Anticancer Activities And Mechanisms Study Of Telekin

With the development of human living environment and living habits changing, the malignant tumor has become one of the factors, which are seriously harmful to human health. Hepatocellular carcinoma (HCC) is one of the most frequent primary tumors in adults with a high mortality rate worldwide. Chemotherapy is currently the most common and effective strategy against HCC in the clinical field. Natural products are the major players in pharmacology in general and in cancer therapy in particular. Telekin, a eudesmane-type sesquiterpene lactone compound solated from Carpesium divaricatum, which has long been used as Chinese folk medicine given its antipyretic, analgesic, vermifugic, and anti-inflammatory properties. Telekin has also been reported to exhibit strong antitumor activity in several human carcinoma cell lines.We performed MTT colorimetric assays to test the cytotoxicity of several compounds isolated from C. divaricatum on several cancer lines and normal cells. Results showed that, telekin displayed the strongest potency required to inhibit tumor cells growth in Smmc-7721, HepG-2, LX-2, A549, and U87cells. Compared with tumor cell lines, telekin had a lower cytotoxicity on normal cells HL-7702. Human hepatocarcinoma cell lines were employed to further evaluate anti-tumor effect and explore anti-cancer mechanisms of telekin. First, telekin displayed significant cytotoxicity in HCCs in a dose-and time-dependent manner. Telekin-induced apoptosis was characterized by chromatin condensation, formation of apoptotic bodies, and exposure of phosphatidylserine on the extracellular surface, as revealed by DAPI nuclear staining and flow cytometry.Flow cytometry analysis showed that telekin induced the loss of mitochondrial membrane potential (MMP), as well as increased the levels of intracellular free calcium and reactive oxygen species (ROS). Additionally, Western Blot results demonstrated that telekin induced the decrease in Apaf-1and Bcl-2expression, increase in Bax expression, release of cytochrome C, and activation of caspase-9and caspase-3in HepG-2cells. These findings indicate that telekin activates the mitochondria-mediated apoptotic pathway in hepatocellular carcinoma cells and may merit further investigation as a potential therapeutic agent for the treatment of hepatocellular carcinoma.Later, we found that telekin inhibits cancer cell proliferation through arresting cell cycle progression before inducing apoptosis in HepG-2cells. Treating HepG-2cells with telekin arrested cells in G2/M phase. This effect of telekin is dose-and time-dependent. Furthermore, our results also showed that the phosphorylation level of p38MAPK increased after stimulation with ROS generation induced by telekin. Previous works suggested that p38MAPK participates not only in inflammatory responses but also in stress-induced signaling, cell proliferation, cell cycle arrest, and apoptosis. It had been reported that p38kinase plays a critical role in the initiation of a G2/M phase delay via controlling the level of Cdc25protein. Activation of p38MAPK could arise G2/M phase cell cycle arrest and suppress cell proliferation.Flow cytometic analysis showed that treating HepG-2cells with telekin arrested cells in G2/M phase. This effect of telekin is dose-and time-dependent. It showed that significant accumulations of G2/M population were observed in HepG-2cells after telekin treatment0μmol/L (8.37%) and15umol/L (50.88%) for24h. In this connection, Western Blot results demonstrated that telekin significantly increased the protein level of phospho-p38, phospho-MAPKAPK-2, phospho-Cdc25A, and phospho-Cdc2, but decreased that of Cdc25A, Cdc2, and Cyclin B1. This effect is dose-dependent. Simultaneously, inhibition of the p38MAPK signaling pathway by SB203580(5μmol/L) significantly prevented G2/M phase cell cycle arrest, and increased the percentage of cell viability. When cells were pretreated with SB203580before exposure of cells to telekin (15μmol/L), the percentage of cell viability (48h) increased from31.67%to61.8%, and the percentage of cells in G2/M phase (24h) decreased from50.88%to17.17%for5μmol/L SB203580. These findings indicated that telekin activated the p38MAPK signaling pathway to induce G2/M phase cell cycle arrest in HepG-2cells.In conclusion, our study demonstrated that telekin-induced apoptosis may including two molecules and mechanisms. Our works indicated that telekin activated the mitochondria-mediated apoptotic pathway, meanwhile operated the p38MAPK pathway induced cell cycle arrest at G2/M-phase. Our findings provide a mechanism for the chemopreventive properties of telekin in Hepatocellular carcinoma cells.