| Paeoniae Radix (PR) is the root of traditional Chinese Herb, Paeonia lactiflora Pallas, which is commonly used to treat liver diseases in China for centuries. Several earlier studies indicated that PR has anticancer growth activities. In this project, the anticancer effects of the PR extracts on human hepatocellular carcinoma HepG2 cells and multidrug-resistance human hepatocellular carcinoma R-HepG2 cells and the underlying mechanisms were studied.;In our study, we found that the aqueous extracts of PR (PRA) exhibited inhibitory effects on the growth of HepG2 and R-HepG2 cells. The IC50 were about 4.6mg/ml and 5.0mg/ml respectively after 48 hours incubation. The induction of DNA fragmentation in both cell lines and accumulation of sub-G1 phase of cell cycle profile in PRA-treated HepG2 cells showed that the cytotoxicity of PRA on HepG2 and R-HepG2 cells was through apoptosis. In vivo studies also showed that PRA was effective in the inhibition of tumor growth in HepG2-bearing nude mice without any cardio- and hepato-toxicity. However, it could not exert any mitogenic effect on isolated lymphocytes in inbred BALB/c mice.;An active component was isolated from the organic extract of Paeoniae Radix (PRO) by assay-guided purification. By Liquid Spray Ionization Mass Spectrometry (LSIMS) and High Resolution Mass Spectrometry (HRMS), the molecular formula of this active component was identified to be C7H6O5. Judging from its 1H NMR spectrum and 13C NMR spectrum, the structure of this active component was then identified to be gallic acid (GA). GA was found to be capable of inducing apoptotic cell death of HepG2 and R-HepG2 cells. The IC50 were 117muM and 205muM after 48 hours incubation respectively. The treatment of GA resulted in mitochondrial membrane depolarization, DNA fragmentation and phosphatidylserine externalisation in both cell lines.;By Real Time Polymerase Chain Reaction (RT-PCR), mRNA of p53, procaspase 9 and procaspase 3 were found to be up-regulated in both HepG2 and R-HepG2 cells upon GA treatments. Moreover, Western blot analysis showed that p53 protein was induced to express, apoptogenic protein cytochrome c was released from mitochondria, activation of caspase 9, caspase 3 and poly (ADP-ribose) polymerase (PARP) in GA treated HepG2 cells.;In conclusion, GA could trigger apoptotic cell death in both HepG2 and R-HepG2 cells which was p53-dependent and involved mitochondria dysfunction. |
