Activation of p53 contributes to pseudolaric acid B-induced senescence in human lung cancer cells in vitro.

Abstract

Pseudolaric acid B (PAB), a diterpene acid isolated from the root bark of Pseudolarix kaempferi Gordon, has shown to exert anti-tumor effects via inducing cell cycle arrest followed by apoptosis in several cancer cell lines. Here we reported that PAB induced a mitotic catastrophe in human lung cancer A549 cells, which resulted in senescence without apoptosis or necrosis. Three human lung cancer cell lines (A549, H460 and H1299 cells) were examined. Cell growth inhibition was assessed with MTT assay. Cell cycle distribution was determined using a flow cytometer. Cell nuclear morphology was observed under a fluorescence microscope. Senescent cells were detected using SA-β-Gal staining. Apoptotic and senescent protein expression was examined using Western blot analysis. The expression of p53 and p21 in the cells was downregulated by siRNAs. Treatment with PAB (5-80 μmol/L) inhibited the growth of A549 cells in dose- and time-dependent manners. Prolonged treatment with PAB (20 μmol/L) caused G2/M arrest at day 1 followed by mitotic catastrophe from day 2, which eventually resulted in cell senescence between days 3 and 4 without cell death (apoptosis or necrosis). Knockdown of p53 expression with siRNA significantly suppressed PAB-induced senescence in A549 cells (p53 wild). Furthermore, PAB-induced senescence was also observed in human lung cancer H460 cells (p53 wild), but not in human lung cancer H1299 cells (p53 null). The anti-tumor action of PAB against human lung cancer A549 cells in vitro involves the induction of senescence through activation of the p53 pathway.