| Pathway description: |
p16-pRb-E2F1:After administration of SAL an upregulation of p16, hypophosphorylation of pRb and down-regulation of the pRb targets Cyclin D1 as well as of E2F1 protein levels were observed indicating that the p16-pRb pathway is regulated by SAL treatment .Similar results were obtained by treating the cells for 6 days. In contrast, LAL treatment mediated no detectable changes of p16,Cyclin D1 and E2F1 expression level. In line with this,SAL treatment led to inhibition of down-stream targets of pRB, Cyclin D1 as well as E2F1 at mRNA level, whereas the p16 mRNA is upregulated by SAL doses. Accordingly, the mRNA level of ID1, an inhibitor of p16 expression, is reduced upon SAL administration . Thus, these data indicate that the p16-pRb-E2F1 pathway is associated with the androgen-mediated cellular senescence.Src-Akt-mTOR :Notably, treatment of LNCaP cells with the Src inhibitor PP2 under SAL conditions reduces the androgen-mediated cellular senescence. In contrast, inhibition of Src without androgens or with LAL has no detectable influence on the SA β-Gal activity in LNCaP cells compared to control. Akt phosphorylation is a well-known pathway of the Src tyrosine kinase and involves signaling molecules such as the PI3K as well as the mammalian target of rapamycin (mTOR). Using inhibitors the role of these factors in androgen-mediated cellular senescence was analyzed. Inhibition of PI3K, an Akt-activating kinase,by the 3-MA inhibitor, reduced the level of androgen-induced cellular senescent cells, which confirms that the Src-Akt signaling pathway is involved in androgen-induced cellular senescence at supraphysiological levels. Similarly, using a specific Akt-kinase inhibitor(Akti) reveals a strong reduction of the SAL-mediated SAβ-Gal activity. A further downstream target of the Src- and the Akt-kinase is mTOR, which is involved in proliferation and cell cycle regulation processes [35]. Rapamycin alone mediates no detectable change in the level of cellular senescence. In contrast, rapamycin co-treated with SAL resulted in reduction of SA β-Gal positive stained cells.The data show that rapamycin reduces the androgen-mediated SA β-Gal activity and suggest that mTOR is partially involved in androgen-mediated cellular senescence. This supports the notion that the Src-Akt-mTOR signaling mediates the androgen-mediated induction of cellular senescence. |