Luteolin By Inhibiting The Activation Of Akt And Promote Of Xiap Degradation Sensitization Fas-induced Hepg2 Cells Apoptosis

Fas, an important cell surface protein of the TNF receptor superfamily that induces apoptosis through binding Fas ligand or anti-Fas antibodies. Unfortunately, not all Fas-expressing cells are sensitive to its stimulus. Systemic treatment with anti-Fas antibodies or rFasL causes severe damage to the liver, so most preclinical studies are now focusing on circumvention of this problem by local administration of FasL, or on the use of inducible FasL-expressing vectors as gene therapy. Therefore it is important to research the mechanisms that counteract the FasL-induced apoptotic process which are still poorly understood.Luteolin, an important flavonoid present in a variety of edible plants, exhibits a wide spectrum of pharmacologic properties such as anticancer, antioxidant, and anti-inflammatory. Luteolin has a variety of anti-cancer effects, cell growth and kinase activity inhibition, apoptosis induction. Furthermore, much more attention has been turned to the chemosensitizing effect of luteolin on cancer cells death.Akt is a serine/threonine protein kinase. Full activation of Akt requires recruitment to the plasma membrane and phosphorylation at Thr308 and Ser473. Akt and its downstream targets promote cell survival and suppress apoptotic death in a major cell types. Akt phosphorylates X-linked inhibitor of apoptosis protein (XIAP) at residue serine87 and inhibits the ubiquitination and degradation.In this study, we found that luteolin synergistically caused the Fas/FasL-induced apoptosis in HepG2 cells. Such potentiation was achieved through inhibiting Akt activation and promoting X-linked Inhibitor of Apoptosis Protein (XIAP) proteasomal degradation, that mediated the survival signals and allow the cells to escape from apoptosis in various human cancers. We showed that low dose luteolin decreased the phosphorylation of Akt at specific site Ser478, whereas the level of total Akt was relatively unchanged. We also found that XIAP, a physiological substrate of Akt, significantly decreased in HepG2 cells treated with luteolin and FasL. To confirm the role of Akt and XIAP in this sensitization, we further tested the effect of LY294002 (a inhibitor of Akt) and transfecting the wild-type Akt plasmid in cells treated with luteolin and FasL. The results showed that luteolin inhibited Akt phosphorylation on Ser473, without altering total Akt levels, which is similar to the effect of LY294002. This effect was abolished by transfecting the wild-type Akt plasmid. In nontransfected cells, almost all cells underwent apoptosis after luteolin and FasL treatment in time-dependent way. In contrast, most Akt-overexpressing cells remained alive. These data indicate that XIAP as a substrate of Akt involved in the sensitization of luteolin on FasL-mediated apoptosis.