increased and paxillin, the molecular downstream of integrin was activated.Pretreatment of U937 cells with the PI3K inhibitor (wortmannin) caused an increase in the level of the autophagy induced by silibinin, and simultaneously the proliferation arrested by silibinin was aggravated. Therefore, these results suggest that PI3K played an important role in regulation of apoptosis and autophagy.