| Summary: | 碩士 === 中國醫藥大學 === 中國醫學研究所碩士班 === 95 === Chrysophanol, an anthraquinone-derivative alkaloid, is extracted from Rheum palmatum L. It has been reported that chrysophanol may exhibit several regulatory functions including anti-inflammation, antiseptic, antioxidation, anti-aging, prevention of atherosclerosis, anti-dementia, and anti-tumor activity. However, the molecular mechanism of anti-tumor effects of chrysophanol has not been well defined. In this study, we used human hepatic carcinoma Hep3B cells and lung carcinoma A549 cells as two cell models to examine the inhibitory effects of chrysophanol on tumor cell growth and migration. Cell viability assay revealed that chrysophanol indeed prevented cell proliferation in these two cell lines. To further investigate the cellular mechanism of chrysophanol, we measured the changes of cellular physiological parameters in chrysophanol-treated cells. We found that the treatment of chrysophanol induced ROS production and increased Ca2+ concentration, results in a significant change of mitochondrial membrane potential and the reduction of ATP concentration in mitochondria. There is no significant change in the expression of Apaf-1 (Apoptosis protease activating factor-1), caspase-8 and caspas-9. Moreover, the chrysophanol-induced cell death in Hep3B cells and A549 cells was accompanied with several characteristics of necrosis, including the necrotic morphology, DNA fragmentation as well as the enhancement of DAPI positive cells. In addition, flow -cytometry analysis showed that the S phase of cell cycle was blocked in chrysophanol-treated cells. Western blotting results also revealed that the expression levels of cell cycle regulatory proteins, such as p21, p27 and p53, were significantly changed in a time-dependent manner. The protein expression of p21 was increased while CDK2 and thymidylate synthase were decreased. We hypothesized that, with the chrysophanol treatment, CDK2 and thymidylate synthase were phosphorylated in response to DNA damage and could not activate cyclin A/CDK2 complex to facilitate the cell cycle progression. In terms of anti-metastasis effects of chrysophanol, our results showed that in chrysophanol-treated cells, the cell migration and invasion were lower than the controls. The expression of PI3K, ERK, and MKK, which are involved in metastasis of cancer cells, was also down-regulated under chrysophanol treatments. Our studies provided the evidence for the inhibitory effects of chrysophanol in cell proliferation and metastasis, and suggested the application of chrysophanol as a potential anticancer reagent in the future.
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